Journal articles: 'Actin depolymerizing factor (ADF)'

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Tokuraku, Kiyotaka, Shinya Okamoto, Miho Katsuki, Hiroyuki Nakagawa, and Susumu Kotani. "The actin-depolymerizing factor destrin has an actin-stabilizing domain." Biochemistry and Cell Biology 79, no. 6 (December 1, 2001): 773–78. http://dx.doi.org/10.1139/o01-176.

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Destrin is a 19 kDa actin-depolymerizing protein of the ADFcofilin family. Destrin was digested with trypsin to a structurally stable 9.2 kDa fragment that contains the actin-binding sequence. The purified 9.2 kDa fragment has an actin filament stabilizing activity, rather than an actin filament depolymerizing activity. The deleted region is probably essential for the actin filament depolymerizing activity of intact destrin. Surprisingly, the 9.2 kDa fragment also has an assembly-promoting activity in the absence of ATP.Key words: actin, destrin, cofilin, ADF.

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Bamburg, J. R., and D. Bray. "Distribution and cellular localization of actin depolymerizing factor." Journal of Cell Biology 105, no. 6 (December 1, 1987): 2817–25. http://dx.doi.org/10.1083/jcb.105.6.2817.

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Actin depolymerizing factor (ADF) is a low molecular mass (19 kD) protein that forms a tightly bound dimeric complex with actin. We have raised a rabbit antiserum to chick brain ADF and used it to analyze the distribution and cellular localization of ADF. We find that ADF is a major constituent of all chick embryonic and most adult tissues examined, accounting for 0.1-0.4% of the total protein. Some tissues have as much as 0.6 mol ADF per mole actin. Adult heart and skeletal muscle are unusual in having very low levels of ADF: less than 0.02% of the soluble protein. During the development of skeletal muscle, ADF levels are maximal up to approximately 11 d in ovo and then decline to reach their adult levels by 14 d posthatching. Brain tissue and cultured cell lines from several other vertebrates, including mammals, all possess proteins of identical size to ADF that are recognized by the ADF antiserum. No proteins are specifically recognized by the ADF antiserum in extracts from Acanthamoeba castellanii or from nerve tissue of several invertebrates. Indirect immunofluorescence shows that ADF is present throughout the cytosol of most cells and at the leading edge of ruffled membranes and in the neuronal growth cone. Its abundance and widespread distribution together with its ability to sequester actin molecules, even those in an already polymerized state, suggest that ADF is a major factor in the regulation of actin filaments in many vertebrate cells.

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Paavilainen, Ville O., Esko Oksanen, Adrian Goldman, and Pekka Lappalainen. "Structure of the actin-depolymerizing factor homology domain in complex with actin." Journal of Cell Biology 182, no. 1 (July 14, 2008): 51–59. http://dx.doi.org/10.1083/jcb.200803100.

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Actin dynamics provide the driving force for many cellular processes including motility and endocytosis. Among the central cytoskeletal regulators are actin-depolymerizing factor (ADF)/cofilin, which depolymerizes actin filaments, and twinfilin, which sequesters actin monomers and caps filament barbed ends. Both interact with actin through an ADF homology (ADF-H) domain, which is also found in several other actin-binding proteins. However, in the absence of an atomic structure for the ADF-H domain in complex with actin, the mechanism by which these proteins interact with actin has remained unknown. Here, we present the crystal structure of twinfilin's C-terminal ADF-H domain in complex with an actin monomer. This domain binds between actin subdomains 1 and 3 through an interface that is conserved among ADF-H domain proteins. Based on this structure, we suggest a mechanism by which ADF/cofilin and twinfilin inhibit nucleotide exchange of actin monomers and present a model for how ADF/cofilin induces filament depolymerization by weakening intrafilament interactions.

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Schwartz, Niles, Melanie Hosford, Ruben M. Sandoval, Mark C. Wagner, Simon J. Atkinson, James Bamburg, and Bruce A. Molitoris. "Ischemia activates actin depolymerizing factor: role in proximal tubule microvillar actin alterations." American Journal of Physiology-Renal Physiology 276, no. 4 (April 1, 1999): F544—F551. http://dx.doi.org/10.1152/ajprenal.1999.276.4.f544.

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Apical membrane of renal proximal tubule cells is extremely sensitive to ischemia, with structural alterations occurring within 5 min. These changes are felt secondary to actin cytoskeletal disruption, yet the mechanism responsible is unknown. Actin depolymerizing factor (ADF), a 19-kDa actin-binding protein, has recently been shown to play an important role in regulation of actin filament dynamics. Because ADF is known to mediate pH-dependent F-actin binding, depolymerization, and severing, and because ADF activation occurs by dephosphorylation, we questioned whether ADF played a role in microvilli microfilament disruption during ischemia. To test our hypothesis, we induced renal ischemia in the rat with the clamp model. Initial immunofluorescence and Western blot studies on cortical tissue documented the presence of ADF in proximal tubule cells. Under physiological conditions, ADF was distributed homogeneously throughout the cytoplasm, primarily in the Triton X-100-soluble fraction, and both phosphorylated (pADF) and nonphosphorylated forms were identified. During ischemia, marked alterations occurred. Intraluminal vesicle/bleb structures contained extremely high concentrations of ADF along with G-actin, but not F-actin. Western blot showed a rapidly occurring duration-dependent dephosphorylation of ADF. At 0–30 min of ischemia, total ADF levels were unchanged, whereas pADF decreased significantly to 72% and 19% of control levels, at 5 and 15 min, respectively. Urine collected under physiological conditions did not contain ADF or actin, whereas urine collected after 30 min of ischemia contained both ADF and actin. Reperfusion was associated with normalization of cellular pADF levels, pADF intracellular distribution, and repair of apical microvilli. These data suggest that activation of ADF during ischemia via dephosphorylation is, in part, responsible for apical actin disruption resulting in microvillar destruction and formation of intraluminal vesicles.

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Schüler, Herwig, Ann-Kristin Mueller, and Kai Matuschewski. "A Plasmodium Actin-depolymerizing Factor That Binds Exclusively to Actin Monomers." Molecular Biology of the Cell 16, no. 9 (September 2005): 4013–23. http://dx.doi.org/10.1091/mbc.e05-02-0086.

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ADF/cofilins (AC) are essential F- and G-actin binding proteins that modulate microfilament turnover. The genome of Plasmodium falciparum, the parasite causing malaria, contains two members of the AC family. Interestingly, P. falciparum ADF1 lacks the F-actin binding residues of the AC consensus. Reverse genetics in the rodent malaria model system suggest that ADF1 performs vital functions during the pathogenic red blood cell stages, whereas ADF2 is not present in these stages. We show that recombinant PfADF1 interacts with monomeric actin but does not bind to actin polymers. Although other AC proteins inhibit nucleotide exchange on monomeric actin, the Plasmodium ortholog stimulates nucleotide exchange. Thus, PfADF1 differs in its biochemical properties from previously known AC proteins and seems to promote turnover exclusively by interaction with actin monomers. These findings provide important insights into the low cytosolic abundance and unique turnover characteristics of actin polymers in parasites of the phylum Apicomplexa.

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Hotulainen, Pirta, Eija Paunola, Maria K. Vartiainen, and Pekka Lappalainen. "Actin-depolymerizing Factor and Cofilin-1 Play Overlapping Roles in Promoting Rapid F-Actin Depolymerization in Mammalian Nonmuscle Cells." Molecular Biology of the Cell 16, no. 2 (February 2005): 649–64. http://dx.doi.org/10.1091/mbc.e04-07-0555.

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Actin-depolymerizing factor (ADF)/cofilins are small actin-binding proteins found in all eukaryotes. In vitro, ADF/cofilins promote actin dynamics by depolymerizing and severing actin filaments. However, whether ADF/cofilins contribute to actin dynamics in cells by disassembling “old” actin filaments or by promoting actin filament assembly through their severing activity is a matter of controversy. Analysis of mammalian ADF/cofilins is further complicated by the presence of multiple isoforms, which may contribute to actin dynamics by different mechanisms. We show that two isoforms, ADF and cofilin-1, are expressed in mouse NIH 3T3, B16F1, and Neuro 2A cells. Depleting cofilin-1 and/or ADF by siRNA leads to an accumulation of F-actin and to an increase in cell size. Cofilin-1 and ADF seem to play overlapping roles in cells, because the knockdown phenotype of either protein could be rescued by overexpression of the other one. Cofilin-1 and ADF knockdown cells also had defects in cell motility and cytokinesis, and these defects were most pronounced when both ADF and cofilin-1 were depleted. Fluorescence recovery after photobleaching analysis and studies with an actin monomer-sequestering drug, latrunculin-A, demonstrated that these phenotypes arose from diminished actin filament depolymerization rates. These data suggest that mammalian ADF and cofilin-1 promote cytoskeletal dynamics by depolymerizing actin filaments and that this activity is critical for several processes such as cytokinesis and cell motility.

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Morgan, TE, RO Lockerbie, LS Minamide, MD Browning, and JR Bamburg. "Isolation and characterization of a regulated form of actin depolymerizing factor." Journal of Cell Biology 122, no. 3 (August 1, 1993): 623–33. http://dx.doi.org/10.1083/jcb.122.3.623.

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Actin depolymerizing factor (ADF) is an 18.5-kD protein with pH-dependent reciprocal F-actin binding and severing/depolymerizing activities. We previously showed developing muscle down-regulates ADF (J. R. Bamburg and D. Bray. 1987. J. Cell Biol. 105: 2817-2825). To further study this process, we examined ADF expression in chick myocytes cultured in vitro. Surprisingly, ADF immunoreactivity increases during the first 7-10 d in culture. This increase is due to the presence of a new ADF species with higher relative molecular weight which reacts identically to brain ADF with antisera raised against either brain ADF or recombinant ADF. We have purified both ADF isoforms from myocytes and have shown by peptide mapping and partial sequence analysis that the new isoform is structurally related to ADF. Immunoprecipitation of both isoforms from extracts of cells prelabeled with [32P]orthophosphate showed that the new isoform is radiolabeled, predominantly on a serine residue, and hence is called pADF. pADF can be converted into a form which comigrates with ADF on 1-D and 2-D gels by treatment with alkaline phosphatase. pADF has been quantified in a number of cells and tissues where it is present from approximately 18% to 150% of the amount of unphosphorylated ADF. pADF, unlike ADF, does not bind to G-actin, or affect the rate or extent of actin assembly. Four ubiquitous protein kinases failed to phosphorylate ADF in vitro suggesting that ADF phosphorylation in vivo is catalyzed by a more specific kinase. We conclude that the ability to regulate ADF activity is important to muscle development since myocytes have both pre- and posttranslational mechanisms for regulating ADF activity. The latter mechanism is apparently a general one for cell regulation of ADF activity.

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Carlier, Marie-France, Valérie Laurent, Jérôme Santolini, Ronald Melki, Dominique Didry, Gui-Xian Xia, Yan Hong, Nam-Hai Chua, and Dominique Pantaloni. "Actin Depolymerizing Factor (ADF/Cofilin) Enhances the Rate of Filament Turnover: Implication in Actin-based Motility." Journal of Cell Biology 136, no. 6 (March 24, 1997): 1307–22. http://dx.doi.org/10.1083/jcb.136.6.1307.

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Actin-binding proteins of the actin depolymerizing factor (ADF)/cofilin family are thought to control actin-based motile processes. ADF1 from Arabidopsis thaliana appears to be a good model that is functionally similar to other members of the family. The function of ADF in actin dynamics has been examined using a combination of physical–chemical methods and actin-based motility assays, under physiological ionic conditions and at pH 7.8. ADF binds the ADPbound forms of G- or F-actin with an affinity two orders of magnitude higher than the ATP- or ADP-Pi– bound forms. A major property of ADF is its ability to enhance the in vitro turnover rate (treadmilling) of actin filaments to a value comparable to that observed in vivo in motile lamellipodia. ADF increases the rate of propulsion of Listeria monocytogenes in highly diluted, ADF-limited platelet extracts and shortens the actin tails. These effects are mediated by the participation of ADF in actin filament assembly, which results in a change in the kinetic parameters at the two ends of the actin filament. The kinetic effects of ADF are end specific and cannot be accounted for by filament severing. The main functionally relevant effect is a 25-fold increase in the rate of actin dissociation from the pointed ends, while the rate of dissociation from the barbed ends is unchanged. This large increase in the rate-limiting step of the monomer-polymer cycle at steady state is responsible for the increase in the rate of actin-based motile processes. In conclusion, the function of ADF is not to sequester G-actin. ADF uses ATP hydrolysis in actin assembly to enhance filament dynamics.

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Rosenblatt, Jody, Brian J. Agnew, Hiroshi Abe, James R. Bamburg, and Timothy J. Mitchison. "Xenopus Actin Depolymerizing Factor/Cofilin (XAC) Is Responsible for the Turnover of Actin Filaments in Listeria monocytogenes Tails." Journal of Cell Biology 136, no. 6 (March 24, 1997): 1323–32. http://dx.doi.org/10.1083/jcb.136.6.1323.

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In contrast to the slow rate of depolymerization of pure actin in vitro, populations of actin filaments in vivo turn over rapidly. Therefore, the rate of actin depolymerization must be accelerated by one or more factors in the cell. Since the actin dynamics in Listeria monocytogenes tails bear many similarities to those in the lamellipodia of moving cells, we have used Listeria as a model system to isolate factors required for regulating the rapid actin filament turnover involved in cell migration. Using a cell-free Xenopus egg extract system to reproduce the Listeria movement seen in a cell, we depleted candidate depolymerizing proteins and analyzed the effect that their removal had on the morphology of Listeria tails. Immunodepletion of Xenopus actin depolymerizing factor (ADF)/cofilin (XAC) from Xenopus egg extracts resulted in Listeria tails that were approximately five times longer than the tails from undepleted extracts. Depletion of XAC did not affect the tail assembly rate, suggesting that the increased tail length was caused by an inhibition of actin filament depolymerization. Immunodepletion of Xenopus gelsolin had no effect on either tail length or assembly rate. Addition of recombinant wild-type XAC or chick ADF protein to XAC-depleted extracts restored the tail length to that of control extracts, while addition of mutant ADF S3E that mimics the phosphorylated, inactive form of ADF did not reduce the tail length. Addition of excess wild-type XAC to Xenopus egg extracts reduced the length of Listeria tails to a limited extent. These observations show that XAC but not gelsolin is essential for depolymerizing actin filaments that rapidly turn over in Xenopus extracts. We also show that while the depolymerizing activities of XAC and Xenopus extract are effective at depolymerizing normal filaments containing ADP, they are unable to completely depolymerize actin filaments containing AMPPNP, a slowly hydrolyzible ATP analog. This observation suggests that the substrate for XAC is the ADP-bound subunit of actin and that the lifetime of a filament is controlled by its nucleotide content.

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Haase, Silvia, Dennis Zimmermann, Maya A. Olshina, Mark Wilkinson, Fabio Fisher, Yan Hong Tan, Rebecca J. Stewart, et al. "Disassembly activity of actin-depolymerizing factor (ADF) is associated with distinct cellular processes in apicomplexan parasites." Molecular Biology of the Cell 26, no. 17 (September 2015): 3001–12. http://dx.doi.org/10.1091/mbc.e14-10-1427.

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Proteins of the actin-depolymerizing factor (ADF)/cofilin family have been shown to be crucial for the motility and survival of apicomplexan parasites. However, the mechanisms by which ADF proteins fulfill their function remain poorly understood. In this study, we investigate the comparative activities of ADF proteins from Toxoplasma gondii and Plasmodium falciparum, the human malaria parasite, using a conditional T. gondii ADF-knockout line complemented with ADF variants from either species. We show that P. falciparum ADF1 can fully restore native TgADF activity, demonstrating functional conservation between parasites. Strikingly, mutation of a key basic residue (Lys-72), previously implicated in disassembly in PfADF1, had no detectable phenotypic effect on parasite growth, motility, or development. In contrast, organelle segregation was severely impaired when complementing with a TgADF mutant lacking the corresponding residue (Lys-68). Biochemical analyses of each ADF protein confirmed the reduced ability of lysine mutants to mediate actin depolymerization via filament disassembly although not severing, in contrast to previous reports. These data suggest that actin filament disassembly is essential for apicomplexan parasite development but not for motility, as well as pointing to genus-specific coevolution between ADF proteins and their native actin.

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Tsai, Cheng-Han, and Yi-Jang Lee. "Focus on ADF/Cofilin: Beyond Actin Cytoskeletal Regulation." ISRN Cell Biology 2012 (February 19, 2012): 1–7. http://dx.doi.org/10.5402/2012/597876.

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Actin depolymerizing factor (ADF)/cofilin, an actin-binding protein ubiquitously expressed in a variety of organisms, is required for regulation of actin dynamics. The activity of ADF/cofilin is dependent on serine 3 phosphorylation by LIM kinase (LIMK), which is regulated by the Rho small GTPase signaling pathway. ADF/cofilin is strongly associated with several important cell biological functions, including cell cycle, morphological maintenance, and locomotion. These functions affect several biological events, including embryogenesis, oncology, nephropathy, and neurodegenerations. Here, we focus on the biochemical and pathophysiological role of ADF/cofilin in mammals.

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Vartiainen, Maria K., Tuija Mustonen, Pieta K. Mattila, Pauli J. Ojala, Irma Thesleff, Juha Partanen, and Pekka Lappalainen. "The Three Mouse Actin-depolymerizing Factor/Cofilins Evolved to Fulfill Cell-Type–specific Requirements for Actin Dynamics." Molecular Biology of the Cell 13, no. 1 (January 2002): 183–94. http://dx.doi.org/10.1091/mbc.01-07-0331.

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Actin-depolymerizing factor (ADF)/cofilins are essential regulators of actin filament turnover. Several ADF/cofilin isoforms are found in multicellular organisms, but their biological differences have remained unclear. Herein, we show that three ADF/cofilins exist in mouse and most likely in all other mammalian species. Northern blot and in situ hybridization analyses demonstrate that cofilin-1 is expressed in most cell types of embryos and adult mice. Cofilin-2 is expressed in muscle cells and ADF is restricted to epithelia and endothelia. Although the three mouse ADF/cofilins do not show actin isoform specificity, they all depolymerize platelet actin filaments more efficiently than muscle actin. Furthermore, these ADF/cofilins are biochemically different. The epithelial-specific ADF is the most efficient in turning over actin filaments and promotes a stronger pH-dependent actin filament disassembly than the two other isoforms. The muscle-specific cofilin-2 has a weaker actin filament depolymerization activity and displays a 5–10-fold higher affinity for ATP-actin monomers than cofilin-1 and ADF. In steady-state assays, cofilin-2 also promotes filament assembly rather than disassembly. Taken together, these data suggest that the three biochemically distinct mammalian ADF/cofilin isoforms evolved to fulfill specific requirements for actin filament dynamics in different cell types.

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Galkin, Vitold E., Albina Orlova, Natalya Lukoyanova, Willy Wriggers, and Edward H. Egelman. "Actin Depolymerizing Factor Stabilizes an Existing State of F-Actin and Can Change the Tilt of F-Actin Subunits." Journal of Cell Biology 153, no. 1 (April 2, 2001): 75–86. http://dx.doi.org/10.1083/jcb.153.1.75.

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Proteins in the actin depolymerizing factor (ADF)/cofilin family are essential for rapid F-actin turnover, and most depolymerize actin in a pH-dependent manner. Complexes of human and plant ADF with F-actin at different pH were examined using electron microscopy and a novel method of image analysis for helical filaments. Although ADF changes the mean twist of actin, we show that it does this by stabilizing a preexisting F-actin angular conformation. In addition, ADF induces a large (∼12°) tilt of actin subunits at high pH where filaments are readily disrupted. A second ADF molecule binds to a site on the opposite side of F-actin from that of the previously described ADF binding site, and this second site is only largely occupied at high pH. All of these states display a high degree of cooperativity that appears to be an integral part of F-actin.

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Nakano, Kentaro, and Issei Mabuchi. "Actin-depolymerizing Protein Adf1 Is Required for Formation and Maintenance of the Contractile Ring during Cytokinesis in Fission Yeast." Molecular Biology of the Cell 17, no. 4 (April 2006): 1933–45. http://dx.doi.org/10.1091/mbc.e05-09-0900.

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The role of the actin-depolymerizing factor (ADF)/cofilin-family protein Adf1 in cytokinesis of fission yeast cells was studied. Adf1 was required for accumulation of actin at the division site by depolymerizing actin at the cell ends, assembly of the contractile ring through severing actin filaments, and maintenance of the contractile ring once formed. Genetic and cytological analyses suggested that it collaborates with profilin and capping protein in the mitotic reorganization of the actin cytoskeleton. Furthermore, it was unexpectedly found that Adf1 and myosin-II also collaborate in assembling the contractile ring. Tropomyosin was shown to antagonize the function of Adf1 in the contractile ring. We propose that formation and maintenance of the contractile ring are achieved by a balanced collaboration of these proteins.

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HUANG, XIAOHONG, YANG XU, GUOWEI GUO, QIANQIAN LIN, ZHONGFENG YE, LIPING YUAN, ZHIYU SUN, and WEI NI. "Molecular characterization of an actin depolymerizing factor from Cryptocaryon irritans." Parasitology 140, no. 5 (January 4, 2013): 561–68. http://dx.doi.org/10.1017/s0031182012001977.

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SUMMARYActin depolymerizing factors regulate actin dynamics involved in cellular processes such as morphogenesis, motility, development and infection. Here, a novel actin depolymerizing factor gene (CiADF2) was cloned from the cDNA library of Cryptocaryon irritans, a parasitic ciliate causing cryptocaryonosis. The full-length cDNA of CiADF2 was 531 bp. Its open reading frame (ORF) was 417 bp, encoding a polypeptide of 138 aa with typical features of the ADF/cofilin family. Reverse transcription-PCR suggested that CiADF2 is expressed in all stages of the life cycle. After site-directed mutagenesis of a non-universal genetic code, the ORF was subcloned in Escherichia coli. The bacteria were induced with the addition of isopropylthio-β-D-galactoside to express a fusion protein of recombinant CiADF2 (rCiADF2) with glutathione S transferase. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot confirmed the predicted molecular mass of rCiADF2 of 16·2 kDa. A mouse antibody against rCiADF2 recognized native CiADF2, and rCiADF2 reacted with mouse antisera against C. irritans trophonts. CiADF2 was abundant in the plasma around cytostomes, suggesting that CiADF2 is involved in ciliate movement. Moreover, rCiADF2 showed F-actin binding and depolymerizing activity. This study will help to clarify the pathogenic biology of the parasite and develop effective control measures for cryptocaryonosis.

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Shukla, Vaibhav Kumar, Ashish Kabra, Diva Maheshwari, Rahul Yadav, Anupam Jain, Sarita Tripathi, Shoichiro Ono, Dinesh Kumar, and Ashish Arora. "Solution structures and dynamics of ADF/cofilins UNC-60A and UNC-60B from Caenorhabditis elegans." Biochemical Journal 465, no. 1 (December 12, 2014): 63–78. http://dx.doi.org/10.1042/bj20140923.

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The relatively flexible vertical orientation of the F-site, as observed for UNC-60B, is associated with higher severing activity, whereas the relatively rigid inclined orientation of the F-site, as observed for UNC-60A, is associated with the weak severing activity of ADF (actin-depolymerizing factor)/cofilins.

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Paavilainen, Ville O., Michael C. Merckel, Sandra Falck, Pauli J. Ojala, Ehmke Pohl, Matthias Wilmanns, and Pekka Lappalainen. "Structural Conservation between the Actin Monomer-binding Sites of Twinfilin and Actin-depolymerizing Factor (ADF)/Cofilin." Journal of Biological Chemistry 277, no. 45 (August 30, 2002): 43089–95. http://dx.doi.org/10.1074/jbc.m208225200.

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Svitkina, Tatyana M., and Gary G. Borisy. "Arp2/3 Complex and Actin Depolymerizing Factor/Cofilin in Dendritic Organization and Treadmilling of Actin Filament Array in Lamellipodia." Journal of Cell Biology 145, no. 5 (May 31, 1999): 1009–26. http://dx.doi.org/10.1083/jcb.145.5.1009.

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The leading edge (∼1 μm) of lamellipodia in Xenopus laevis keratocytes and fibroblasts was shown to have an extensively branched organization of actin filaments, which we term the dendritic brush. Pointed ends of individual filaments were located at Y-junctions, where the Arp2/3 complex was also localized, suggesting a role of the Arp2/3 complex in branch formation. Differential depolymerization experiments suggested that the Arp2/3 complex also provided protection of pointed ends from depolymerization. Actin depolymerizing factor (ADF)/cofilin was excluded from the distal 0.4 μm of the lamellipodial network of keratocytes and in fibroblasts it was located within the depolymerization-resistant zone. These results suggest that ADF/cofilin, per se, is not sufficient for actin brush depolymerization and a regulatory step is required. Our evidence supports a dendritic nucleation model (Mullins, R.D., J.A. Heuser, and T.D. Pollard. 1998. Proc. Natl. Acad. Sci. USA. 95:6181–6186) for lamellipodial protrusion, which involves treadmilling of a branched actin array instead of treadmilling of individual filaments. In this model, Arp2/3 complex and ADF/cofilin have antagonistic activities. Arp2/3 complex is responsible for integration of nascent actin filaments into the actin network at the cell front and stabilizing pointed ends from depolymerization, while ADF/cofilin promotes filament disassembly at the rear of the brush, presumably by pointed end depolymerization after dissociation of the Arp2/3 complex.

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Reymann, Anne-Cécile, Cristian Suarez, Christophe Guérin, Jean-Louis Martiel, Christopher J. Staiger, Laurent Blanchoin, and Rajaa Boujemaa-Paterski. "Turnover of branched actin filament networks by stochastic fragmentation with ADF/cofilin." Molecular Biology of the Cell 22, no. 14 (July 15, 2011): 2541–50. http://dx.doi.org/10.1091/mbc.e11-01-0052.

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Cell motility depends on the rapid assembly, aging, severing, and disassembly of actin filaments in spatially distinct zones. How a set of actin regulatory proteins that sustains actin-based force generation during motility work together in space and time remains poorly understood. We present our study of the distribution and dynamics of Arp2/3 complex, capping protein (CP), and actin-depolymerizing factor (ADF)/cofilin in actin “comet tails,” using a minimal reconstituted system with nucleation-promoting factor (NPF)-coated beads. The Arp2/3 complex concentrates at nucleation sites near the beads as well as in the first actin shell. CP colocalizes with actin and is homogeneously distributed throughout the comet tail; it serves to constrain the spatial distribution of ATP/ADP-Pi filament zones to areas near the bead. The association of ADF/cofilin with the actin network is therefore governed by kinetics of actin assembly, actin nucleotide state, and CP binding. A kinetic simulation accurately validates these observations. Following its binding to the actin networks, ADF/cofilin is able to break up the dense actin filament array of a comet tail. Stochastic severing by ADF/cofilin loosens the tight entanglement of actin filaments inside the comet tail and facilitates turnover through the macroscopic release of large portions of the aged actin network.

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Abe, H., T. Obinata, L. S. Minamide, and J. R. Bamburg. "Xenopus laevis actin-depolymerizing factor/cofilin: a phosphorylation-regulated protein essential for development." Journal of Cell Biology 132, no. 5 (March 1, 1996): 871–85. http://dx.doi.org/10.1083/jcb.132.5.871.

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Two cDNAs, isolated from a Xenopus laevis embryonic library, encode proteins of 168 amino acids, both of which are 77% identical to chick cofilin and 66% identical to chick actin-depolymerizing factor (ADF), two structurally and functionally related proteins. These Xenopus ADF/cofilins (XADs) differ from each other in 12 residues spread throughout the sequence but do not differ in charge. Purified GST-fusion proteins have pH-dependent actin-depolymerizing and F-actin-binding activities similar to chick ADF and cofilin. Similarities in the developmental and tissue specific expression, embryonic localization, and in the cDNA sequence of the noncoding regions, suggest that the two XACs arise from allelic variants of the pseudotetraploid X. laevis. Immunofluorescence localization of XAC in oocyte sections with an XAC-specific monoclonal antibody shows it to be diffuse in the cortical cytoplasm. After fertilization, increased immunostaining is observed in two regions: along the membrane, particularly that of the vegetal hemisphere, and at the interface between the cortical and animal hemisphere cytoplasm. The cleavage furrow and the mid-body structure are stained at the end of first cleavage. Neuroectoderm derived tissues, notochord, somites, and epidermis stain heavily either continuously or transiently from stages 18-34. A phosphorylated form of XAC (pXAC) was identified by 2D Western blotting, and it is the only species found in oocytes. Dephosphorylation of >60% of the pXAC occurs within 30 min after fertilization. Injection of one blastomere at the 2 cell stage, either with constitutively active XAC or with an XAC inhibitory antibody, blocked cleavage of only the injected blastomere in a concentration-dependent manner without inhibiting nuclear division. The cleavage furrow of eggs injected with constitutively active XAC completely regressed. Blastomeres injected with neutralized antibody developed normally. These results suggest that XAC is necessary for cytokinesis and that its activity must be properly regulated for cleavage to occur.

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Ressad, Fariza, Dominique Didry, Gui-Xian Xia, Yan Hong, Nam-Hai Chua, Dominique Pantaloni, and Marie-France Carlier. "Kinetic Analysis of the Interaction of Actin-depolymerizing Factor (ADF)/Cofilin with G- and F-Actins." Journal of Biological Chemistry 273, no. 33 (August 14, 1998): 20894–902. http://dx.doi.org/10.1074/jbc.273.33.20894.

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Chang, Chun-Yuan, Jyh-Der Leu, and Yi-Jang Lee. "The Actin Depolymerizing Factor (ADF)/Cofilin Signaling Pathway and DNA Damage Responses in Cancer." International Journal of Molecular Sciences 16, no. 2 (February 13, 2015): 4095–120. http://dx.doi.org/10.3390/ijms16024095.

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Mohri, Kurato, Kanako Ono, Robinson Yu, Sawako Yamashiro, and Shoichiro Ono. "Enhancement of Actin-depolymerizing Factor/Cofilin-dependent Actin Disassembly by Actin-interacting Protein 1 Is Required for Organized Actin Filament Assembly in the Caenorhabditis elegans Body Wall Muscle." Molecular Biology of the Cell 17, no. 5 (May 2006): 2190–99. http://dx.doi.org/10.1091/mbc.e05-11-1016.

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Regulated disassembly of actin filaments is involved in several cellular processes that require dynamic rearrangement of the actin cytoskeleton. Actin-interacting protein (AIP) 1 specifically enhances disassembly of actin-depolymerizing factor (ADF)/cofilin-bound actin filaments. In vitro, AIP1 actively disassembles filaments, caps barbed ends, and binds to the side of filaments. However, how AIP1 functions in the cellular actin cytoskeletal dynamics is not understood. We compared biochemical and in vivo activities of mutant UNC-78 proteins and found that impaired activity of mutant UNC-78 proteins to enhance disassembly of ADF/cofilin-bound actin filaments is associated with inability to regulate striated organization of actin filaments in muscle cells. Six functionally important residues are present in the N-terminal β-propeller, whereas one residue is located in the C-terminal β-propeller, suggesting the presence of two separate sites for interaction with ADF/cofilin and actin. In vitro, these mutant UNC-78 proteins exhibited variable alterations in actin disassembly and/or barbed end-capping activities, suggesting that both activities are important for its in vivo function. These results indicate that the actin-regulating activity of AIP1 in cooperation with ADF/cofilin is essential for its in vivo function to regulate actin filament organization in muscle cells.

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Ortega-Ortega, Yolanda, Janet Carrasco-Castilla, Marco A. Juárez-Verdayes, Roberto Toscano-Morales, Citlali Fonseca-García, Noreide Nava, Luis Cárdenas, and Carmen Quinto. "Actin Depolymerizing Factor Modulates Rhizobial Infection and Nodule Organogenesis in Common Bean." International Journal of Molecular Sciences 21, no. 6 (March 13, 2020): 1970. http://dx.doi.org/10.3390/ijms21061970.

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Actin plays a critical role in the rhizobium–legume symbiosis. Cytoskeletal rearrangements and changes in actin occur in response to Nod factors secreted by rhizobia during symbiotic interactions with legumes. These cytoskeletal rearrangements are mediated by diverse actin-binding proteins, such as actin depolymerization factors (ADFs). We examined the function of an ADF in the Phaseolus vulgaris–rhizobia symbiotic interaction (PvADFE). PvADFE was preferentially expressed in rhizobia-inoculated roots and nodules. PvADFE promoter activity was associated with root hairs harbouring growing infection threads, cortical cell divisions beneath root hairs, and vascular bundles in mature nodules. Silencing of PvADFE using RNA interference increased the number of infection threads in the transgenic roots, resulting in increased nodule number, nitrogen fixation activity, and average nodule diameter. Conversely, overexpression of PvADFE reduced the nodule number, nitrogen fixation activity, average nodule diameter, as well as NODULE INCEPTION (NIN) and EARLY NODULIN2 (ENOD2) transcript accumulation. Hence, changes in ADFE transcript levels affect rhizobial infection and nodulation, suggesting that ADFE is fine-tuning these processes.

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Tada, Hirobumi, Tomoyuki Miyazaki, Kiwamu Takemoto, Kenkichi Takase, Susumu Jitsuki, Waki Nakajima, Mayu Koide, et al. "Neonatal isolation augments social dominance by altering actin dynamics in the medial prefrontal cortex." Proceedings of the National Academy of Sciences 113, no. 45 (October 25, 2016): E7097—E7105. http://dx.doi.org/10.1073/pnas.1606351113.

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Social separation early in life can lead to the development of impaired interpersonal relationships and profound social disorders. However, the underlying cellular and molecular mechanisms involved are largely unknown. Here, we found that isolation of neonatal rats induced glucocorticoid-dependent social dominance over nonisolated control rats in juveniles from the same litter. Furthermore, neonatal isolation inactivated the actin-depolymerizing factor (ADF)/cofilin in the juvenile medial prefrontal cortex (mPFC). Isolation-induced inactivation of ADF/cofilin increased stable actin fractions at dendritic spines in the juvenile mPFC, decreasing glutamate synaptic AMPA receptors. Expression of constitutively active ADF/cofilin in the mPFC rescued the effect of isolation on social dominance. Thus, neonatal isolation affects spines in the mPFC by reducing actin dynamics, leading to altered social behavior later in life.

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von Blume, Julia, Juan M. Duran, Elena Forlanelli, Anne-Marie Alleaume, Mikhail Egorov, Roman Polishchuk, Henrik Molina, and Vivek Malhotra. "Actin remodeling by ADF/cofilin is required for cargo sorting at the trans-Golgi network." Journal of Cell Biology 187, no. 7 (December 21, 2009): 1055–69. http://dx.doi.org/10.1083/jcb.200908040.

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Knockdown of the actin-severing protein actin-depolymerizing factor (ADF)/cofilin inhibited export of an exogenously expressed soluble secretory protein from Golgi membranes in Drosophila melanogaster and mammalian tissue culture cells. A stable isotope labeling by amino acids in cell culture mass spectrometry–based protein profiling revealed that a large number of endogenous secretory proteins in mammalian cells were not secreted upon ADF/cofilin knockdown. Although many secretory proteins were retained, a Golgi-resident protein and a lysosomal hydrolase were aberrantly secreted upon ADF/cofilin knockdown. Overall, our findings indicate that inactivation of ADF/cofilin perturbed the sorting of a subset of both soluble and integral membrane proteins at the trans-Golgi network (TGN). We suggest that ADF/cofilin-dependent actin trimming generates a sorting domain at the TGN, which filters secretory cargo for export, and that uncontrolled growth of this domain causes missorting of proteins. This type of actin-dependent compartmentalization and filtering of secretory cargo at the TGN by ADF/cofilin could explain sorting of proteins that are destined to the cell surface.

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Wioland, Hugo, Antoine Jegou, and Guillaume Romet-Lemonne. "Torsional stress generated by ADF/cofilin on cross-linked actin filaments boosts their severing." Proceedings of the National Academy of Sciences 116, no. 7 (January 28, 2019): 2595–602. http://dx.doi.org/10.1073/pnas.1812053116.

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Proteins of the actin depolymerizing factor (ADF)/cofilin family are the central regulators of actin filament disassembly. A key function of ADF/cofilin is to sever actin filaments. However, how it does so in a physiological context, where filaments are interconnected and under mechanical stress, remains unclear. Here, we monitor and quantify the action of ADF/cofilin in different mechanical situations by using single-molecule, single-filament, and filament network techniques, coupled to microfluidics. We find that local curvature favors severing, while tension surprisingly has no effect on cofilin binding and weakly enhances severing. Remarkably, we observe that filament segments that are held between two anchoring points, thereby constraining their twist, experience a mechanical torque upon cofilin binding. We find that this ADF/cofilin-induced torque does not hinder ADF/cofilin binding, but dramatically enhances severing. A simple model, which faithfully recapitulates our experimental observations, indicates that the ADF/cofilin-induced torque increases the severing rate constant 100-fold. A consequence of this mechanism, which we verify experimentally, is that cross-linked filament networks are severed by cofilin far more efficiently than nonconnected filaments. We propose that this mechanochemical mechanism is critical to boost ADF/cofilin’s ability to sever highly connected filament networks in cells.

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Roy-Zokan, Eileen M., Kelly A. Dyer, and Richard B. Meagher. "Phylogenetic Patterns of Codon Evolution in the ACTIN-DEPOLYMERIZING FACTOR/COFILIN (ADF/CFL) Gene Family." PLOS ONE 10, no. 12 (December 30, 2015): e0145917. http://dx.doi.org/10.1371/journal.pone.0145917.

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Wen, Zhexing, Liang Han, James R. Bamburg, Sangwoo Shim, Guo-li Ming, and James Q. Zheng. "BMP gradients steer nerve growth cones by a balancing act of LIM kinase and Slingshot phosphatase on ADF/cofilin." Journal of Cell Biology 178, no. 1 (July 2, 2007): 107–19. http://dx.doi.org/10.1083/jcb.200703055.

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Bone morphogenic proteins (BMPs) are involved in axon pathfinding, but how they guide growth cones remains elusive. In this study, we report that a BMP7 gradient elicits bidirectional turning responses from nerve growth cones by acting through LIM kinase (LIMK) and Slingshot (SSH) phosphatase to regulate actin-depolymerizing factor (ADF)/cofilin-mediated actin dynamics. Xenopus laevis growth cones from 4–8-h cultured neurons are attracted to BMP7 gradients but become repelled by BMP7 after overnight culture. The attraction and repulsion are mediated by LIMK and SSH, respectively, which oppositely regulate the phosphorylation-dependent asymmetric activity of ADF/cofilin to control the actin dynamics and growth cone steering. The attraction to repulsion switching requires the expression of a transient receptor potential (TRP) channel TRPC1 and involves Ca2+ signaling through calcineurin phosphatase for SSH activation and growth cone repulsion. Together, we show that spatial regulation of ADF/cofilin activity controls the directional responses of the growth cone to BMP7, and Ca2+ influx through TRPC tilts the LIMK-SSH balance toward SSH-mediated repulsion.

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Huang, Jun, Wei Sun, Jiaxin Ren, Ruichun Yang, Jingsheng Fan, Yunfeng Li, Xin Wang, Shija Joseph, Wenbin Deng, and Lihong Zhai. "Genome-Wide Identification and Characterization of Actin-Depolymerizing Factor (ADF) Family Genes and Expression Analysis of Responses to Various Stresses in Zea Mays L." International Journal of Molecular Sciences 21, no. 5 (March 4, 2020): 1751. http://dx.doi.org/10.3390/ijms21051751.

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Actin-depolymerizing factor (ADF) is a small class of actin-binding proteins that regulates the dynamics of actin in cells. Moreover, it is well known that the plant ADF family plays key roles in growth, development and defense-related functions. Results: Thirteen maize (Zea mays L., ZmADFs) ADF genes were identified using Hidden Markov Model. Phylogenetic analysis indicated that the 36 identified ADF genes in Physcomitrella patens, Arabidopsis thaliana, Oryza sativa japonica, and Zea mays were clustered into five groups. Four pairs of segmental genes were found in the maize ADF gene family. The tissue-specific expression of ZmADFs and OsADFs was analyzed using microarray data obtained from the Maize and Rice eFP Browsers. Five ZmADFs (ZmADF1/2/7/12/13) from group V exhibited specifically high expression in tassel, pollen, and anther. The expression patterns of 13 ZmADFs in seedlings under five abiotic stresses were analyzed using qRT-PCR, and we found that the ADFs mainly responded to heat, salt, drought, and ABA. Conclusions: In our study, we identified ADF genes in maize and analyzed the gene structure and phylogenetic relationships. The results of expression analysis demonstrated that the expression level of ADF genes was diverse in various tissues and different stimuli, including abiotic and phytohormone stresses, indicating their different roles in plant growth, development, and response to external stimulus. This report extends our knowledge to understand the function of ADF genes in maize.

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Wang, Jie, Xiao-Lin Kou, Cheng Chen, Mei Wang, Cui Qi, Jing Wang, Wei-Yan You, Gang Hu, Jiong Chen, and Jun Gao. "Hippocampal Wdr1 Deficit Impairs Learning and Memory by Perturbing F-actin Depolymerization in Mice." Cerebral Cortex 29, no. 10 (December 22, 2018): 4194–207. http://dx.doi.org/10.1093/cercor/bhy301.

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Abstract WD repeat protein 1 (Wdr1), known as a cofactor of actin-depolymerizing factor (ADF)/cofilin, is conserved among eukaryotes, and it plays a critical role in the dynamic reorganization of the actin cytoskeleton. However, the function of Wdr1 in the central nervous system remains elusive. Using Wdr1 conditional knockout mice, we demonstrated that Wdr1 plays a significant role in regulating synaptic plasticity and memory. The knockout mice exhibited altered reversal spatial learning and fear responses. Moreover, the Wdr1 CKO mice showed significant abnormalities in spine morphology and synaptic function, including enhanced hippocampal long-term potentiation and impaired long-term depression. Furthermore, we observed that Wdr1 deficiency perturbed actin rearrangement through regulation of the ADF/cofilin activity. Taken together, these results indicate that Wdr1 in the hippocampal CA1 area plays a critical role in actin dynamics in associative learning and postsynaptic receptor availability.

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Ontivero, Marta, Gustavo Martínez Zamora, Sergio Salazar, Juan Carlos Díaz Ricci, and Atilio Pedro Castagnaro. "Isolation of a strawberry gene fragment encoding an actin depolymerizing factor-like protein from genotypes resistant to Colletotrichum acutatum." Genome 54, no. 12 (December 2011): 1041–44. http://dx.doi.org/10.1139/g11-068.

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Actin depolymerizing factors (ADFs) have been recently implicated in plant defense against pathogenic fungi, associated with the cytoskeletal rearrangements that contribute to establish an effective barrier against fungal ingress. In this work, we identified a DNA fragment corresponding to a part of a gene predicted to encode an ADF-like protein in genotypes of Fragaria ananassa resistant to the fungus Colletotrichum acutatum . Bulked segregant analysis combined with AFLP was used to identify polymorphisms linked to resistance in hybrids derived from the cross between the resistant cultivar ‘Sweet Charlie’ and the susceptible cultivar ‘Pájaro’. The sequence of one out of three polymorphic bands detected showed significant BLASTX hits to ADF proteins from other plants. Two possible exons were identified and bioinformatic analysis revealed the presence of the ADF homology domain with two actin-binding sites, an N-terminal phosphorylation site, and a nuclear localization signal. In addition to its possible application in strawberry breeding programs, these finding may contribute to investigate the role of ADFs in plant resistance against fungi.

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Herde, Michel K., Eckhard Friauf, and Marco B. Rust. "Developmental expression of the actin depolymerizing factor ADF in the mouse inner ear and spiral ganglia." Journal of Comparative Neurology 518, no. 10 (May 15, 2010): 1724–41. http://dx.doi.org/10.1002/cne.22298.

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Léna, J. Y., J. R. Bamburg, A. Rabié, and C. Faivre-Sarrailh. "Actin-depolymerizing factor (ADF) in the cerebellum of the developing rat: A quantitative and immunocytochemical study." Journal of Neuroscience Research 30, no. 1 (September 1991): 18–27. http://dx.doi.org/10.1002/jnr.490300104.

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Ashworth, Sharon L., Sarah E. Wean, Silvia B. Campos, Constance J. Temm-Grove, Erica L. Southgate, Bernadette Vrhovski, Peter Gunning, Ron P. Weinberger, and Bruce A. Molitoris. "Renal ischemia induces tropomyosin dissociation-destabilizing microvilli microfilaments." American Journal of Physiology-Renal Physiology 286, no. 5 (May 2004): F988—F996. http://dx.doi.org/10.1152/ajprenal.00168.2003.

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Ischemic-induced cell injury results in rapid duration-dependent actin-depolymerizing factor (ADF)/cofilin-mediated disruption of the apical microvilli microfilament cores. Because intestinal microvillar microfilaments are bound and stabilized in the terminal web by the actin-binding protein tropomyosin, we questioned whether a protective effect of tropomyosin localization to the terminal web of the proximal tubule microfilament cores is disrupted during ischemic injury. With tropomyosin-specific antibodies, we examined rat cortical sections under physiological conditions and following ischemic injury by confocal microscopy. In addition, Western blot analysis of cortical extracts and urine was undertaken. Our studies demonstrated the presence of tropomyosin isoforms in the proximal tubule microvillar terminal web under physiological conditions and their dissociation in response to 25 min of ischemic injury. This correlated with the excretion of tropomyosin-containing plasma membrane vesicles in urine from ischemic rats. In addition, we noted increased tropomyosin Triton X-100 solubility following ischemia in cortical extracts. These studies suggest tropomyosin binds to and stabilizes the microvillar microfilament core in the terminal web under physiological conditions. With the onset of ischemic injury, we propose that tropomyosin dissociates from the microfilament core providing access to microfilaments in the terminal web for F-actin binding, severing and depolymerizing actions of ADF/cofilin proteins.

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Goode, Bruce L., David G. Drubin, and Pekka Lappalainen. "Regulation of the Cortical Actin Cytoskeleton in Budding Yeast by Twinfilin, a Ubiquitous Actin Monomer-sequestering Protein." Journal of Cell Biology 142, no. 3 (August 10, 1998): 723–33. http://dx.doi.org/10.1083/jcb.142.3.723.

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Here we describe the identification of a novel 37-kD actin monomer binding protein in budding yeast. This protein, which we named twinfilin, is composed of two cofilin-like regions. In our sequence database searches we also identified human, mouse, and Caenorhabditis elegans homologues of yeast twinfilin, suggesting that twinfilins form an evolutionarily conserved family of actin-binding proteins. Purified recombinant twinfilin prevents actin filament assembly by forming a 1:1 complex with actin monomers, and inhibits the nucleotide exchange reaction of actin monomers. Despite the sequence homology with the actin filament depolymerizing cofilin/actin-depolymerizing factor (ADF) proteins, our data suggests that twinfilin does not induce actin filament depolymerization. In yeast cells, a green fluorescent protein (GFP)–twinfilin fusion protein localizes primarily to cytoplasm, but also to cortical actin patches. Overexpression of the twinfilin gene (TWF1) results in depolarization of the cortical actin patches. A twf1 null mutation appears to result in increased assembly of cortical actin structures and is synthetically lethal with the yeast cofilin mutant cof1-22, shown previously to cause pronounced reduction in turnover of cortical actin filaments. Taken together, these results demonstrate that twinfilin is a novel, highly conserved actin monomer-sequestering protein involved in regulation of the cortical actin cytoskeleton.

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Ashworth, Sharon L., Ruben M. Sandoval, Melanie Hosford, James R. Bamburg, and Bruce A. Molitoris. "Ischemic injury induces ADF relocalization to the apical domain of rat proximal tubule cells." American Journal of Physiology-Renal Physiology 280, no. 5 (May 1, 2001): F886—F894. http://dx.doi.org/10.1152/ajprenal.2001.280.5.f886.

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Breakdown of proximal tubule cell apical membrane microvilli is an early-occurring hallmark of ischemic acute renal failure. Intracellular mechanisms responsible for these apical membrane changes remain unknown, but it is known that actin cytoskeleton alterations play a critical role in this cellular process. Our laboratory previously demonstrated that ischemia-induced cell injury resulted in dephosphorylation and activation of the actin-binding protein, actin depolymerizing factor [(ADF); Schwartz, N, Hosford M, Sandoval RM, Wagner MC, Atkinson SJ, Bamburg J, and Molitoris BA. Am J Physiol Renal Fluid Electrolyte Physiol 276: F544–F551, 1999]. Therefore, we postulated that ischemia-induced ADF relocalization from the cytoplasm to the apical microvillar microfilament core was an early event occurring before F-actin alterations. To directly investigate this hypothesis, we examined the intracellular localization of ADF in ischemic rat cortical tissues by immunofluorescence and quantified the concentration of ADF in brush-border membrane vesicles prepared from ischemic rat kidneys by using Western blot techniques. Within 5 min of the induction of ischemia, ADF relocalized to the apical membrane region. The length of ischemia correlated with the time-related increase in ADF in isolated brush-border membrane vesicles. Finally, depolymerization of microvillar F-actin to G-actin was documented by using colocalization studies for G- and F-actin. Collectively, these data indicate that ischemia induces ADF activation and relocalization to the apical domain before microvillar destruction. These data further suggest that ADF plays a critical role in microvillar microfilament destruction and apical membrane damage during ischemia.

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Ono, Shoichiro, Kazumi Nomura, Sadae Hitosugi, Domena K. Tu, Jocelyn A. Lee, David L. Baillie, and Kanako Ono. "The two actin-interacting protein 1 genes have overlapping and essential function for embryonic development in Caenorhabditis elegans." Molecular Biology of the Cell 22, no. 13 (July 2011): 2258–69. http://dx.doi.org/10.1091/mbc.e10-12-0934.

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Disassembly of actin filaments by actin-depolymerizing factor (ADF)/cofilin and actin-interacting protein 1 (AIP1) is a conserved mechanism to promote reorganization of the actin cytoskeleton. We previously reported that unc-78, an AIP1 gene in the nematode Caenorhabditis elegans, is required for organized assembly of sarcomeric actin filaments in the body wall muscle. unc-78 functions in larval and adult muscle, and an unc-78–null mutant is homozygous viable and shows only weak phenotypes in embryos. Here we report that a second AIP1 gene, aipl-1 (AIP1-like gene-1), has overlapping function with unc-78, and that depletion of the two AIP1 isoforms causes embryonic lethality. A single aipl-1–null mutation did not cause a detectable phenotype. However, depletion of both unc-78 and aipl-1 arrested development at late embryonic stages due to severe disorganization of sarcomeric actin filaments in body wall muscle. In vitro, both AIPL-1 and UNC-78 preferentially cooperated with UNC-60B, a muscle-specific ADF/cofilin isoform, in actin filament disassembly but not with UNC-60A, a nonmuscle ADF/cofilin. AIPL-1 is expressed in embryonic muscle, and forced expression of AIPL-1 in adult muscle compensated for the function of UNC-78. Thus our results suggest that enhancement of actin filament disassembly by ADF/cofilin and AIP1 proteins is critical for embryogenesis.

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Nomura, Kazumi, and Shoichiro Ono. "ATP-dependent regulation of actin monomer–filament equilibrium by cyclase-associated protein and ADF/cofilin." Biochemical Journal 453, no. 2 (June 28, 2013): 249–59. http://dx.doi.org/10.1042/bj20130491.

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CAP (cyclase-associated protein) is a conserved regulator of actin filament dynamics. In the nematode Caenorhabditis elegans, CAS-1 is an isoform of CAP that is expressed in striated muscle and regulates sarcomeric actin assembly. In the present study, we report that CAS-2, a second CAP isoform in C. elegans, attenuates the actin-monomer-sequestering effect of ADF (actin depolymerizing factor)/cofilin to increase the steady-state levels of actin filaments in an ATP-dependent manner. CAS-2 binds to actin monomers without a strong preference for either ATP– or ADP–actin. CAS-2 strongly enhances the exchange of actin-bound nucleotides even in the presence of UNC-60A, a C. elegans ADF/cofilin that inhibits nucleotide exchange. UNC-60A induces the depolymerization of actin filaments and sequesters actin monomers, whereas CAS-2 reverses the monomer-sequestering effect of UNC-60A in the presence of ATP, but not in the presence of only ADP or the absence of ATP or ADP. A 1:100 molar ratio of CAS-2 to UNC-60A is sufficient to increase actin filaments. CAS-2 has two independent actin-binding sites in its N- and C-terminal halves, and the C-terminal half is necessary and sufficient for the observed activities of the full-length CAS-2. These results suggest that CAS-2 (CAP) and UNC-60A (ADF/cofilin) are important in the ATP-dependent regulation of the actin monomer–filament equilibrium.

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Chaudhry, Faisal, Christophe Guérin, Matthias von Witsch, Laurent Blanchoin, and Christopher J. Staiger. "Identification of Arabidopsis Cyclase-associated Protein 1 as the First Nucleotide Exchange Factor for Plant Actin." Molecular Biology of the Cell 18, no. 8 (August 2007): 3002–14. http://dx.doi.org/10.1091/mbc.e06-11-1041.

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The actin cytoskeleton powers organelle movements, orchestrates responses to abiotic stresses, and generates an amazing array of cell shapes. Underpinning these diverse functions of the actin cytoskeleton are several dozen accessory proteins that coordinate actin filament dynamics and construct higher-order assemblies. Many actin-binding proteins from the plant kingdom have been characterized and their function is often surprisingly distinct from mammalian and fungal counterparts. The adenylyl cyclase-associated protein (CAP) has recently been shown to be an important regulator of actin dynamics in vivo and in vitro. The disruption of actin organization in cap mutant plants indicates defects in actin dynamics or the regulated assembly and disassembly of actin subunits into filaments. Current models for actin dynamics maintain that actin-depolymerizing factor (ADF)/cofilin removes ADP–actin subunits from filament ends and that profilin recharges these monomers with ATP by enhancing nucleotide exchange and delivery of subunits onto filament barbed ends. Plant profilins, however, lack the essential ability to stimulate nucleotide exchange on actin, suggesting that there might be a missing link yet to be discovered from plants. Here, we show that Arabidopsis thaliana CAP1 (AtCAP1) is an abundant cytoplasmic protein; it is present at a 1:3 M ratio with total actin in suspension cells. AtCAP1 has equivalent affinities for ADP– and ATP–monomeric actin (Kd ∼ 1.3 μM). Binding of AtCAP1 to ATP–actin monomers inhibits polymerization, consistent with AtCAP1 being an actin sequestering protein. However, we demonstrate that AtCAP1 is the first plant protein to increase the rate of nucleotide exchange on actin. Even in the presence of ADF/cofilin, AtCAP1 can recharge actin monomers and presumably provide a polymerizable pool of subunits to profilin for addition onto filament ends. In turnover assays, plant profilin, ADF, and CAP act cooperatively to promote flux of subunits through actin filament barbed ends. Collectively, these results and our understanding of other actin-binding proteins implicate CAP1 as a central player in regulating the pool of unpolymerized ATP–actin.

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Ashworth, Sharon L., Erica L. Southgate, Ruben M. Sandoval, Peter J. Meberg, James R. Bamburg, and Bruce A. Molitoris. "ADF/cofilin mediates actin cytoskeletal alterations in LLC-PK cells during ATP depletion." American Journal of Physiology-Renal Physiology 284, no. 4 (April 1, 2003): F852—F862. http://dx.doi.org/10.1152/ajprenal.00210.2002.

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Ischemic injury induces actin cytoskeleton disruption and aggregation, but mechanisms affecting these changes remain unclear. To determine the role of actin-depolymerizing factor (ADF)/ cofilin participation in ischemic-induced actin cytoskeletal breakdown, we utilized porcine kidney cultured cells, LLC-PKA4.8, and adenovirus containing wild-type (wt), constitutively active, and inactive Xenopus ADF/cofilin linked to green fluorescence protein [XAC(wt)-GFP] in an ATP depletion model. High adenoviral infectivity (70%) in LLC-PKA4.8 cells resulted in linearly increasing XAC(wt)-GFP and phosphorylated (p)XAC(wt)-GFP (inactive) expression. ATP depletion rapidly induced dephosphorylation, and, therefore, activation, of endogenous pcofilin as well as pXAC(wt)-GFP in conjunction with the formation of fluorescent XAC(wt)-GFP/actin aggregates and rods. No significant actin cytoskeletal alterations occurred with short-term ATP depletion of LLC-PKA4.8 cells expressing GFP or the constitutively inactive mutant XAC(S3E)-GFP, but cells expressing the constitutively active mutant demonstrated nearly instantaneous actin disruption with aggregate and rod formation. Confocal image three-dimensional volume reconstructions of normal and ATP-depleted LLC-PKA4.8 cells demonstrated that 25 min of ATP depletion induced a rapid increase in XAC(wt)-GFP apical and basal signal in addition to XAC-GFP/actin aggregate formation. These data demonstrate XAC(wt)-GFP participates in ischemia-induced actin cytoskeletal alterations and determines the rate and extent of these ATP depletion-induced cellular alterations.

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Ostrowska, Zofia, and Joanna Moraczewska. "Cofilin – a protein controlling dynamics of actin filaments." Postępy Higieny i Medycyny Doświadczalnej 71, no. 1 (May 5, 2017): 0. http://dx.doi.org/10.5604/01.3001.0010.3818.

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Cofilins are evolutionary conserved proteins present in all Eukaryotic cells. Their primary function is dynamic reorganization of actin cytoskeleton. Two cofilin isoforms are known: cofilin 1, present in all studied non-muscle cells and in embryonic muscle cells, and cofilin 2, which dominates in mature skeletal and cardiac muscles. Polypeptide chains of both isoforms fold into a structure homological to a conservative ADF (actin depolymerizing factor) domain, which is characteristic of actin depolymerizing factor. In cofilin molecule two actin-binding sites were found. One site binds monomeric and filamentous actin, the second one interacts only with the filament. Binding of cofilin to actin filament causes a change in the orientation of subunits, which results in filament severing. This increases number of ends which can either elongate or shorten the filament, depending on the conditions. Cofilin interactions with monomeric actin decreases availability of polymerization-competent actin subunits. Cofilin activity is controlled by phosphorylation, binding membrane phospholipids, local pH and oxidative stress. Under conditions of oxidative stress oxidation of cysteine residues leads to formation of dimers, which are able to cross-link actin filaments. Stable actin-cofilin rods save cellular ATP, which is not used during active polymerization process. This facilitates faster cell recovery from the stress. The final cellular reaction on the environmental stimuli is a resultant of cofilin activity and activities of other actin-binding proteins, which function either synergistically or antagonistically. Due to the central role in the regulation of actinfilaments dynamics, cofilin is involved in development of cancer, neurodegenerative diseases, congenital myopathies and cardiomyopathies.

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Nevalainen, Elisa M., Aneta Skwarek-Maruszewska, Attila Braun, Markus Moser, and Pekka Lappalainen. "Two biochemically distinct and tissue-specific twinfilin isoforms are generated from the mouse Twf2 gene by alternative promoter usage." Biochemical Journal 417, no. 2 (December 23, 2008): 593–600. http://dx.doi.org/10.1042/bj20080608.

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Twf (twinfilin) is an evolutionarily conserved regulator of actin dynamics composed of two ADF-H (actin-depolymerizing factor homology) domains. Twf binds actin monomers and heterodimeric capping protein with high affinity. Previous studies have demonstrated that mammals express two Twf isoforms, Twf1 and Twf2, of which at least Twf1 also regulates cytoskeletal dynamics by capping actin filament barbed-ends. In the present study, we show that alternative promoter usage of the mouse Twf2 gene generates two isoforms, which differ from each other only at their very N-terminal region. Of these isoforms, Twf2a is predominantly expressed in non-muscle tissues, whereas expression of Twf2b is restricted to heart and skeletal muscle. Both proteins bind actin monomers and capping protein, as well as efficiently capping actin filament barbed-ends. However, the N-terminal ADF-H domain of Twf2b interacts with ADP-G-actin with a 5-fold higher affinity than with ATP-G-actin, whereas the corresponding domain of Twf2a binds ADP-G-actin and ATP-G-actin with equal affinities. Taken together, these results show that, like Twf1, mouse Twf2 is a filament barbed-end capping protein, and that two tissue-specific and biochemically distinct isoforms are generated from the Twf2 gene through alternative promoter usage.

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Ono, Shoichiro. "The Caenorhabditis elegans unc-78 Gene Encodes a Homologue of Actin-Interacting Protein 1 Required for Organized Assembly of Muscle Actin Filaments." Journal of Cell Biology 152, no. 6 (March 19, 2001): 1313–20. http://dx.doi.org/10.1083/jcb.152.6.1313.

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Assembly and maintenance of myofibrils require dynamic regulation of the actin cytoskeleton. In Caenorhabditis elegans, UNC-60B, a muscle-specific actin depolymerizing factor (ADF)/cofilin isoform, is required for proper actin filament assembly in body wall muscle (Ono, S., D.L. Baillie, and G.M. Benian. 1999. J. Cell Biol. 145:491–502). Here, I show that UNC-78 is a homologue of actin-interacting protein 1 (AIP1) and functions as a novel regulator of actin organization in myofibrils. In unc-78 mutants, the striated organization of actin filaments is disrupted, and large actin aggregates are formed in the body wall muscle cells, resulting in defects in their motility. Point mutations in unc-78 alleles change conserved residues within different WD repeats of the UNC-78 protein and cause less severe phenotypes than a deletion allele, suggesting that these mutations partially impair the function of UNC-78. UNC-60B is normally localized in the diffuse cytoplasm and to the myofibrils in wild type but mislocalized to the actin aggregates in unc-78 mutants. Similar Unc-78 phenotypes are observed in both embryonic and adult muscles. Thus, AIP1 is an important regulator of actin filament organization and localization of ADF/cofilin during development of myofibrils.

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Maciver, Sutherland K., Brian J. Pope, Sue Whytock, and Alan G. Weeds. "The effect of two actin depolymerizing factors (ADF/cofilins) on actin filament turnover : pH sensitivity of F-actin binding by human ADF, but not of Acanthamoeba actophorin." European Journal of Biochemistry 256, no. 2 (September 1998): 388–97. http://dx.doi.org/10.1046/j.1432-1327.1998.2560388.x.

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Suurna, Maria V., Sharon L. Ashworth, Melanie Hosford, Ruben M. Sandoval, Sarah E. Wean, Bijal M. Shah, James R. Bamburg, and Bruce A. Molitoris. "Cofilin mediates ATP depletion-induced endothelial cell actin alterations." American Journal of Physiology-Renal Physiology 290, no. 6 (June 2006): F1398—F1407. http://dx.doi.org/10.1152/ajprenal.00194.2005.

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Ischemia and sepsis lead to endothelial cell damage, resulting in compromised microvascular flow in many organs. Much remains to be determined regarding the intracellular structural events that lead to endothelial cell dysfunction. To investigate potential actin cytoskeletal-related mechanisms, ATP depletion was induced in mouse pancreatic microvascular endothelial cells (MS1). Fluorescent imaging and biochemical studies demonstrated a rapid and progressive increase in F-actin along with a decrease in G-actin at 60 min. Confocal microscopic analysis showed ATP depletion resulted in destruction of actin stress fibers and accumulation of F-actin aggregates. We hypothesized these actin alterations were secondary to dephosphorylation/activation of actin-depolymerizing factor (ADF)/cofilin proteins. Cofilin, the predominant isoform expressed in MS1 cells, was rapidly dephosphorylated/activated during ATP depletion. To directly investigate the role of cofilin activation on the actin cytoskeleton during ischemia, MS1 cells were infected with adenoviruses containing the cDNAs for wild-type Xenopus laevis ADF/cofilin green fluorescent protein [XAC(wt)-GFP], GFP, and the constitutively active and inactive isoforms XAC(S3A)-GFP and XAC(S3E)-GFP. The rate and extent of cortical actin destruction and actin aggregate formation were increased in ATP-depleted XAC(wt)-GFP- and XAC(S3A)-GFP-expressing cells, whereas increased actin stress fibers were observed in XAC(S3E)-GFP-expressing cells. To investigate the upstream signaling pathway of ADF/cofilin, LIM kinase 1-GFP (LIMK1-GFP) was expressed in MS1 cells. Cells expressing LIMK1-GFP protein had higher levels of phosphorylated ADF/cofilin, increased stress fibers, and delayed F-actin cytoskeleton destruction during ATP depletion. These results strongly support the importance of cofilin regulation in ischemia-induced endothelial cell actin cytoskeleton alterations leading to cell damage and microvascular dysfunction.

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Anyanful, Akwasi, Kanako Ono, Robert C. Johnsen, Hinh Ly, Victor Jensen, David L. Baillie, and Shoichiro Ono. "The RNA-binding protein SUP-12 controls muscle-specific splicing of the ADF/cofilin pre-mRNA in C. elegans." Journal of Cell Biology 167, no. 4 (November 15, 2004): 639–47. http://dx.doi.org/10.1083/jcb.200407085.

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Tissue-specific alternative pre-mRNA splicing is essential for increasing diversity of functionally different gene products. In Caenorhabditis elegans, UNC-60A and UNC-60B, nonmuscle and muscle isoforms of actin depolymerizing factor (ADF)/cofilin, are expressed by alternative splicing of unc-60 and regulate distinct actin-dependent developmental processes. We report that SUP-12, a member of a new family of RNA recognition motif (RRM) proteins, including SEB-4, regulates muscle-specific splicing of unc-60. In sup-12 mutants, expression of UNC-60B is decreased, whereas UNC-60A is up-regulated in muscle. sup-12 mutations strongly suppress muscle defects in unc-60B mutants by allowing expression of UNC-60A in muscle that can substitute for UNC-60B, thus unmasking their functional redundancy. SUP-12 is expressed in muscle and localized to the nuclei in a speckled pattern. The RRM domain of SUP-12 binds to several sites of the unc-60 pre-mRNA including the UG repeats near the 3′-splice site in the first intron. Our results suggest that SUP-12 is a novel tissue-specific splicing factor and regulates functional redundancy among ADF/cofilin isoforms.

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Moriyama, Kenji, and Ichiro Yahara. "Human CAP1 is a key factor in the recycling of cofilin and actin for rapid actin turnover." Journal of Cell Science 115, no. 8 (April 15, 2002): 1591–601. http://dx.doi.org/10.1242/jcs.115.8.1591.

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Cofilin-ADF (actin-depolymerizing factor) is an essential driver of actin-based motility. We discovered two proteins, p65 and p55, that are components of the actin-cofilin complex in a human HEK293 cell extract and identified p55 as CAP1/ASP56, a human homologue of yeast CAP/SRV2(cyclase-associated protein). CAP is a bifunctional protein with an N-terminal domain that binds to Ras-responsive adenylyl cyclase and a C-terminal domain that inhibits actin polymerization. Surprisingly, we found that the N-terminal domain of CAP1, but not the C-terminal domain, is responsible for the interaction with the actin-cofilin complex. The N-terminal domain of CAP1 was also found to accelerate the depolymerization of F-actin at the pointed end,which was further enhanced in the presence of cofilin and/or the C-terminal domain of CAP1. Moreover, CAP1 and its C-terminal domain were observed to facilitate filament elongation at the barbed end and to stimulate ADP-ATP exchange on G-actin, a process that regenerates easily polymerizable G-actin. Although cofilin inhibited the nucleotide exchange on G-actin even in the presence of the C-terminal domain of CAP1, its N-terminal domain relieved this inhibition. Thus, CAP1 plays a key role in speeding up the turnover of actin filaments by effectively recycling cofilin and actin and through its effect on both ends of actin filament.

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Shukla, Vaibhav Kumar, Ashish Kabra, Rahul Yadav, Shoichiro Ono, Dinesh Kumar, and Ashish Arora. "NMR assignments of actin depolymerizing factor (ADF) like UNC-60A and cofilin like UNC-60B proteins of Caenorhabditis elegans." Biomolecular NMR Assignments 9, no. 2 (December 11, 2014): 261–65. http://dx.doi.org/10.1007/s12104-014-9588-5.

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Purde, Vedud, Florian Busch, Elena Kudryashova, Vicki H. Wysocki, and Dmitri S. Kudryashov. "Oligomerization Affects the Ability of Human Cyclase-Associated Proteins 1 and 2 to Promote Actin Severing by Cofilins." International Journal of Molecular Sciences 20, no. 22 (November 12, 2019): 5647. http://dx.doi.org/10.3390/ijms20225647.

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Actin-depolymerizing factor (ADF)/cofilins accelerate actin turnover by severing aged actin filaments and promoting the dissociation of actin subunits. In the cell, ADF/cofilins are assisted by other proteins, among which cyclase-associated proteins 1 and 2 (CAP1,2) are particularly important. The N-terminal half of CAP has been shown to promote actin filament dynamics by enhancing ADF-/cofilin-mediated actin severing, while the central and C-terminal domains are involved in recharging the depolymerized ADP–G-actin/cofilin complexes with ATP and profilin. We analyzed the ability of the N-terminal fragments of human CAP1 and CAP2 to assist human isoforms of “muscle” (CFL2) and “non-muscle” (CFL1) cofilins in accelerating actin dynamics. By conducting bulk actin depolymerization assays and monitoring single-filament severing by total internal reflection fluorescence (TIRF) microscopy, we found that the N-terminal domains of both isoforms enhanced cofilin-mediated severing and depolymerization at similar rates. According to our analytical sedimentation and native mass spectrometry data, the N-terminal recombinant fragments of both human CAP isoforms form tetramers. Replacement of the original oligomerization domain of CAPs with artificial coiled-coil sequences of known oligomerization patterns showed that the activity of the proteins is directly proportional to the stoichiometry of their oligomerization; i.e., tetramers and trimers are more potent than dimers, which are more effective than monomers. Along with higher binding affinities of the higher-order oligomers to actin, this observation suggests that the mechanism of actin severing and depolymerization involves simultaneous or consequent and coordinated binding of more than one N-CAP domain to F-actin/cofilin complexes.

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Journal articles on the topic 'Actin depolymerizing factor (ADF)'

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