Calponin 3 regulates actin cytoskeleton rearrangement in trophoblastic cell fusion.
Cell-cell fusion is an intriguing differentiation process, essential for placental development and maturation. A proteomic approach identified a cytoplasmic protein, calponin 3 (CNN3), related to the fusion of BeWo choriocarcinoma cells. CNN3 was expressed in cytotrophoblasts in human placenta. CNN3 gene knockdown promoted actin cytoskeletal rearrangement and syncytium formation in BeWo cells, suggesting CNN3 to be a negative regulator of trophoblast fusion. Indeed, CNN3 depletion promoted BeWo cell fusion. CNN3 at the cytoplasmic face of cytoskeleton was dislocated from F-actin with forskolin treatment and diffused into the cytoplasm in a phosphorylation-dependent manner. Phosphorylation sites were located at Ser293/296 in the C-terminal region, and deletion of this region or site-specific disruption of Ser293/296 suppressed syncytium formation. These CNN3 mutants were colocalized with F-actin and remained there after forskolin treatment, suggesting that dissociation of CNN3 from F-actin is modulated by the phosphorylation status of the C-terminal region unique to CNN3 in the CNN family proteins. The mutant missing these phosphorylation sites displayed a dominant negative effect on cell fusion, while replacement of Ser293/296 with aspartic acid enhanced syncytium formation. These results indicated that CNN3 regulates actin cytoskeleton rearrangement which is required for the plasma membranes of trophoblasts to become fusion competent.
Shibukawa
Yukinao
Y
Department of Molecular Medicine, Osaka Medical Center and Research Institute for Maternal and Child Health, 840 Murodo-cho, Izumi, Osaka 594-1101, Japan.
Yamazaki
Natsuko
N
Kumasawa
Keiichi
K
Daimon
Etsuko
E
Tajiri
Michiko
M
Okada
Yuka
Y
Ikawa
Masahito
M
Wada
Yoshinao
Y
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002459:Cell Fusion; D045744:Cell Line, Tumor; D002822:Choriocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005576:Colforsin / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism*; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D011485:Protein Binding / Q000187:drug effects; D034622:RNA Interference; D014327:Trophoblasts / Q000378:metabolism* / Q000473:pathology; D000096985:Calponins
D000199:Actins; D002135:Calcium-Binding Proteins; D005576:Colforsin; D008164:Luminescent Proteins; D008840:Microfilament Proteins; D000096985:Calponins