ISOLATION AND CHARACTERIZATION OF A NEVER CYTOKINESIS-DEFICIENT MUTANT IN DICTYOSTELIUM-DISCOIDEUM

Cytokinesis is a dramatic event in the life of any cell during which n umerous mechanisms must coordinate the legitimate and complete mechani cal separation into two daughter cells. We have used Dictyostelium dis coideum as a model system to study this highly orchestrated event thro ugh genetic analysis. Transformants were generated using a method of i nsertional mutagenesis known as restriction enzyme-mediated integratio n (REMI) and subsequently screened for defects in cytokinesis. Mutants isolated in a similar screen suffered a disruption in the myosin II h eavy chain gene, a protein known to be essential for cytokinesis and i n a novel gene encoding a rho-like protein termed racE [Larochelle et al., 1996]. In the screen reported here we isolated a third type of mu tant, called 10BH2, which also had a complete defect in cytokinesis. 1 0BH2 mutant cells are able to propagate on tissue culture plates by fr agmenting into smaller cells by a process known as traction-mediated c ytofission. However, when grown in suspension culture, 10BH2 cells fai l to divide and become large and multinucleate. Phenotypic characteriz ation of the mutant cells showed that other cytoskeletal functions are preserved. The distribution of myosin and actin is identical to wild type cells. The cells can chemotax, phagocytose, cap crosslinked recep tors, and contract normally. However, the 10BH2 mutants are unable to complete the Dictyostelium developmental program beyond the finger sta ge. The mutant cells contain functional genes for myosin II heavy and light chains and the racE gene. Thus, based on these findings, we conc lude that 10BH2 represents a novel cytokinesis-deficient mutant. (C) 1 996 Wiley-Liss, Inc.