Localization of a mammalian homolog of diaphanous, mDia1, to the mitotic spindle in HeLa cells.

摘要: mDia1 is a mammalian homolog of Drosophila diaphanous and works as an effector the small GTPase Rho. It member formin homology (FH) proteins contains Rho-binding domain FH3 region in its N terminus, FH1 containing polyproline stretches middle FH2 C terminus. Several lines evidence indicate that are essential cytokinesis. present large amount cytoplasm both interphase mitotic cells. Using instantaneous fixation method preferentially extracts soluble components, we have analyzed localization HeLa Immunocytochemistry using polyclonal anti-mDia1 antibody revealed specific immunofluorescence localized to spindle. This was seen from prophase telophase. Western blot analysis also detected immunoreactivity spindle fraction isolated Consistently, expression full-length fusion protein with green fluorescence (GFP) GFP again Expression fusions various truncated mutants identified this determined by 173 amino acid-long sequence between region, which C-terminal part region. Point mutation Leu(434) Leu(455) Neither electroporation botulinum C3 exoenzyme nor microinjection Val14RhoA into cells affected endogenous spindle, suggesting localizes independent Rho activity. The study has thus established suggests role spindle-cleavage furrow interaction during cell division.