Differential activities, subcellular distribution and tissue expression patterns of three members of Slingshot family phosphatases that dephosphorylate cofilin.
BACKGROUND
Cofilin, a key regulator of actin filament dynamics, is inactivated by phosphorylation at Ser-3 by LIM-kinases and is reactivated by dephosphorylation by a family of protein phosphatases, termed Slingshot (SSH).


RESULTS
We have identified two novel isoforms of SSHs, termed SSH-2L and SSH-3L and characterized them in comparison with SSH-1L that was previously reported. SSH-1L and SSH-2L, but not SSH-3L, tightly bound to and co-localized with actin filaments. When expressed in cultured cells, SSH-1L, SSH-2L and SSH-3L decreased the level of Ser-3-phosphorylated cofilin (P-cofilin) in cells and suppressed LIM-kinase-induced actin reorganization, although SSH-3L was less effective than SSH-1L and SSH-2L. In cell-free assays, SSH-1L and SSH-2L efficiently dephosphorylated P-cofilin, whereas SSH-3L did do so only weakly. Using deleted mutants of SSH-1L and SSH-2L, we found that the N-terminal and C-terminal extracatalytic regions are critical for cofilin-phosphatase and F-actin-binding activities, respectively. In situ hybridization analyses revealed characteristic patterns of expression of each of the mouse Ssh genes in both neuronal and non-neuronal tissues; in particular, expression of Ssh-3 in epithelial tissues is evident.


CONCLUSIONS
SSH-1L, SSH-2L and SSH-3L have the potential to dephosphorylate P-cofilin, but subcellular distribution, F-actin-binding activity, specific phosphatase activity and expression patterns significantly differ, which suggests that they have related but distinct functions in various cellular and developmental events.
Ohta
Yusaku
Y
Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi, Japan.
Kousaka
Kazuyoshi
K
Nagata-Ohashi
Kyoko
K
Ohashi
Kazumasa
K
Muramoto
Aya
A
Shima
Yasuyuki
Y
Niwa
Ryusuke
R
Uemura
Tadashi
T
Mizuno
Kensaku
K
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001483:Base Sequence; D001921:Brain / Q000196:embryology / Q000201:enzymology* / Q000378:metabolism; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D054461:Lim Kinases; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D051339:Actin Depolymerizing Factors; D000199:Actins; D000911:Antibodies, Monoclonal; D007527:Isoenzymes; D008840:Microfilament Proteins; D012333:RNA, Messenger; D011494:Protein Kinases; C515892:LIMK1 protein, human; D054461:Lim Kinases; C515893:Limk1 protein, mouse; D010744:Phosphoric Monoester Hydrolases