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

doi:10.1083/jcb.147.3.559

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© The Rockefeller University Press, 0021-9525/1999//559 $5.00
The Journal of Cell Biology, Volume 147, Number 3, , 1999 559-576

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. Chung^a and Richard A. Firtel^a

^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

We have identified a Dictyostelium discoideum gene encodinga serine/threonine kinase, PAKa, a putative member of the Ste20/PAKfamily of p21-activated kinases, with a kinase domain and along NH₂-terminal regulatory domain containing an acidic segment,a polyproline domain, and a CRIB domain. PAKa colocalizes withmyosin II to the cleavage furrow of dividing cells and the posteriorof 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 cellmovement, suppressing lateral pseudopod extension, and properretraction of the posterior of chemotaxing cells. paka nullcells are defective in myosin II assembly, as the myosin IIcap in the posterior of chemotaxing cells and myosin II assemblyinto cytoskeleton upon cAMP stimulation are absent in thesecells, while constitutively active PAKa leads to an upregulationof myosin II assembly. PAKa kinase activity against histone2B is transiently stimulated and PAKa incorporates into thecytoskeleton with kinetics similar to those of myosin II assemblyin response to chemoattractant signaling. However, PAKa doesnot phosphorylate myosin II. We suggest that PAKa is a majorregulator of myosin II assembly, but does so by negatively regulatingmyosin II heavy chain kinase.

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

1.used in this paper: GFP, green fluorescent protein; H2B, histone2B; MIHCK, myosin I heavy chain kinase; myrPAKa, a PAKa carryingthe NH2-terminal myristoylation signal from Src; PAKaCA, a putativeconstitutively 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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