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Master of Science (MS)
Molecular Cell and Developmental Biology (Medicine)
Date of Defense
First Committee Member
Pedro Salas - Committee Chair
Second Committee Member
Teresa Zimmers - Committee Member
Third Committee Member
David Helfman - Mentor
Fourth Committee Member
Joyce Slingerland - Outside Committee Member
We have previously reported that non-muscle myosin II (NMMII) and myosin light chain kinase (MLCK) are required for oncogenic Ras signaling to ERK in Ki-Ras transformed rat fibroblsasts (Helfman and Pawlak, J. Cell Biochem. 95(5), 1069-80, 2005). Here I examine if MLCK plays a role in ERK signaling in various tumor derived human epithelial cell lines. I also determined whether genetic inhibition of NMMII isoforms IIA and IIB, or MLCK also inhibits ERK activation in the MCF 10A human breast epithelial cell line expressing oncogenic H-Ras. Inhibition of MLCK by pharmacological inhibitors such as ML-7 and ML-9 was used to determine the role of MLCK in ERK signaling in an array of H/K-Ras transformed and tumor derived cell lines: T-24 bladder carcinoma, HCT 116 colon carcinoma, and MCF 10A Ras breast cancer cells. Genetic inhibition was carried out using specific siRNA targeted towards MLCK and NMMIIA or IIB. The knock down of NMM IIA and IIB did not inhibit active ERK, which suggested either a redundant function of NMM IIC or an alternate substrate for MLCK. Inhibition of MLCK by ML-7/ML-9 reduced activated ERK in all H/K-Ras transformed, or human tumor derived cell lines we tested. The possible mechanism of how MLCK could play a role in ERK signaling was tested by co-immunoprecipitation (co-IP) of MAPK scaffolding proteins with MLCK. That the ERK scaffold KSR1 regulates ERK signaling in MCF 10A Ras, was demonstrated through inhibition of KSR1 with siRNA. Moreover, KSR was shown to interact with MLCK because it was found to co-precipitate with MLCK.
Signaling; Cytoskeleton; Cancer
Khan, Protiti, "The Role of Myosin Light Chain Kinase and Non Muscle Myosin II In Ras Signaling to ERK" (2008). Open Access Theses. 177.