Centrosome Regulation in Development and Dysregulation in Disease

National Heart, Lung, And Blood Institute

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Abstract

One of our goals is to better understand the role of centrosomes during brain development. Proper brain patterning requires stem cells to accurately produce and position cells throughout development and until the adult tissue is formed. This is also true in Drosophila, which relies on proper asymmetric division of their neural stem cells, or neuroblasts (NBs) to produce the adult fly brain. Previous work on centrosome NBs uncovered a specialized centrosome maturation cycle important for invariant NB division. Briefly, the two centrosomes are asymmetric in interphase: the apical centrosome is mature and anchored to the cortex while the basal centrosome is inactivated and becomes mobile. Importantly, the active apical centrosome is always inherited by the new NB, while the basal centrosome is inherited by the differentiated cell. During this last funding period (1 year), we investigated the mechanism by which this asymmetric centrosome behavior is achieved and its importance for proper brain development. To begin our studies, we sought to identify all the molecular differences between the apical and basal centrosomes We generated antibodies and GFP transgenic animals that allowed us to screen a panel of candidate proteins that might be asymmetrically enriched on one of the centrosomes. Our analysis identified PLP as the only protein enriched on the basal centrosome, which lead to the hypothesis that PLP might serve as a negative regulator of basal centrosome activity. Mutations in Pericentrin (the human ortholog of PLP) cause primordial dwarfism type II; therefore, detailed understanding of its function and regulation is clinically relevant. We found that loss of PLP leads to precocious activation of the basal centrosome, defects in centrosome number inheritance, and defects in mitotic spindle formation. This work supports a role for PLP as a negative regulator of PCM recruitment in interphase, which is distinct from its known positive regulatory role of PCM recruitment in mitosis. We went on to show that PLP performs this function by preventing the critical maturation kinase Polo from localizing to the basal centrosome during interphase. We favor a model where PLP acts on the centrosome by physically shielding the docking of promaturation factors until mitotic entry. A biochemical modification and/or conformational change would then allow PLP to transition to a positive regulator in mitosis that scaffolds other PCM factors. Another possible direct function for PLP might be to differentially modulate the behavior of proteins at the apical and basal centrosomes. It will be critical to examine how the dynamics of other centrosome proteins are affected by the loss of PLP. Given the importance of interphase centrosome asymmetry in stem cell division, we believe that an in depth understanding of the interplay between PCM proteins is a critical area of research. Our current work aims to determine what functions upstream and downstream of PLP in this regulatory pathway.