STIL Microcephaly Mutations Interfere with APC/C-Mediated Degradation and Cause Centriole Amplification

Abstract

Our results reveal a role for Cdk1 in STIL dissociation from centrosomes during early mitosis, with implications for the timing of cartwheel disassembly. Additionally, we propose that centriole amplification triggered by STIL stabilization is the underlying cause of microcephaly in human patients with corresponding STIL mutations.

Quantitative analysis of human centrosome architecture by targeted proteomics and fluorescence imaging

“…Similarly, the number of Sas‐6 molecules at centrosomes rose from ~346 in G1/S arrested cells to 440 in G2/M cells (Fig 5B) and those of STIL from 167 to 318 (Fig 5C). Neither Sas‐6‐EGFP nor STIL‐EGFP could be detected at centrosomes in mitotic cells (Fig 5B and C), consistent with ubiquitin‐dependent proteolytic degradation of both proteins (Strnad et al , 2007; Tang et al , 2011a; Arquint et al , 2012; Vulprecht et al , 2012; Arquint & Nigg, 2014). …”

“…However, these numbers represent an underestimate, because ~50% of centrosomes purified from asynchronously growing KE37 cells are derived from G1‐phase cells. In contrast to most other proteins analyzed here, including the protein used for calibration (γ‐tubulin), both Sas‐6 and STIL show marked cycle regulation and very similar cell cycle profiles; as a consequence, they are barely detectable at most G1‐phase centrosomes (Strnad et al , 2007; Arquint & Nigg, 2014; Keller et al , 2014). Thus, an approximate doubling of the above numbers likely provides a better estimate of the absolute abundance of Sas‐6 and STIL on those centrosomes (S and G2 phases) that are positive for these proteins, resulting in 170 and 72 molecules for Sas‐6 and STIL, respectively.…”

The Centrosome and Its Duplication Cycle

“…Similar reductions occur for CPAP, STIL, and human Sas-6 and each of these molecules is targeted by APC/C-Cdh1 or Cdc20 (Strnad et al 2007;Tang et al 2009Tang et al , 2011Puklowski et al 2011;Arquint et al 2012;Arquint and Nigg 2014). Interplay between APC/C and SCF pathways has also been reported to regulate levels of Sas-6 (Puklowski et al 2011).…”

“…Other genes for centrosomal proteins that are mutated in microcephaly include the CNN homolog, CDK5RAP2 (MCPH3), required for centrosome maturation and DNA damage-induced G 2 arrest (Barr et al 2010;Lizarraga et al 2010), and CEP63 with its partner Cep152, which participate in centriole assembly (Sir et al 2011). Centriole amplification has also been shown to be one cause of microcephaly in human patients that is triggered by mutants that stabilize STIL by removing a KEN destruction box (Arquint and Nigg 2014).…”

Small organelle, big responsibility: the role of centrosomes in development and disease

“…A missense MCPH mutation weakens the ability of CPAP to bind STIL and impairs centriole production [32][33][34][35]. By contrast, MCPH-causing mutations in STIL do not interfere with centriole formation, but prevent cell cycledependent degradation of the protein, thereby triggering centriole amplification [36]. Thus, MCPH mutations seem to target essential components of the centriole biogenesis pathway.…”