PP2A Phosphatase Acts upon SAS-5 to Ensure Centriole Formation in C. elegans Embryos

Abstract

Centrosome duplication occurs once per cell cycle and ensures that the two resulting centrosomes assemble a bipolar mitotic spindle. Centriole formation is fundamental for centrosome duplication. In Caenorhabditis elegans, the evolutionarily conserved proteins SPD-2, ZYG-1, SAS-6, SAS-5, and SAS-4 are essential for centriole formation, but how they function is not fully understood. Here, we demonstrate that Protein Phosphatase 2A (PP2A) is also critical for centriole formation in C. elegans embryos. We find that PP2A subunits genetically and physically interact with the SAS-5/SAS-6 complex. Furthermore, we show that PP2A-mediated dephosphorylation promotes centriolar targeting of SAS-5 and ensures SAS-6 delivery to the site of centriole assembly. We find that PP2A is similarly needed for the presence of HsSAS-6 at centrioles and for centriole formation in human cells. These findings lead us to propose that PP2A-mediated loading of SAS-6 proteins is critical at the onset of centriole formation.

The Centrosome and Its Duplication Cycle

“…The close cooperation between SAS-5 and SAS-6 in centriole duplication in C. elegans is also seen with the human and Drosophila counterparts of SAS-5, STIL, and Ana2, respectively (Stevens et al 2010;Kitagawa et al 2011a;Tang et al 2011;Arquint et al 2012;Vulprecht et al 2012). Overexpression of SAS-6 in Drosophila syncytial embryos led to the de novo formation of tube-or vesicle-like structures that are surrounded by microtubule asters (Rodrigues- …”

“…However, coexpression of Sas-6 and Ana2 in Drosophila spermatocytes leads to the assembly of cartwheel-like structures (Stevens et al 2010). Overexpression of STIL or Sas-6 in other systems leads to centriole overduplication (Kitagawa et al 2011a;Tang et al 2011;Arquint et al 2012;Vulprecht et al 2012). Both Ana2 and STIL localize to the innermost region of the centriole Dzhindzhev et al 2014), raising the possibility that Ana2/STIL might be part of the cartwheel structure.…”

“…In C. elegans, PP2A's association with the SAS-5 -SAS-6 complex is instrumental in targeting SAS-5 and, hence, SAS-6 to the centriole (Kitagawa et al 2011a), and its activity protects SAS-5 as well as ZYG-1 from degradation by the proteasome (Song et al 2011). PP2A is similarly required for human Sas-6 to localize to centrioles (Kitagawa et al 2011a), although it is not clear whether this is mediated through STIL.…”

Analysis of centriole elimination duringC. elegansoogenesis

“…Overall, we conclude that SPD-2, ZYG-1, SAS-6 and SAS-4 are required for centrosome duplication in proliferating GCs, as anticipated from the sterility associated with spd-2(oj29) and zyg-1(it25) (O'Connell et al, 1998;Kemp et al, 2007), as well as in sas-6(ok2554) and sas-4(tm3951) (Kitagawa et al, 2011).…”

“…The following genotypes were used: sas-6(ok2554) (Kitagawa et al, 2011), zyg-1(it25) (Kemp et al, 2007), spd-2(oj29) (O'Connell et al, 1998), sas-5(t2033) , sas-5(gk400) (http://www.wormbase.org), sas-4(tm3951) (Kitagawa et al, 2011), cki-2(ok2105) (Buck et al, 2009), fog-1(q253) (Barton and Kimble, 1990) (shifted to the restrictive temperature of 25°C as L1/L2), fog-2(q71) (Schedl and Kimble, 1988), fog-3(q470) (Ellis and Kimble, 1994), tra-1(e1076) (Schedl et al, 1989), her-1(hv1y101) unc-42 and her-1(hv1y101) unc-42 dpy-11;lon-2 (Francis et al, 1995b), analyzed as described (Bean et al, 2004), mog-1(q233) (Graham and Kimble, 1993), fem-3(q96gf) (Barton et al, 1987), cgh-1(ok492) (Audhya et al, 2005), glo-1(zu391) (Hermann et al, 2005), rrf-1(pk1417) (Sijen et al, 2001), GFP-SAS-4 (Kirkham et al, 2003), GFP-SAS-5 , GFP-SAS-6 , GFP-SPD-2 (Kemp et al, 2004), GFP-TBG-1 (Oegema et al, 2001), GFP -TAC-1 (Bellanger and Gönczy, 2003), VIT-2-GFP (Grant and Hirsh, 1999), NMY-2-GFP (Nance et al, 2003).…”