NSF Award
9905070
The overall objective of this project is to study the involvement of the heterotrimeric G protein subunit, Gi(alpha2), in cellular differentiation. The hematopoietic cell lines K562 and U937 will be used as experimental paradigms. Differentiation will be induced in these cells with sodium butyrate and phorbol myristic acetate to achieve different terminal phenotypes. The project will accomplish three specific objectives. First, pertussis toxin and antisense methods to inactivate Gi(alpha2) will be used to test the hypothesis that differentiation of K562 cells (measured by hemoglobin expression) requires Gi(alpha2). Second, the idea that transcriptional mechanisms are responsible for changes in Gi(alpha2) levels will be verified and confirmed. Third, the probability that Gi(alpha2) or related G protein subunits may be translocated to the nucleus during differentiation will be assessed. G protein subunits will be detected by immunoblotting with affinity-purified antibodies. Nuclear translocation will be assayed by cell fractionation methods as well as by immunofluorescence and confocal microscopy. Conventional wisdom of how G proteins mediate signal transduction has not traditionally considered their translocation to the nuclear compartment as a potential mode of action. Therefore, this project has the potential of revealing novel roles for G alpha subunits that do not involve their traditional role in activating transmembrane effectors near their site of formation. The project will also provide training of students of ethnic minority origin, who are historically under-represented in the scientific community.
