Protein phosphatase 5 is a negative regulator of separase function during cortical granule exocytosis inC. elegans

Abstract

SummaryMutations in the Caenorhabditis elegans separase gene, sep-1, are embryonic lethal. Newly fertilized mutant embryos have defects in polar body extrusion, fail to undergo cortical granule exocytosis, and subsequently fail to complete cytokinesis. Chromosome nondisjunction during the meiotic divisions is readily apparent after depletion of sep-1 by RNAi treatment, but much less so in hypomorphic mutant embryos. To identify factors that influence the activity of separase in cortical granule exocytosis and cytokinesis, we carried out a genetic suppressor screen. A mutation in the protein phosphatase 5 (pph-5) gene was identified as an extragenic suppressor of sep-1. This mutation suppressed the phenotypes of hypomorphic separase mutants but not RNAi depleted animals. Depletion of pph-5 caused no phenotypes on its own, but was effective in restoring localization of mutant separase to vesicles and suppressing cortical granule exocytosis and cytokinesis phenotypes. The identification of PPH-5 as a suppressor of separase suggests that a new phospho-regulatory pathway plays an important role in regulating anaphase functions of separase.

Rab6 is required for the exocytosis of cortical granules and the recruitment of separase to the granules during the oocyte-to-embryo transition in Caenorhabditis elegans

“…However, no experimental support is currently available for the hypothesis that RAB-6.1 must be phosphorylated for its localization to the cortical granules. Interestingly, however, the depletion of protein phosphatase 5 (PPH-5) suppresses the localization defect for two of the three sep-1 alleles that have a specific defect in SEP-1 localization to the cortical granules (Bembenek et al, 2007;Richie et al, 2011). It has been proposed that phosphorylation is involved in the regulation of SEP-1 localization, and that SEP-1 is a plausible target of the phosphorylation (Richie et al, 2011).…”

“…Interestingly, however, the depletion of protein phosphatase 5 (PPH-5) suppresses the localization defect for two of the three sep-1 alleles that have a specific defect in SEP-1 localization to the cortical granules (Bembenek et al, 2007;Richie et al, 2011). It has been proposed that phosphorylation is involved in the regulation of SEP-1 localization, and that SEP-1 is a plausible target of the phosphorylation (Richie et al, 2011). Alternatively, RAB-6.1 may be a target for phosphoregulation, and PPH-5 depletion may lead to increased levels of the GTPbound form of RAB-6.1 and of the RAB-6.1 protein on cortical granules in order to suppress some of the sep-1 alleles.…”

“…CDK-1, APC/ C, and SEP-1). Further, the localization of SEP-1 on cortical granules is likely critical for exocytosis, because sep-1 alleles defective for cortical granule exocytosis are defective for SEP-1 localization on cortical granules, and a suppressor for the exocytosis defect also rescues the SEP-1 localization (Richie et al, 2011). However, the molecule that recruits separase to cortical granules remains unidentified.…”

Biology and insights into the role of cohesin protease separase in human malignancies

“…Knockdown of separase using RNAi inhibits degranulation in addition to causing abnormal chromosomal segregation (Bembenek et al, 2007). Several mutations of separase with little effect on chromosomal segregation reduce its localization to cortical granules (Bembenek et al, 2007;Richie et al, 2011), suggesting that the functions of separase in membrane trafficking and chromosomal segregation are independent.…”

“…The C-terminus of separase, which contains the proteolytic active site, is conserved from yeast to humans. In addition to its canonical role in sister-chromatid separation, separase is involved in other functions, such as centrosome duplication (Nakamura, Arai & Fujita, 2009;Tsou et al, 2009;Schockel et al, 2011;Lee & Rhee, 2012;Matsuo et al, 2012), DNA damage repair (Nagao, Adachi & Yanagida, 2004;McAleenan et al, 2013), and membrane trafficking (Bembenek et al, 2007;Richie et al, 2011), etc. Because separase is involved in numerous important cell processes, its activity is tightly regulated (Ciosk et al, 1998;Zou et al, 1999;Stemmann et al, 2001;Gorr, Boos & Stemmann, 2005;Holland & Taylor, 2006).…”