Date of Award


Document Type

Open Access Master's Thesis

Degree Name

Master of Science in Biological Sciences (MS)

Administrative Home Department

Department of Biological Sciences

Advisor 1

Paul Goetsch

Committee Member 1

Ebenezer Tumban

Committee Member 2

Zhiying Shan


Misexpression of germline genes like Cancer Testis (CT) genes, called a soma-to-germline transformation, is a phenomenon linked to tumorigenesis. However, the mechanisms underlying this phenomenon are poorly understood. A soma-to-germline transformation in Caenorhabditis elegans occurs due to the loss function of the highly conserved DREAM (Dp, Retinoblastoma (Rb)-like, E2F, and MuvB) transcriptional repressor complex. In mammalian cells, the DREAM complex (Muvb core complex, E2F4/5, DP1/2, and p130/p107 proteins), as well as the Retinoblastoma protein (pRb), are implicated in transcriptional repression of cell cycle genes in quiescence or G0. We hypothesize that the expression of CT genes in malignant cells occurs because of the loss of activities of DREAM complex or pRb, similar to how the soma-to-germline transformation occurs in C. elegans. Thus, we expect that cancer cells that express CT genes will either fail to arrest in G0 or display defective repression of key cell cycle genes. To test sensitivity cells to arrest in response to limiting growth conditions, we did flow cytometry to measure the DNA content of 10 cell lines. We found that seven cell lines arrested in G0/G1, indicating that these cells have downregulated CT genes expression, and DREAM or Rb is involved in repressing cell cycle in G0/G1. This result also suggested that in the cells that did not arrest, both DREAM and Rb are inactive. Next, to test the mRNA expression of CT genes of all the cell lines, we did mRNA analysis of CT genes. We found that CT genes are expressed in proliferating cells of both cells that can arrest under limiting growth conditions and the cells that do not, indicating we did not observe a difference in mRNA expression between cells that arrest and cells that do not. To test if CT misexpression is associated with dysfunction in either Rb or DREAM, we did further analyses to test early and late cell cycle genes expression in SW480, NCI-H1299, and 8MGBA, comparing the expression of cell cycle cells in proliferating cells to arrested cells in G0/G1. We found that mRNA expression of early cell cycle genes MCM5, ORC1, and CDC45 are downregulated, as expected since DREAM and Rb regulation of these genes overlap. However, DREAM solely regulates G2/M genes like CCNB2, PBK, and BUB1. We found that G2/M genes were not in significantly downregulated in NCI-H1299 and SW-480 cells, suggesting that DREAM is dysfunctional in these cell lines. As a secondary test of DREAM and Rb function, we performed luciferase reporter assays with promoters of DREAM and Rb target genes in SW480 and NCI-H1299 cell lines. Surprisingly, we observed that Rb is dysfunctional in SW480 cells. Together, these studies will facilitate future studies into the link between cell cycle regulation and CT upregulation in cancer cells.