Control of centrosome biogenesis

Division Of Basic Sciences - Nci

Investigators

Linked publications, trials & patents

Abstract

Centrosomes are minute multifunctional membrane-less organelles present in only two copies in a typical cycling animal cell. In the core of a centrosome is a nine-fold symmetrical cylindrical microtubule-based structure called a centriole. The centriole organizes the second major part of the centrosome called pericentriolar material, which is a large and dynamic multiprotein complex and the site of most centrosomal functions. Centrosomes perform vital cellular functions such as microtubule nucleation, organization of the bipolar mitotic spindle pole during cell division, cellular signaling, and they influence tissue architecture and cell motility. Centrioles also organize sensory and motile cilia, which are critically important for development and tissue homeostasis. Supernumerary and/or structurally aberrant centrioles and centrosomes can initiate tumorigenesis, promote tumor invasiveness, and are present in almost all types of tumors. Centriolar or ciliary defects are, in addition, an underlying cause of genetic disorders known as ciliopathies. Therefore, we continue to direct our research efforts toward understanding the molecular mechanisms that regulate centriole and centrosome number, their architecture. Specifically, we elucidated the localization and the dynamics of a centrosomal protein CPAP (also known as CENPJ), in human cells. CPAP is critical for centriole and centrosome assembly, and its mutations have been found in genetic diseases such as microcephaly and Seckel syndrome. Therefore, understanding how CPAP is regulated and how it affects centrosome and cilia formation is of paramount importance. We have generated a model system in which we can inducibly replace endogenous version of CPAP protein with various disease-associated CPAP mutants and study their effects on centrosome and cilia assembly, cell division and cell proliferation. We have identified and are now characterizing CPAP mutants involved in centriole elongation, maturation, and aberrant centriole reduplication. In addition, we have characterized centriolar defects that occur under suboptimal levels of CPAP protein, which include the lack of complete centriole walls, structural instability, and breakage. We have also demonstrated that CPAP is a highly dynamic within the centriole, but that it may not directly participate in the formation of mitotic spindles and cell division, as previously suggested. In another line of research, we have explored how various cell cycle kinases (Plk1, Cdk2, Plk4), which are frequently upregulated in cancer, regulate centriole and centrosome number, and ensure centriole and centrosome homeostasis and how their dysregulation leads to centrosome amplification and reduplication. Manuscripts reporting new findings are being prepared for publication. In addition, we studied the basic principles of centriole elongation in human cells. Proper centriole structuring is a prerequisite for the subsequent proper centrosome and cilia functioning. However, strikingly little is still known about the mechanisms that regulate centriole assembly. Partially, this is because centrioles are small, sub-resolutional, structures which are difficult to study using conventional microscopy. So, in 2022, we have invested in development of super resolution protocols that would allow us to further increase imaging resolution, providing us with in-depth view of centrosomal architecture and localization of its components during centriole assembly. The current resolution that we can achieve using a combination expansion microscopy and STED is nearing 10 nm, which is unprecedented. Some of the results using this imaging approach have been published. We are currently using this approach to dissect the process of centriole elongation. Specifically, we are exploring how individual cell cycle regulators affect centriole elongation during specific parts of the cell cycle. The final goal of this study is to understand how healthy cells maintain relatively uniform centriole and centrosome size, and to identify how structurally defective centrioles and centrosomes occur in tumors.