Vertebrate nonmuscle myosin II isoforms rescue small interfering RNA-induced defects in COS-7 cell cytokinesis

J Biol Chem. 2005 May 20;280(20):19594-9. doi: 10.1074/jbc.M501573200. Epub 2005 Mar 17.


RNA interference (RNAi) treatment of monkey COS-7 cells, a cell line that lacks nonmuscle myosin heavy chain II-A (NMHC II-A) but contains NMHC II-B and II-C, was used to investigate the participation of NMHC isoforms in cytokinesis. We specifically suppressed the expression of NMHC II-B or II-C using 21 nucleotide small interfering RNA (siRNA) duplexes. Down-regulation of NMHC II-B protein expression to 10.2 +/- 0.7% inhibited COS-7 cell proliferation by 50% in the RNAi-treated cells compared with control cells. Moreover, whereas 8.7 +/- 1.0% of control cells were multinucleated, 62.4 +/- 8.8% of the NMHC II-B RNAi-treated cells were multinucleated 72 h after transfection. The RNAi-treated cells had increased surface areas and, unlike control cells, lacked actin stress fibers. Treatment of the COS-7 cells with NMHC II-C siRNA decreased NMHC II-C expression to 5.2 +/- 0.1% compared with the endogenous content of II-C; however, down-regulation of NMHC II-C did not cause increased multinucleation. Immunoblot analysis using a pan-myosin antibody showed that the content of NMHC II-C was less than one-twentieth the amount of NMHC II-B, thereby explaining the lack of response to II-C siRNA. Introducing green fluorescent protein (GFP)-tagged NMHC II isoforms into II-B siRNA-treated cells resulted in reduction of multinucleation from 62.4 +/- 8.8% to 17.8 +/- 2.2% using GFP-NMHC II-B, to 29.8 +/- 7.4% using GFP-NMHC II-A, and to 34.1 +/- 8.6% using NMHC II-C-GFP. These studies have shown that expression of endogenous NMHC II-C in COS-7 cells is insufficient for normal cytokinesis and that exogenous NMHC II-A and NMHC II-C can, at least partially, rescue the defect in cytokinesis due to the loss of NMHC II-B.

MeSH terms

  • Actins / metabolism
  • Animals
  • Base Sequence
  • COS Cells
  • Chlorocebus aethiops
  • Cytokinesis
  • Gene Expression
  • Myosin Type II / antagonists & inhibitors
  • Myosin Type II / genetics*
  • Myosin Type II / metabolism*
  • Nonmuscle Myosin Type IIA / antagonists & inhibitors
  • Nonmuscle Myosin Type IIA / genetics
  • Nonmuscle Myosin Type IIA / metabolism
  • Nonmuscle Myosin Type IIB / antagonists & inhibitors
  • Nonmuscle Myosin Type IIB / genetics
  • Nonmuscle Myosin Type IIB / metabolism
  • Protein Isoforms / antagonists & inhibitors
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • RNA Interference
  • RNA, Small Interfering / genetics*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Transfection


  • Actins
  • Protein Isoforms
  • RNA, Small Interfering
  • Recombinant Proteins
  • Myosin Type II
  • Nonmuscle Myosin Type IIA
  • Nonmuscle Myosin Type IIB