THE INVOLVEMENT OF THE RHO GENE-PRODUCT, A SMALL MOLECULAR-WEIGHT GTP-BINDING PROTEIN, IN POLYPLOIDIZATION OF A HUMAN MEGAKARYOCYTIC CELL-LINE, CMK

The role of rho proteins, which are ras p21-related small GTP-binding proteins, in megakaryocyte endomitosis was examined using a botulinum C3 exoenzyme (C3), a rho inactivating enzyme. The megakaryocytic leuke mia cell line CMK expressed high levels of rhoA and rhoC mRNAs, wherea s rhoB mRNA was expressed at a very low level. The addition of C3 to t he culture medium caused ADP-ribosylation of the rho proteins in CMK c ells in a dose- and time-dependent manner. This procedure also induced a higher frequency of polyploid cells with increased glycoprotein (GP ) IIb/IIIa antigens on the cells. This effect of C3 on both ploidy and the antigen expression was abolished by prior incubation of C3 with a n anti-C3 monoclonal antibody. Cytochalasin B, an actin polymerization inhibitor, also induced polyploid cells; however, it did not stimulat e the expression of GP IIb/IIIa antigens in CMK cells. This finding su ggests that C3-induced increase in the expression of GP IIb/IIIa antig ens was not through the actin microfilament disassembly. The present s tudy suggests that the I ho p21 is a partly regulatory component in po lyploidization and GP IIb/IIIa antigen expression of a human megakaryo cytic cell line, CMK.