4 research outputs found
On the regulation of centriole duplication in human cells : exploring the interactions of polo-like kinase 4 with the centrosomal proteins Cep192 and STIL
No full textCentrioles duplicate once in each cell cycle to give rise to two centrosomes that form the spindle poles during mitosis. Aberrant centriole duplication can result in the formation of supernumerary centrosomes, leading to incorrect spindle assembly and chromosome segregation errors, thereby possibly contributing to carcinogenesis (Ganem et al., 2009; Nigg, 2002; Zyss and Gergely, 2008). Thus, to ensure genome stability, centriole duplication has to be precisely regulated. Polo-like kinase 4 (PLK4) is a key regulator of centriole duplication (Bettencourt-Dias et al., 2005; Habedanck et al., 2005). PLK4 is characterized by an N-terminal Ser/Thr kinase domain and three C-terminal Polo-boxes (PB1-PB3) (Slevin et al., 2012). The PB1-PB2 domain is required for PLK4's centrosomal localization and binding to Cep152 (Cizmecioglu et al., 2010; Hatch et al., 2010; Slevin et al., 2012). In contrast to PB1-PB2, no binding partners have been described for PB3.
Here, we identify Cep192 and STIL as novel interaction partners of PLK4-PB1-PB2 and PLK4-PB3, respectively. In the first part of this study, we reveal that Cep192 directly binds PB1-PB2 via a short region within its N-terminus, which contains conserved patches of acidic residues. We show that also in the case of Cep152 a short N-terminal acidic region is critical for the binding to PB1-PB2. These acidic regions of Cep192 and Cep152 enable electrostatic interactions with positively charged residues of the PB1-PB2 domain in order to promote PLK4 centriolar recruitment (Sonnen et al., 2013). In the second part of this study, we identify STIL as the first known binding partner of PLK4-PB3. We show that the coiled-coil motif of STIL (STIL-CC) is necessary and sufficient for this interaction and thus important for centriole duplication. Based on a collaboration for crystallographic and NMR analyses, we furthermore demonstrate that PB3 adopts a canonical PB fold, and that the PLK4-PB3/STIL-CC binding mimics coiled-coil formation. Analysis of structure-guided STIL mutants suggests a dual binding mode of STIL-CC to PB3 and L1 of PLK4 (linker between the catalytic domain and the PB domains). Taken together, we propose a speculative model for the initial steps of procentriole assembly according to which PLK4 is recruited to centrioles by electrostatic interactions between PB1-PB2 and Cep192/Cep152, and thereafter is stabilized and activated via STIL-CC binding to PB3 and L1
STIL binding to Polo-box 3 of PLK4 regulates centriole duplication
No full textPolo-like kinases (PLK) are eukaryotic regulators of cell cycle progression, mitosis and cytokinesis; PLK4 is a master regulator of centriole duplication. Here, we demonstrate that the SCL/TAL1 interrupting locus (STIL) protein interacts via its coiled-coil region (STIL-CC) with PLK4 in vivo. STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region. Structure determination of free PLK4-PB3 and its STIL-CC complex via NMR and crystallography reveals a novel mode of Polo-box-peptide interaction mimicking coiled-coil formation. In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding. We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication
Centrosomes as signalling centres
No full textCentrosomes-as well as the related spindle pole bodies (SPBs) of yeast-have been extensively studied from the perspective of their microtubule-organizing roles. Moreover, the biogenesis and duplication of these organelles have been the subject of much attention, and the importance of centrosomes and the centriole-ciliary apparatus for human disease is well recognized. Much less developed is our understanding of another facet of centrosomes and SPBs, namely their possible role as signalling centres. Yet, many signalling components, including kinases and phosphatases, have been associated with centrosomes and spindle poles, giving rise to the hypothesis that these organelles might serve as hubs for the integration and coordination of signalling pathways. In this review, we discuss a number of selected studies that bear on this notion. We cover different processes (cell cycle control, development, DNA damage response) and organisms (yeast, invertebrates and vertebrates), but have made no attempt to be comprehensive. This field is still young and although the concept of centrosomes and SPBs as signalling centres is attractive, it remains primarily a concept-in need of further scrutiny. We hope that this review will stimulate thought and experimentation
Human Cep192 and Cep152 cooperate in Plk4 recruitment and centriole duplication
No full textPolo-like kinase 4 (Plk4) is a key regulator of centriole duplication, but the mechanism underlying its recruitment to mammalian centrioles is not understood. In flies, Plk4 recruitment depends on Asterless, whereas nematodes rely on a distinct protein, Spd-2. Here, we have explored the roles of two homologous mammalian proteins, Cep152 and Cep192, respectively, in the centriole recruitment of human Plk4. We demonstrate that Cep192 plays a key role in centrosome recruitment of both Cep152 and Plk4. Double-depletion of Cep192 and Cep152 completely abolishes Plk4 binding to centrioles as well as centriole duplication, indicating that the two proteins cooperate. Most importantly, we show that Cep192 binds Plk4 through an N-terminal extension that is specific to the largest isoform. The Plk4 binding regions of Cep192 and Cep152 (residues 190-240 and 1-46, respectively) are rich in negatively charged amino acids, suggesting that Plk4 localization to centrioles depends on electrostatic interactions with the positively charged polo-box domain. We conclude that cooperation between Cep192 and Cep152 is crucial for centriole recruitment of Plk4 and centriole duplication during the cell cycle
