This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Myo2p, a member of the class V myosins in budding yeast, functions in polarity establishment, in secretory vesicle trafficking in vacuolar inheritance, and in nuclear segregation, golgi, and peroxisome inheritance. Although Myo2p is involved in many processes, only two docking protein complexes are known. Myo2p binds to the vacuolar membrane through the -Vac17p-Vac8p complex, and Kar9p serves as the linker between Myo2p and spindle microtubule to allow nuclear segregation. We have several lines of evidence that show that Myo2p is part of an RNA-protein complex (RNP). The goal of this collaboration is to identify novel proteins in this complex, including linkers of Myo2p to its RNA cargo. We have partially purified the Myo2p RNP complex via differential centrifugation experiments. This complex is present in a high speed pellet fraction (P3). The following experiments show that Myo2p is part of an RNP: 1. On sucrose velocity gradients, Myo2p from P3 fractionation shifts dramatically with the addition of RNase. Myo2p does not bind to actively translating ribosomes, as cycloheximide treatment has no effect on the fractionation pattern. The goal of this collaboration is to identify novel proteins in this complex, including linkers of Myo2p to its RNA cargo.
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