Mechanisms underlying centriole morphogenesis

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PROJECT SUMMARY/ ABSTRACT The heart of the centrosome, the microtubule organizing center, is composed of two centrioles. The two centrioles are not equal. The older of the two, called the mother centriole, differs structurally from the younger, daughter centriole. Mother centriole-specific structures confer the unique capability to nucleate the primary cilium, an organelle that serves as the cell’s antenna. Consequently, defects in centriolar proteins can cause human ciliopathies, diseases caused by disrupted ciliary function. Despite being universal features of vertebrate cells, how the mother and daughter centrioles differ and how centrioles are built remain mysterious. I uncovered a complex of proteins comprised of CEP90, MNR and OFD1 (which I have named DISCO for DIStal Centriole cOmplex) required for proper centriole morphogenesis. Mutations in DISCO components cause Joubert and Orofaciodigital syndromes, disorders of brain, face and limb development. By studying this novel centriolar complex, I seek to understand how centrioles are built, and how they are remodeled to support cilium assembly. Using an innovative combination of expansion and structured illumination microscopy (Ex-SIM), I will define how components of this complex structure the distal centriole and how human disease-associated mutations disrupt this sub-compartment (Aim 1). MNR and OFD1 control centriole length by an unknown mechanism. Using super-resolved imaging and biochemical assays, I will uncover molecular mechanisms by which centriole length is established by MNR and OFD1 (Aim 2). CEP90 and MNR are also components of centriolar satellites, poorly understood membrane-less granules surrounding the centrosome. I have found that centriolar satellites display hallmarks of phase separation. Using live-imaging and in vitro biochemical reconstitution, I will test the hypothesis that CEP90, MNR and OFD1 are trafficked to the centriole by phase- separated centriolar satellites to support ciliogenesis (Aim 3). With the help of an outstanding advisory committee, I will train in advanced imaging and biophysical techniques that will allow me to address fundamental questions on how centrioles and cilia are built. Spanning both the mentored and independent phases, these studies will illuminate how human disease-associated proteins build and modify centrioles to allow cilium biogenesis, and create a strong foundation for an independent research career studying the role of centrioles and cilia in human development and disease.