PROJECT SUMMARY/ABSTRACT
There is a fundamental gap in our knowledge to explain the intrinsic ability of centrioles to duplicate and
function as microtubule-organizing centers. To fill this gap, we require a thorough molecular
characterization of the very first steps in centriole assembly and pericentriolar material (PCM)
recruitment; this deficit impedes our understanding of the etiology of centrosome-linked diseases. During
mitosis, centriole function is activated and their duplication initiated by of the kinases, Polo and Polo-like
kinase 4 (Plk4), respectively. Normally, upon mitotic entry, cells contain two centrosomes, each
containing a mother-daughter centriole pair that undergoes Polo-dependent `maturation', the process of
recruiting additional PCM, allowing them to facilitate spindle assembly. As cells exit mitosis, each
daughter cell inherits a centriole pair which has been modified by Plk4 to duplicate. Thus, Polo kinases
control both the formation and duplication of functional centrosomes, and consequently, alterations in
Polo kinase activity can dramatically influence spindle assembly and proper chromosome segregation.
Our long-term goal is to better understand how cells couple cell cycle progression with centrosome
function and duplication to ensure accurate distribution of the genome during division. The objective of
this application is to understand how Polo and Plk4 are activated on mitotic centrioles to promote PCM
recruitment and centriole duplication. Drawn from our preliminary data, our central hypothesis is that
maturation and duplication are intrinsic behaviors of centrioles because Sas4, a centriole-surface protein,
controls the activities of both master-regulators, Polo and Plk4. The rationale for the proposed research
is that understanding the evolutionarily conserved fundamental mechanisms of centriole biology has the
potential to translate into therapeutic strategies to address centriole-linked human disease. This
hypothesis will be tested in three specific aims: 1) Examine a mechanism of centrosome maturation
whereby Sas4 activates Polo kinase to generate a mitotic platform for PCM recruitment; 2) Determine if
Sas4 regulates Plk4 stability, and whether Sas4 phosphorylation by Plk4 controls the Plk4/Sas4
centriolar pattern to restrict daughter centriole assembly to a single site; and 3) Determine if
phosphorylation of Asl and Sas4 by Plk4 induces formation of a daughter centriole initiation complex to
recruit Ana2. The approach is innovative, in the applicant's opinion, because it represents a new and
substantive departure of the status quo by shifting focus to the role of the centriole surface protein, Sas4,
as a key activator and substrate for both Polo and Plk4. The proposed research is significant because a
thorough understanding of centriole biogenesis and function is intimately linked to our success in
preventing/treating centriole-associated diseases (including ciliopathy, birth defects, neurodevelopmental
disorders, and cancer).