The small GTPase Rab4A interacts with the central region of cytoplasmic dynein light intermediate chain-1

Rab4 belongs to the Rab family of small GTPases involved in the regulation of intracellular transport, and has been localized to early endosomes. We h ave employed the yeast two-hybrid system to identify proteins that specific ally interact with Rab4AQ67L, a GTPase-deficient mutant form of Rab4A. Scre ening a mouse embryo cDNA library identified a clone (M449) that interacted with Rab4A in a nucleotide-dependent fashion. Data base searches identifie d this clone as the mouse cytoplasmic dynein light intermediate chain-1 (LI C-1). Based on this finding, the full-length equivalent human cytoplasmic d ynein LIC-1 was isolated by PCR. When Rab4A was overexpressed together with either M449 or dynein LIC-1 in HeLa cells, the proteins were found to colo calize in the perinuclear region. We characterize the localization of both overexpressed human dynein LIC-1 and the endogenous protein with respect to microtubules and show that it concentrates to the microtubule-organizing c enter and mitotic spindle. Additionally, GFPRab4A endosomes localize to mic rotubules and are redistributed by nocodazole treatment. This is the first described interaction between cytoplasmic dynein, a retrograde motor protei n, and a Rab protein. (C) 2001 Academic Press.