Identification of Smooth Muscle Gamma Actin in Prostate Epithelia
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Prostate cancer is the second most common form of cancer in American men. Though broad treatments are available, these often have deleterious effects on the patient. One approach which may result in a more tailored treatment for prostate cancer resides in the understanding of the expression of smooth muscle gamma actin (SMGA) in the physiology of cancerous prostate epithelial cells. It is important to focus on SMGA due to its increased expression in the prostate epithelia during cancer progression. Previous research indicates that SMGA protein is present in the prostate epithelia, but as of yet, it has not been detected by traditional methods. The goal of this study is to determine the discrepancy between SMGA protein found in the prostate epithelia from that found in the prostate smooth muscle. In order to demonstrate SMGA protein is in fact present in prostate epithelia, we subjected lysates derived from a prostate cancer cell line, PC-3, to Western blot analysis, immunoprecipitation, and mass spectroscopy. We used monoclonal muscle actin isoform-specific antibody, HUC1-1, to determine that these isoforms were present in lysate samples. As experimental controls, actin purified from chicken gizzard smooth muscle cells was run in parallel to the lysate on the same gel. We next used HUC1-1 in immunoprecipitation experiments to precipitate actin from the lysate. We found that its band matched to the control of purified actin which migrated at 43 kDa on the gel system. The band from the precipitated actin was then cut from the gel and subjected to mass spectroscopy analysis for sequencing. These experiments identified multiple peptides that are diagnostic of SMGA. However, we did not detect the amino terminus of the prostate epithelial specific-SMGA. Suggested studies would be: 1) to sequence the amino acids at the N-terminus using Edman degradation and compare the sequenced protein to SMGA found in mature smooth muscle; and in the long term, 2) synthesize an antibody specific to prostate epithelial specific-SMGA to determine its function in cancerous prostate.
Kolcun, Georgina Mary (2014). Identification of Smooth Muscle Gamma Actin in Prostate Epithelia. Honors and Undergraduate Research. Available electronically from