Actin tyrosine dephosphorylation by the Src homology 1-containing protein tyrosine phosphatase is essential for actin depolymerization after membrane IgM cross-linking.
Src homology protein 1 (SHP-1) plays an important role in B cell Ag receptor (BCR) differentiation, proliferation, survival, and apoptosis. After BCR stimulation in apoptotic cells, SHP-1 has been shown to be recruited to phosphorylated immunoreceptor tyrosine-based inhibitory motifs present in receptors such as CD22 and CD72. However, the substrates of SHP-1 in the chicken B cell line, DT40, have remained undefined. To identify SHP-1 substrates in DT40, we used a trapping mutant, SHP-1 C/S (a catalytically inactive form). Cross-linking of BCR induced hyperphosphorylation of approximately 44-kDa protein in C/S transfectants. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis revealed that this was actin (cytoplasmic type 5) carrying three immunoreceptor tyrosine-based inhibitory motif-like sequences. SHP-1 was shown to bind to one of these sequences in synthetic peptide binding experiment. Thus, actin is a direct SHP-1 substrate. Furthermore, more SHP-1 molecules translocate into lipid rafts, and their association with actin was increased after BCR stimulation. In C/S transfectants, actin polymerization induced by membrane IgM ligation was sustained to a greater extent for a longer time compared with wild-type transfectants. Therefore, actin dephosphorylation by SHP-1 is essential for actin depolymerization after BCR stimulation. Our data suggest that SHP-1 plays a pivotal role in reorganization of cytoskeletal architecture inducing actin dephosphorylation. These results clearly demonstrate the direct interaction of SHP-1 with actin.
Baba
Takeshi
T
Research Institute for Biological Sciences, Tokyo University of Science, Chiba, Japan.
Fusaki
Noemi
N
Shinya
Nobuko
N
Iwamatsu
Akihiro
A
Hozumi
Nobumichi
N
D000199:Actins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002645:Chickens; D003432:Cross-Linking Reagents / Q000378:metabolism*; D003545:Cysteine / Q000235:genetics; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D021962:Membrane Microdomains / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011108:Polymers / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism*; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism*; D012694:Serine / Q000235:genetics; D013379:Substrate Specificity / Q000235:genetics; D014162:Transfection; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism*; D018909:src Homology Domains / Q000235:genetics
D000199:Actins; D003432:Cross-Linking Reagents; D047908:Intracellular Signaling Peptides and Proteins; D011108:Polymers; D011947:Receptors, Antigen, B-Cell; D014443:Tyrosine; D012694:Serine; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; D003545:Cysteine