Examining Intracellular Phosphorylation Gradients During Cell Division

<p>The dynamic cellular reorganization needed for successful mitosis requires spatial regulatory cues. I examine this problem at two different levels. First, I analyze a phosphorylation gradient for substrates of the chromosomal passenger complex (CPC). CPC is a conserved regulator involved in key mitotic events such as chromosome-microtubule attachment and spindle midzone formation. Previously, spatial phosphorylation gradients have been reported for CPC substrates, raising the possibility that CPC-dependent signaling establishes order on the micron-length scale in dividing cells. However, this hypothesis has not been tested, largely because of incomplete characterization of the CPC-dependent phosphorylation dynamics. Here I examine the spatiotemporal dynamics of CPC-dependent phosphorylation along microtubules throughout mitosis using a Forster resonance energy transfer-based sensor. I find that a CFC-substrate phosphorylation gradient, with highest phosphorylation levels between the two spindle poles, emerges when a cell enters mitosis. After anaphase onset, the gradient emerges and persists until cell cleavage. Selective mislocalization of the CPC during anaphase suppresses gradient formation, but overall substrate phosphorylation levels remain unchanged. Under these conditions, the spindle midzone fails to organize and function properly. My findings suggest a model in which the CPC establishes phosphorylation gradients to coordinate the spatiotemporal dynamics needed for error-free cell division. Second, I examine the contribution of a microtubule crosslinking protein PRC1 to microtubule organization during cytokinesis. I find that PRC1 depletion leads to abnormal elongation of anaphase spindle, which depends on mic rotubules. I also find that the dynamics of growing microtubule plus-ends imaged by EBl-GFP is not significantly altered. Based on these findings, I propose a model for how PRC1 contributes to the length control of the anaphase spindle.</p>

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Thesis

Year

2012

Author dc:creator

• Tan, Lei

Contributors dc:contributor

• Tarun Kapoor

• mitosis
• chromosomal passenger complex
• phosphorylation
• PRCI protein
• anaphase spindle
• cytokinesis
• Life Sciences

Repository record dc:identifier

https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/250

OAI identifier oai:identifier

oai:digitalcommons.rockefeller.edu:student_theses_and_dissertations-1252