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  • br Transparency document br Main Text The

    2019-10-09


    Transparency document
    Main Text The centrosome is a non-membrane-bound organelle found in most animal cells. It has several important functions, including control of cilia formation, microtubule organization and nucleation, spindle assembly and transport of organelles and vesicles. Centrosomes consist of two centrioles, which are barrel-shaped microtubule-based structures surrounded by an electron-dense matrix, the pericentriolar material (PCM). Duplication of centrioles must occur in coordination with DNA synthesis only once per cell cycle and duplication involves the assembly of a new daughter centriole (procentriole) next to the proximal end of the mother centriole [1]. In human cells, hSAS-6, STIL, CEP135, CPAP, γ-tubulin, and CP110 have been identified as essential factors required at the onset of centriole biogenesis for centriole formation 2, 3. The newly formed procentrioles then start to elongate, increase in length during S phase, and are fully assembled in G2 phase of the cell cycle. A new study by Li et al.[4] now reports a role for deubiquitination in the stabilization of CP110 during S/G2 phase. The protein CP110 was first identified as a Cdk2 substrate required for centrosome overduplication in S-phase-arrested cells [5]. It is associated with the growing centriolar distal tips, forming a cap beneath which centrioles elongate through the insertion of α/β-tubulin [2]. Depletion of CP110 or overexpression of CPAP was shown to induce centriole elongation, suggesting that CPAP and CP110 play opposing roles in controlling centriole length 6, 7, 8. CP110 in a complex with its regulator Cep97 also plays an additional role in preventing cilia formation through the inhibition of centriole to LiCl mass conversion [9]. CP110 protein levels are tightly controlled during the cell cycle in order to prevent centriole duplication errors. CP110 physically associates with the F-box protein cyclin F on centrioles during the G2 phase of the cell cycle and is then ubiquitinated by the SCFCyclinF E3 ubiquitin ligase complex, leading to its degradation [10]. The new study by Li et al.[4] now reports that deubiquitination is also an important mechanism for the regulation of CP110 protein levels during the cell cycle. Polyubiquitinated proteins that are marked for degradation can be stabilized in response to the activity of deubiquitinating enzymes (DUBs) — proteases that remove the polyubiquitin chain. Around 100 active DUBs are found in the human genome [11] and, as a result of their ability to reverse ubiquitination, these enzymes control a broad range of key cellular processes. Li et al.[4] report that the DUB USP33 (ubiquitin-specific protease 33, also known as VDU1) binds to CP110 and specifically deubiquitinates CP110 in a cell-cycle-dependent manner, thus counteracting SCFCyclinF ubiquitin ligase activity (Figure 1). Although several SCF ubiquitin ligase complexes have been recently shown to regulate centrosome biogenesis [12], DUBs had not been demonstrated to be involved in the regulation of this process until now. Both USP33 and the highly related USP20 (also known as VDU2) interact with CP110, but USP33 has a greater impact on CP110 levels than USP20. USP33 and USP20 are mainly localized to the cytoplasm, in particular to the endoplasmic reticulum [13]. USP33 additionally localizes to the proximal end of centrioles, whereas CP110 is a distal-end-capping protein, suggesting that only a subpopulation of CP110 interacts with USP33. USP33 localizes to centrosomes primarily during S and G2 phase of the cell cycle similar to CP110. Centriolar targeting of USP33 is at least in part dependent on CP110 because depletion of CP110 reduced USP33 localization to the centrosome. USP33 specifically deubiquitinates CP110 both in vitro and in vivo. The amino-terminal domain of CP110 interacts with the catalytic domain of USP33 and is required for USP33-mediated deubiquitination.