The mechanism by which Rab GTPases are targeted to and activated at specific organelles remains a poorly understood process. It is known that Rab specific guanine nucleotide exchange factors (GEFs) play an important role in this process. Here we present studies that investigate a novel mechanism of Rab GEF regulation, whereby subunit exchange modifies its Rab specificity. In addition, there are examples of GEF independent Rab targeting, suggesting that other factors have this activity. In our work, we identify an ABC transporter that acts in Rab organelle recruitment. C. elegans gut granules are lysosome-related organelles (LROs) found only within intestinal cells whose biogenesis requires the activity of conserved complexes that act in LRO formation in mammals. It is known that the CCZ-1/SAND-1 complex acts as a GEF to activate RAB-7 and promote early endosome to late endosome maturation and RAB-5 to RAB-7 conversion. Our studies suggest that CCZ-1, but not SAND-1, functions with GLO-3 as a GEF for the Rab32/38 related Rab GLO-1 which functions in LRO biogenesis. We show that
ccz-1(-) and
glo-3(-), but not
sand-1(-), mutants exhibit defects in gut granule biogenesis and the cytoplasmic accumulation of GLO-1.
ccz-1(-) and
sand-1(-) mutants display defects in conventional endosome biogenesis that are not exhibited by
glo-1(-) and
glo-3(-) mutants. A fast guanine nucleotide exchange GLO-1 mutant restores gut granules, which contain GLO-1, in
ccz-1(-) and
glo-3(-) mutants, but not other mutants defective in gut granule formation. Moreover, while
ccz-1(-) displays protein trafficking defects distinct from
glo-1(-) and
glo-3(-),
glo-1(-);
sand-1(-) and
glo-3(-);
sand-1(-) double mutants resemble
ccz-1(-). These results are consistent with CCZ-1 acting with SAND-1 as a GEF for RAB-7 in endosome maturation and with GLO-3 as a GEF for GLO-1 in gut granule biogenesis. We screened our collection of gut granule biogenesis mutants for cytoplasmic accumulation of GLO-1 and identified a phenotype similar to
ccz-1(-) and
glo-3(-) in
wht-2(-) mutants. WHT-2 is an ABCG family membrane transporter, which we find is localized to the gut granule membrane.
wht-2(-) mutants contain partially formed gut granules, that lack GLO-1. Interestingly, WHT-2 functions in the polarized distribution of gut granules, but not other organelles, in embryonic intestinal cells. We find that the fast nucleotide exchange GLO-1 mutant does not restore the gut granule association of GLO-1 or gut granule formation in
wht-2(-) mutants. However, WHT-2 is required for the ability of GLO-1 fast nucleotide exchange mutants to bypass the requirement of GLO-3 but not CCZ-1 in gut granule biogenesis. Together, these data point to a GEF independent role of WHT-2 in GLO-1 organelle recruitment and activation.