RRC ID 41523
Author Gerashchenko BI, Ueda K, Hino M, Hosoya H.
Title Phosphorylation at threonine-18 in addition to phosphorylation at serine-19 on myosin-II regulatory light chain is a mitosis-specific event.
Journal Cytometry
Abstract BACKGROUND:Cell division is an inevitable and vitally indispensable event in cell life, when the nucleus and cytoskeleton undergo profound reorganization. Cytoplasmic division (cytokinesis) is known to occur immediately after the end of nuclear division, when the nuclear envelope breaks down, and chromosomes condense and segregate, but its driving mechanism remains enigmatic. Myosin, particularly myosin-II, is thought to be required for cytokinesis as a force-generating element, the activity of which is mainly regulated through phosphorylations on its 20-kDa regulatory light chains (RLCs).
MATERIALS AND METHODS:Multiparameter flow cytometric analysis was performed on fixed HeLa S3 cells (suspension culture cells) sequentially stained with the polyclonal antibody (termed PP1) against both phosphorylated sites (serine-19 and threonine-18) on the RLC, and with propidium iodide for DNA. "Positive" cells were sorted, followed by their microscopic examination. Fluorescence microscopy was employed to visualize the cell-cycle-dependent distribution of immunolabeled diphosphorylated RLCs in both HeLa S3 and adherent HeLa cells.
RESULTS AND CONCLUSIONS:Doubly phosphorylated myosin RLCs were highly expressed in mitotic cells, suggesting the positive regulatory role of diphosphorylation in the redistribution of RLCs between daughter cells and then in cytokinesis. The increased immunofluorescence signal from the phosphorylated forms of RLC, together with flow cytometry, provides a clue with which to investigate the mechanisms governing the function of nonmuscle myosins during various cell motile events, including cytokinesis.
Volume 47(3)
Pages 150-7
Published 2002-3-1
DOI 10.1002/cyto.10059
PII 10.1002/cyto.10059
PMID 11891719
MeSH Amino Acid Sequence / physiology Antibody Specificity / immunology Cell Cycle / physiology Cell Division / physiology Cytoskeleton DNA / analysis Fluorescent Antibody Technique HeLa Cells Humans Mitosis / physiology* Myosin Light Chains / metabolism* Myosin Type II / metabolism* Phosphorylation Serine / metabolism Threonine / metabolism
Times Cited 8
WOS Category BIOCHEMICAL RESEARCH METHODS CELL BIOLOGY
Resource
Human and Animal Cells