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J. Cell Biol. 147 (3): 559-576

Copyright © 1999 by the Rockefeller University Press.

Original Article

Paka, a Putative Pak Family Member, Is Required for Cytokinesis and the Regulation of the Cytoskeleton in Dictyostelium discoideum Cells during Chemotaxis

Chang Y. Chunga, and Richard A. Firtela

a Department of Biology, Center for Molecular Genetics, University of California San Diego, La Jolla, California 92093-0634
Center for Molecular Genetics, Room 225, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0634.(858) 534-7073(858) 534-2788

rafirtel{at}ucsd.edu

Abstract: We have identified a Dictyostelium discoideum gene encoding a serine/threonine kinase, PAKa, a putative member of the Ste20/PAK family of p21-activated kinases, with a kinase domain and a long NH2-terminal regulatory domain containing an acidic segment, a polyproline domain, and a CRIB domain. PAKa colocalizes with myosin II to the cleavage furrow of dividing cells and the posterior of polarized, chemotaxing cells via its NH2-terminal domain. paka null cells are defective in completing cytokinesis in suspension. PAKa is also required for maintaining the direction of cell movement, suppressing lateral pseudopod extension, and proper retraction of the posterior of chemotaxing cells. paka null cells are defective in myosin II assembly, as the myosin II cap in the posterior of chemotaxing cells and myosin II assembly into cytoskeleton upon cAMP stimulation are absent in these cells, while constitutively active PAKa leads to an upregulation of myosin II assembly. PAKa kinase activity against histone 2B is transiently stimulated and PAKa incorporates into the cytoskeleton with kinetics similar to those of myosin II assembly in response to chemoattractant signaling. However, PAKa does not phosphorylate myosin II. We suggest that PAKa is a major regulator of myosin II assembly, but does so by negatively regulating myosin II heavy chain kinase.

Key Words: Dictyostelium discoideum • PAK • myosin II • chemotaxis • Rac/Cdc42

1.used in this paper: GFP, green fluorescent protein; H2B, histone 2B; MIHCK, myosin I heavy chain kinase; myrPAKa, a PAKa carrying the NH2-terminal myristoylation signal from Src; PAKaCA, a putative constitutively active PAKa containing only the kinase domain; PAKaDN, kinase dead, putative dominant, negative mutant of PAKa; PAKaG+K, a PAKa mutant containing the CRIB and kinase domains

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