The cell polarity proteins Boi1p and Boi2p stimulate vesicle fusion at the plasma membrane of yeast cells

Abstract

Eukaryotic cells can direct secretion to defined regions of their plasma membrane. These regions are distinguished by an elaborate architecture of proteins and lipids that are specialized to capture and fuse post-Golgi vesicles. Here, we show that the proteins Boi1p and Boi2p are important elements of this area of active exocytosis at the tip of growing yeast cells. Cells lacking Boi1p and Boi2p accumulate secretory vesicles in their buds. The essential PH domains of Boi1p and Boi2p interact with Sec1p, a protein required for SNARE complex formation and vesicle fusion. Sec1p loses its tip localization in cells depleted of Boi1p and Boi2p but overexpression of Sec1p can partially compensate for their loss. The capacity to simultaneously bind phospholipids, Sec1p, multiple subunits of the exocyst, Cdc42p and the module for generating active Cdc42p identify Boi1p and Boi2p as essential mediators between exocytosis and polar growth.

An actin nucleation complex catalyzes filament formation at sites of exocytosis

“…Additionally removing the SAM domain of Boi1 (boi1 ∆ 299 ) and thus any residual interactions with the Bud6-Bni1 complex further lowered the amount of actin cables and enhanced the number of delocalized actin patches to similar levels as found in cells expressing only Boi1 ∆ 414 . The impaired vesicle fusion of a ∆ boi1∆boi2 strain can be suppressed by overexpression of the t-SNARE Sso1 (Kustermann et al 2017). The still disorganized actin structure of this strain confirms that actin organization and vesicle docking are distinct activities of Boi1/2 ( Fig.…”

“…By focusing our analysis on the boi1 Δ 86-178 -allele, we tried to separate its influence on actin filament formation from the protein`s two other main functions, the localization of the Bem1-Cdc24 complex and the formation of the tethering and docking complex (Bender et al 1996;Kustermann et al 2017). The former activity is located on a short binding motif in the middle of the sequence of Boi1, whereas the latter activity locates on the membrane-and Sec1-binding C-terminal PH domain, and the Exo84-, Sec3binding N-terminal SH3 domain (Bender et al 1996;Kustermann et al 2017). We propose that the concentration of all three activities in one protein coordinates vesicle fusion with trafficking and enables the control of both activities through Rho-GTPases ( Fig.…”

“…Pop1 complements the essential function of Boi1/2 in vesicle fusion (Kustermann et al 2017). These findings indicate that the molecules and mechanisms involved in the transfer of secretory vesicles from their long-distance carrier to cortical actin structures are quite conserved.…”

“…Boi1 and Boi2 are homologous proteins that assist in the tethering and fusion of post-Golgi vesicles at the plasma membrane and thus qualify as candidates for organizing actin filaments in the bud (Kustermann et al 2017;Masgrau et al 2017). Both proteins were already shown to interact with Bud6 in vivo (Kustermann et al 2017). As Boi1 and Boi2 perform their essential functions redundantly, we performed our molecular analysis predominantly on Boi1 (Kustermann et al 2017).…”

“…Both proteins were already shown to interact with Bud6 in vivo (Kustermann et al 2017). As Boi1 and Boi2 perform their essential functions redundantly, we performed our molecular analysis predominantly on Boi1 (Kustermann et al 2017). Split-Ub interaction analysis in strains lacking either BUD6 or BNI1 confirmed that both proteins interact independently of each other with Boi1 ( Fig.…”