Phosphorylation of the Rab exchange factor Sec2p directs a switch in regulatory binding partners

Abstract

Sec2p is a guanine nucleotide exchange factor that promotes exocytosis by activating the Rab GTPase Sec4p. Sec2p is highly phosphorylated, and we have explored the role of phosphorylation in the regulation of its function. We have identified three phosphosites and demonstrate that phosphorylation regulates the interaction of Sec2p with its binding partners Ypt32p, Sec15p, and phosphatidyl-inositol-4-phosphate. In its nonphosphorylated form, Sec2p binds preferentially to the upstream Rab, Ypt32p-GTP, thus forming a Rab guanine nucleotide exchange factor cascade that leads to the activation of the downstream Rab, Sec4p. The nonphosphorylated form of Sec2p also binds to the Golgi-associated phosphatidyl-inositol-4-phosphate, which works in concert with Ypt32p-GTP to recruit Sec2p to Golgi-derived secretory vesicles. In contrast, the phosphorylated form of Sec2p binds preferentially to Sec15p, a downstream effector of Sec4p and a component of the exocyst tethering complex, thus forming a positivefeedback loop that prepares the secretory vesicle for fusion with the plasma membrane. Our results suggest that the phosphorylation state of Sec2p can direct a switch in its regulatory binding partners that facilitates maturation of the secretory vesicle and helps to promote the directionality of vesicular transport.

Activation of Rab8 guanine nucleotide exchange factor Rabin8 by ERK1/2 in response to EGF signaling

“…Phosphorylation of Rabin8 at S272 promotes binding to the downstream exocyst subunit Sec15 and inhibits binding to phosphatidylserine. In the budding yeast Saccharomyces cerevisiae, the Rabin8 homolog Sec2p is phosphorylated at multiple sites (35). Although the responsible kinases remain unknown, phosphorylation of Sec2p switches its binding from Ypt32p and PI(4)P at the Golgi to Sec15p.…”

“…Although the responsible kinases remain unknown, phosphorylation of Sec2p switches its binding from Ypt32p and PI(4)P at the Golgi to Sec15p. Therefore, the phosphorylation is thought to facilitate maturation of the secretory vesicle and promote their directional transport (35). In Candida albicans, Sec2p was shown to be phosphorylated by the cell cycle kinase Cdc28, and this phosphorylation is important for hyphal development (38).…”

Novel topography of the Rab11-effector interaction network within a ciliary membrane targeting complex

“…Remarkably (HxRxxS/T) is absent from Sec2, nevertheless S186 and S188 in the linker region are also phosphorylated by another kinase 80 . Clustal2w was used for the alignment.…”

“…region directs a switch in binding from Ypt32 toSec15 79,80 . This mode of action is not conserved, as Sec15 is a common effector for Rab11 and Rab864,65 , which differs from Ypt32 that does not associate with the exocyst complex.…”

Septin-Dependent Assembly of the Exocyst Is Essential for Plant Infection by Magnaporthe oryzae

“…In the budding yeast Saccharomyces cerevisiae, the polarisome complex consists of Peanut Shmoo 2, Spindle Pole Antigen 2 (Spa2), and the formin Bud Neck Involved 1, which nucleates F-actin cables to sites of polarized growth (Sheu et al, 1998;Sagot et al, 2002;Evangelista et al, 2003). Post-Golgi secretory vesicles are then delivered to the tip via the Rab GTPase Sec4, which is activated through its guanine nucleotide exchange factor (GEF), Sec2 (Stalder et al, 2013), and binds to the exocyst complex through Sec15 interaction (Salminen and Novick, 1989). The exocyst complex is an evolutionarily conserved octameric protein complex, comprising Sec3p, Sec5p, Sec6p, Sec8p, Sec10p, Sec15p, Exo70p, and Exo84p required for vesicle docking to the plasma membrane (TerBush et al, 1996;Guo et al, 1999;He and Guo, 2009).…”