Plk4 Phosphorylates Ana2 to Trigger Sas6 Recruitment and Procentriole Formation

Abstract

SummaryCentrioles are 9-fold symmetrical structures at the core of centrosomes and base of cilia whose dysfunction has been linked to a wide range of inherited diseases and cancer [1]. Their duplication is regulated by a protein kinase of conserved structure, the C. elegans ZYG-1 or its Polo-like kinase 4 (Plk4) counterpart in other organisms [2–4]. Although Plk4’s centriolar partners and mechanisms that regulate its stability are known, its crucial substrates for centriole duplication have never been identified. Here we show that Drosophila Plk4 phosphorylates four conserved serines in the STAN motif of the core centriole protein Ana2 to enable it to bind and recruit its Sas6 partner. Ana2 and Sas6 normally load onto both mother and daughter centrioles immediately after their disengagement toward the end of mitosis to seed procentriole formation. Nonphosphorylatable Ana2 still localizes to the centriole but can no longer recruit Sas6 and centriole duplication fails. Thus, following centriole disengagement, recruitment of Ana2 and its phosphorylation by Plk4 are the earliest known events in centriole duplication to recruit Sas6 and thereby establish the architecture of the new procentriole engaged with its parent.

The Centrosome and Its Duplication Cycle

“…It has been suggested that recruitment of Asl onto the daughter centriole in Drosophila embryos occurs only once mother and daughter have separated at the end of mitosis and that this provides a duplication license (Novak et al 2014). However, it is notable that in cultured cells, the incorporation of Ana2 and Sas-6 onto the daughter's procentriole once it has disengaged from the mother in telophase appears to mark the very first event in the duplication process (Dzhindzhev et al 2014).…”

“…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. Indeed, the C. elegans SAS-5 physically binds a narrow central region of the SAS-6 coiled-coil domain, and is able to prevent the coiled-coil dimer from forming a tetramer in vitro (Qiao et al 2012).…”

“…Both Ana2 and Sas-6 become recruited to the daughter centriole once it has disengaged from the mother at the end of mitosis. If the Plk4 sites in Ana2 are mutated to nonphosphorylatable residues, it can still bind to the daughter centriole but it cannot recruit Sas-6, and centriole duplication fails (Dzhindzhev et al 2014). In contrast to Drosophila, where some Sas-6 remains at the core of the centriole once it is incorporated, in human cells, Sas-6 is destroyed during G 1 (Strnad et al 2007) and is transiently recruited to the lumen of the mother centriole in S phase (Fong et al 2014).…”

“…Indeed, the C. elegans SAS-5 physically binds a narrow central region of the SAS-6 coiled-coil domain, and is able to prevent the coiled-coil dimer from forming a tetramer in vitro (Qiao et al 2012). Moreover, once phosphorylated by Plk4, Drosophila Ana2 becomes able to bind Sas-6 (see below) (Dzhindzhev et al 2014). Although SAS-6 and SAS-5/Ana2/STIL are components of the innermost part of the centriole, SAS-4 colocalizes with its microtubule wall (zone II in Drosophila cells; Fu and Glover 2012).…”

“…The protein at the innermost core of the centriole is Sas-6 (zone I in Drosophila; Sonnen et al 2012;Dzhindzhev et al 2014). Indications for its importance in establishing ninefold symmetry first came from studies of loss of its function in Chlamydomonas, Paramecium, and from Drosophila, in which loss of, or abnormalities in, the cartwheel structure were observed Rodrigues-Martins et al 2007a;Jerka-Dziadosz et al 2010).…”