Rab11 is phosphorylated by classical and novel protein kinase C isoenzymes upon sustained phorbol ester activation.

PAVAROTTI M,; CAPMANY A, ; VITALE N.; COLOMBO MI,; DAMIANI, MT

BIOLOGY OF THE CELL

PORTLAND PRESS LTD

Lugar: Londres; Año: 2012 vol. 104 p. 102 - 115

0248-4900

Rab11 is a small GTPase that controls diverse intracellular trafficking pathways but the molecular machinery that regulates the participation of Rab11 in those different transport events is poorly understood. In resting cells, Rab11 localizes at the endocytic recycling compartment whereas the different Protein Kinase C (PKC) isoforms display a cytosolic distribution. Sustained phorbol ester stimulation induces the translocation of the classical PKCá and PKCâII isoenzymes to the endocytic recycling compartment enriched in Rab11, and results in transferrin recycling inhibition. In contrast, novel PKCä and PKCå isoenzymes neither redistribute to the perinucleus nor modify transferrin recycling transport after phorbol esters stimulation. Whereas several Rabs have been shown to be phosphorylated, there is no evidence that Rab11 as a kinase substrate yet. In this report, we show that Rab11 appears phosphorylated in vivo in phorbol esters-stimulated cells. A bioinformatic analysis of Rab11 allowed us to identify several high-probability Ser/Thr kinase phosphorylation sites. Our results demonstrate that classical Protein Kinase C (PKCá and PKCâII but not PKCâI) directly phosphorylate Rab11 in vitro. In addition, novel PKCå and PKCç but not PKCä isoenzymes also phosphorylate Rab11. Mass spectroscopy analysis revealed that Ser 177 is Rab11 residue to be phosphorylated in vitro by either PKCâII or PKCå. In agreement, the phosphomimetic mutant, Rab11 S177D, retains transferrin at the endocytic recycling compartment without PMA stimulus. Hence this report shows for the first time that Rab11 is differentially phosphorylated by distinct PKC isoenzymes and this post-translational modification might be a regulatory mechanism of intracellular trafficking.