Southern California Conferences for Undergraduate Research

Southern California Conferences for Undergraduate Research

Over-expression of Mutant Rheb Proteins and mTOR Signaling

Authors:

Kusum Kumar, Laura Milbrandt

Mentor:

Nitika Parmar, Associate Professor of Biology, California State University Channel Islands

The insulin/Rheb/mTOR pathway has been implicated in a variety of cancers. The activation of mTOR (Mammalian Target of Rapamycin) is regulated by a small G-protein, Rheb. In mammalian systems there are two Rheb genes- Rheb1 and Rheb2. Although the protein sequences of these two genes share a 51% homology, the two proteins are found to be distinct with regard to amino acids residues involved in Rheb regulation. Residues involved in Rheb1 function include serine 130 and lysine 45. We made Rheb1 and Rheb2 mutants targeting amino acids at this position as well as other pertinent positions (threonine20, serine 44 and glutamine130). Wild-type and mutant plasmids were over-expressed in HEK293 cells and effect on phosphorylation of S6 was monitored. Distinct differences were observed between wild-type and mutant protein impacts on S6 phosphorylation. Cells which were expressing the mutant proteins were grown in enriched and serum deprived media followed by stimulation. They showed variations in phospho-S6 status indicating that certain mutant forms may offer a growth advantage to the cells under starved conditions. In addition, cell viability analysis indicated that over-expression of both wild-type and mutant Rheb proteins inhibited cell growth. No major differences were observed between the wild-type and mutant protein effects. This is contrary to what has been observed previously for Rheb which has been shown to have a positive impact on growth. Our results so far are unique and warrant a deeper investigation of Rheb’s impact on cellular growth.


Presented by:

Laura Milbrandt

Date:

Saturday, November 17, 2012

Poster:

22

Room:

Broome Library

Presentation Type:

Poster Presentation

Discipline:

Biology
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