Date of Award

2015

Embargo Period

8-1-2024

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Oral Health Sciences

College

College of Graduate Studies

First Advisor

Vasmsi Gangaraju

Second Advisor

Viswanathan Palanisamy

Third Advisor

Andrew G. Jakymiw

Fourth Advisor

Dennis K. Watson

Fifth Advisor

Lauren E. Ball

Abstract

Post-transcriptional gene regulation plays an important role in controlling gene expression patterns in mammalian cells. CUG-binding and Embryonic Lethal vision-type RNA-binding protein 3 (ETR)-like Factor (CELF1), also known as CUGBP1, is a RNA-binding protein involved in post-transcriptional gene regulation through nuclear (alternative splicing) and cytoplasmic (mRNA turnover and translation) mRNA processing events. Primarily, CELF1 is known for its contribution to the development of myotonic dystrophy (DM1). But, published observations from our laboratory and others determined that CELF1 is overexpressed in head and neck squamous cell carcinoma (HNSCC) as well as multiple other cancers. Unpublished proteomic pulsed-stable isotope labeling by amino acids in cell culture (pSILAC) from our laboratory, has identified approximately putative 1350 CELF1 target proteins were controlled at the mRNA translation level in UM74B oral cancer cells. The pSILAC data and subsequent ribosome profiling validation experiments revealed that the protein expression of MARCKS (Myrisoylated alanine-rich kinase C substrate) is significantly reduced upon CELF1 knockdown in comparison to control oral cancer cells. MARCKS is known to play an important role in cell shape, cell motility, secretion, transmembrane transport, and regulation of the cell cycle in cancer cells. We have determined that MARKCS protein is overexpressed in HNSCC cell lines compared to primary normal oral keratinocytes. Moreover, CELF1 directly controls the expression of MARCKS, but its mRNA levels remain unchanged in oral cancer cells. CELF1 ribonucleoprotien immunoprecipitation (RNP-IP) experiments determined that MARCKS mRNA is directly associated with CELF1. The cell migratory role of MARCKS was evaluated by a scratch wound healing assay and transwell migration assay. Depletion of CELF1 and MARCKS independently resulted in reduction of cell migration. Moreover, overexpression of CELF1 in normal non-malignant human oral keratinocyte cells resulted in overexpression of MARCKS and enhanced cell migration. Finally, shRNA-mediated reduction of CELF1 in association with exogenously expressed MARCKS in oral cancer cells, we were able to rescue the CELF1-mediated cell migration phenotype. In conclusion, our results demonstrate that CELF1 controls cell migration through regulating MARCKS protein translation in oral cancer cells.

Rights

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