Centrosomes, cilia and centriolar satellites [electronic resource] : characterizing the role of Cep72 in centriolar satellite function

Centrosomes and cilia are evolutionarily conserved organelles with important functions in cell signaling, organismal development and disease. Defects in centrosome and cilium function are associated with a set of phenotypically-related syndromes called ciliopathies. To discover new centrosome/cilium components, candidate genes were identified by virtue of their transcriptional upregulation in multiciliated tracheal epithelial cells and selected for further study based on their subcellular localization. Using this approach, Cep72 was identified as a component of centriolar satellites - poorly understood centrosome-associated structures defined by the protein PCM1. A subset of ciliopathy-associated proteins that includes BBS4, Cep290 and OFD1 associate with centriolar satellites, yet how this association affects their function is unknown. Cep72 was determined to be a PCM1-interacting protein and its association with centriolar satellites was characterized in detail. Cep72 was found to interact with Cep290 and overexpression/depletion experiments demonstrated that Cep72 facilitates recruitment of Cep290, but not BBS4, to centriolar satellites. Loss of satellites by depletion of PCM1 resulted in cytoplasmic disperal of BBS4 and redistribution of Cep72 and Cep290 from centriolar satellites to the centrosome. In contrast to the reported function of centriolar satellites in facilitating the localization of proteins to the centrosome, the effects of PCM1 depletion on Cep72 and Cep290 localization suggest that association with centriolar satellites can also restrict centrosomal localization for some proteins. During ciliogenesis, BBS4 transitions from centriolar satellites to the primary cilium, however regulation of this transition is not understood. Depletion of Cep72 or Cep290 prevented transition of BBS4 localization, causing BBS4 to remain localized to centriolar satellites in ciliated cells. Depletion of PCM1 or Cep72 in developing zebrafish embryos resulted in phenotypes consistent with centrosome/cilium defects, suggesting centriolar satellites influence cilium function during development. Given these results, we propose that centriolar satellites function in modulating the ciliary localization of BBS4 and possibly other ciliogenesis factors, via a mechanism that involves Cep290 as an effector of that regulation. The distribution of centriolar satellites around the centrosome during interphase requires microtubules and dynein. During the cellular stress response to misfolded proteins, protein aggregates are transported in a microtubule and dynein-dependent manner to structures surrounding the centrosome referred to as aggresomes. We observed PCM1-dependent recruitment of centriolar satellite proteins to aggresomes, which prompted further investigation of the potential involvement of PCM1 in aggresome formation. Depletion of PCM1 had modest effects on aggresome formation and cell viability in response to misfolded proteins. However the influence of PCM1 on centrosome function and microtubule organization could not be ruled out as possible explanation for these effects.

Advisor: Stearns Tim primary advisor. Thesis advisor
Kopito Ron Rieger advisor. Thesis advisor
Nachury Maxence
Nelson W. J. (W. James) advisor. Thesis advisor