Mechanisms of spindle formation – Mitosis

Mitosis is a key stage during the life of a cell. It is the stage where a bipolar spindle structure is organized to segregate duplicated chromosomes into the two daughter cells. Spindle organization and function require exquisite precision, robustness and fidelity. Defects associated with the spindle can lead to defects in chromosomal segregation, or aneuploidy, which has been correlated with some types of cancer. The spindle is a macromolecular machine made of microtubules, microtubule-associated proteins (MAPs), molecular motors and other regulatory proteins. Of intense interest have been molecular motors, which perform work such as cross-linking and sliding microtubules apart to form the bipolar spindle, or to depolymerize microtubules to maintain proper spindle lengths, or to carry chromosomes to opposite spindle poles. Surprisingly, while we have learned much about the motors involved in mitosis, we still know very little about the MAPs and other regulatory proteins and how they coordinate with motors to bring about proper spindle formation.
My lab uses the relatively simple fission yeast Schizosaccharomyces pombe and human cultured cells to address conserved mechanisms of spindle organization and function. This particular project focuses on how the initial bipolar spindle is formed at the start of mitosis, the stage termed prophase. We focus on the MAPs that contribute to spindle formation. Using fission yeast as a gene discovery tool, we have begun to define the roles of a new gene we called psr1+ (poles separation regulator 1). Our work indicates that psr1p organizes the initial bipolar spindle during prophase. Psr1-deletion leads to high frequency of monopolar spindles and massive subsequent chromosome segregation defects. Interestingly, fission yeast psr1+ appears to have a human functional homolog. We have begun to characterize a novel human gene we called PSR1. In HeLa cells, siRNA of PSR1 also leads to high frequency of monopolar spindles and subsequent chromosome segregation defects.
Based on strong preliminary data that fission yeast psr1p localizes to the spindle pole body (SPB) and interacts with the conserved SUN-domain protein sad1p and the microtubule +TIP EB1 protein mal3p, we propose to test the model where psr1p organizes the initial bipolar spindle at prophase. The model suggests that psr1p is anchored to the spindle pole by sad1p and captures plus ends of microtubules coming from the opposite spindle pole by binding to mal3p, thus establishing the bipolar spindle. Fission yeast and human psr1+/PSR1 are novel genes that function in organizing the initial biopolar spindle at the start of mitosis. Our model for psr1+/PSR1 represents a new conceptual framework to study spindle organization and function.
This proposal aims to combine modern cell and molecular biology techniques in fission yeast and human cultured cells, biochemistry, high-resolution optical live-cell and in vitro imaging, and innovative microfluidic techniques to control cellular microenvironment, to test the new model and reach a mechanistic understanding of bipolar spindle formation. I am an established expert in using fission yeast to study conserved mechanisms of spindle organization and function (Loiodice et al, 2005 Mol Biol Cell; Janson et al, 2007 Cell; Fu et al, 2009 Dev Cell). My collaborator Matthieu Piel is an established expert on using human cells to study mechanisms of mitosis (Fink et al, 2011 Nat Cell Biol). This places us in an excellent position to make major discoveries concerning psr1+/PSR1 and their conserved function.