In our first renewal application (funded from May 1,1998-April 30, 2002), we proposed to examine the role of Wnt signaling during endoderm induction in C. elegans. We have since shown that a conserved MAP Kinase module converges with Wnt signaling to polarize endoderm potential within a single embryonic cell called EMS. We also showed that Wnt signaling induces a rotation of the mitotic spindle in EMS, such that its posterior daughter inherits endoderm potential. Our findings provide important mechanistic insights about a key signal transduction pathway that is widely conserved, being involved in several important human cancers and in many different developmental processes. Four other cell divisions in the early embryo are intrinsically asymmetric. In each case, as in EMS, the mitotic spindle rotates to align with the axis of polarization. We have identified new mutants with mitotic spindle positioning defects in two intrinsically polarized embryonic cells called P0 and P1. We propose new specific aims that focus largely on intrinsic polarity in these two cells. We are investigating a gene we discovered called spn-4, which encodes a conserved RNA binding protein that acts downstream of the PAR polarity proteins to regulate mitotic spindle orientation and cell polarity in P1. We also proposed to study another mutant we recently identified, called spn-5, that exhibits P1 spindle orientation defects, and four new mutants (spn-6-9) with defects in P0 mitotic spindle positioning. Finally, we propose a novel and powerful screen to identify genes required for spindle orientation and asymmetric cell division. We are using RNA interference to identify loci throughout the genome that, when reduced in function, can enhance or suppress conditional mutants grown at just-viable or just-lethal temperatures. Our long-term goal is to attain a comprehensive view of the pathways that regulate cell polarity in the early C. elegans embryo.
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