Mutations in the SAC1 gene suppress defects in yeast Golgi and yeast actin function.

Abstract

Abstract. The budding mode of Saccharomyces cerevisiae cell growth demands that a high degree of secretory polarity be established and directed toward the emerging bud. We report here our demonstration that mutations in SAC1, a gene identified by virtue of its allele-specific genetic interactions with yeast actin defects, were also capable of suppressing secl4 lethalities associated with yeast Golgi defects. Moreover, these sad suppressor properties also extended to sec6 and sec9 secretory vesicle defects. The genetic data are consistent with the notion that SAClp modulates both secretory pathway and actin cytoskeleton function. On this basis, we suggest that SAClp may represent one aspect of the mechanism whereby secretory and cytoskeletal activities are coordinated, so that proper spatial regulation of secretion might be achieved.

The Sac1 Phosphoinositide Phosphatase Regulates Golgi Membrane Morphology and Mitotic Spindle Organization in Mammals

“…Ablation of ySac1 PIP phosphatase function evokes pleiotropic phenotypes that include a cold sensitivity for growth, Ins auxotrophy, allele-specific genetic interactions with mutations in the single yeast actin structural gene, and bypass for the normally essential cellular requirement for Sec14, the major yeast PtdIns/phosphatidylcholine transfer protein (Cleves et al, 1989;Novick et al, 1989;Whitters et al, 1993). Mutations that compromise ySac1 enzymatic activity phenocopy sac1 null mutations, whereas substitutions outside the ySac1 catalytic motif evoke defects that are likely regulatory in nature (Nemoto et al, 2000;Li et al, 2002).…”

“…The SAC domain derives from the yeast Sac1 protein (ySac1; Cleves et al, 1989), and it represents a signature for PIP phosphatase catalytic activity . PIP phosphatases such as phosphatase and tensin homolog (mutated in multiple advanced cancers 1) (PTEN) (Maehama et al, 2001), synaptojanins (Cremona et al, 1999), and synaptojanin-like proteins (Srinivasan et al, 1997;Stolz et al, 1998) all harbor SAC domains.…”

“…First, ySac1 degrades a PtdIns-4-P pool produced by the Stt4 PtdIns 4-OH kinase, one of three PtdIns 4-OH kinases in this organism (Nemoto et al, 2000;Foti et al, 2001). Second, ySac1 is an integral membrane protein of endoplasmic reticulum (ER) and Golgi membranes (Cleves et al, 1989;Whitters et al, 1993;Nemoto et al, 2000). This disposition is determined by two C-terminal membrane anchor sequences with the large catalytic domain Abbreviations used: BrdU, bromodeoxyuridine; BSA, bovine serum albumin; ER, endoplasmic reticulum; ES, embryonic stem; FACS, fluorescence-activated cell sorting; GAG, glycosaminoglycan; GFP, green fluorescent protein; Ins, inositol; LOF, loss-of-function; MT, microtubule; PBS, phosphate-buffered saline; PIP, phosphoinositide; PITP, phosphatidylinositol transfer protein; PtdIns, phosphatidylinositol; RFP, red fluorescent protein; RT-PCR, reverse transcriptase-polymerase chain reaction; TGN, trans-Golgi network; VSV-G, vesicular stomatitis virus glycoprotein.…”

“…Spatial and temporal control of such reactions requires a finely coordinated balance between the activities of the lipid kinases that produce PIPs and the activities of enzymes that degrade them. PIP turnover is catalyzed by two general classes of enzymes: phospholipases and PIP phosphatases.Although experimental scrutiny of the phospholipases has historically been more intense, recent demonstrations of the significant biological functions played by PTEN, the myotubularins, and synaptojanins highlight the general importance of PIP phosphatases (Wishart et al, 2001;Wishart and Dixon, 2002;Wenk and De Camilli, 2004).The SAC domain derives from the yeast Sac1 protein (ySac1; Cleves et al, 1989), and it represents a signature for PIP phosphatase catalytic activity . PIP phosphatases such as phosphatase and tensin homolog (mutated in multiple advanced cancers 1) (PTEN) (Maehama et al, 2001), synaptojanins (Cremona et al, 1999), and synaptojanin-like proteins (Srinivasan et al, 1997;Stolz et al, 1998) all harbor SAC domains.…”

“…The prototypical member of this family, ySac1, catalyzes the dephosphorylation of PIPs with the intriguing exception that PtdIns-4,5-P 2 is not a substrate Rivas et al, 1999;Hughes et al, 2000). In keeping with the multiplicity of PIP signaling functions in eukaryotes, Sac1 loss-of-function (LOF) elicits pleitotropic effects in yeast that reflect broad-ranging alterations in multiple aspects of lipid metabolism (Cleves et al, 1989;Novick et al, 1989;Rivas et al, 1999;Tahirovic et al, 2005). Such metabolic alterations contribute to mislocalization of PIP binding proteins from the Golgi complex to inappropriate compartments (Li et al, 2002).…”