Deconstructing Cell Architecture: Exploring centriole structure, function, and position in the green alga Chlamydomonas reinhardtii

An enduring question in cell biology is how internal cell geometry is established and maintained through endless rounds of cell division. In many cell types, organelles and sub-cellular networks are partitioned into fine spatial arrangements. Precise cell geometry often plays a functional role in signaling, growth, division, and in establishing cell polarity. One sub-cellular structure that is precisely localized is the centriole. Centrioles are non-membrane-bound organelles composed of nine triplet microtubule blades arranged around a central cartwheel structure. Centrioles are found as a pair, comprised of a mother and a daughter, which is duplicated during each cell cycle. Mother centrioles are so-called because they were assembled in a previous cell cycle to the daughter centriole. Centrioles have two main functions in the cell. First, centrioles together with pericentriolar material comprise the centrosome, the major microtubule-organizing center of the cell. Second, centrioles serve as basal bodies to nucleate the assembly of cilia. Although originally named for their centralized location, centrioles are repositioned to more peripheral sites during cell-state transitions such as wound healing, cell migration, and cell growth. Despite the clear importance of centriole positioning to the cell and the organism, almost nothing is known about this process. Chlamydomonas reinhardtii is a unicellular green alga that has served as an ideal model organism in which to study centriole and cilia biology. Chlamydomonas cells have centrioles that are structurally and molecularly similar to those of vertebrates. Chlamydomonas cells also have robust cell geometry, facilitating the measurement of the positions of intracellular structures. Here, I use Chlamydomonas to understand centriole positioning. Specifically, I developed a novel screen to identify mutants with defects in centriole positioning. From this screen, I identified a number of interesting mutants with defects in cilia and centriole assembly, number, and position. I use these mutants to demonstrate that the mother centriole plays an instructive role in defining cell geometry, directing the positioning of many sub-cellular structures including the daughter centriole and the nucleus. I also use these mutants to understand spindle position, implicating the centrioles as essential for properly orienting the mitotic spindle. Additionally, I demonstrate that one of these mutants, asq2, carries a mutation in the conserved gene TBCCd1, which may regulate template-driven centriole assembly