Alternative splicing of the human Rab6A gene generates two close but functionally different isoforms

Analysis of the human Rab6A gene structure reveals the presence of a duplic ated exon, and incorporation of either of the two exons by alternative spli cing is shown to generate two Rab6 isoforms named Rab6A and Rab6A', which d iffer in only three amino acid residues located in regions flanking the PM3 GTP-binding domain of the proteins. These isoforms are ubiquitously expres sed at similar levels, exhibit the same GTP-binding properties, and are loc alized to the Golgi apparatus. Overexpression of the GTP-bound mutants of R ab6A (Rab6A Q72L) or Rab6A' (Rab6A' Q72L) inhibits secretion in HeLa cells, but overexpression of Rab6A' Q72L does not induce the redistribution of Go lgi proteins into the endoplasmic reticulum. This suggests that Rab6A' is n ot able to stimulate Golgi-to-endoplasmic reticulum retrograde transport, a s described previously for Rab6A. In addition, Rab6A' interacts with two Ra b6A partners, GAPCenA and "clone 1," but not: with the kinesin-like protein Rabkinesin-6, a Golgi-associated Rab6A effector. Interestingly, we found t hat the functional differences between Rab6A and Rab6A' are contingent on o ne amino acid (T or A at position 87). Therefore, limited amino acid substi tutions within a Rab protein introduced by alternative splicing could repre sent a mechanism to generate functionally different isoforms that interact with distinct sets of effectors.