Reprogramming of Rat Hepatoma Cells with Fibroblast-specific Protein Twist1

In mammalian development, a complex system comprised of regulatory signals causes tissue distinction with unique structures as well as functions. Gene expression controls these functions through specific combinations of transcription factors and cofactors that influence cell differentiation by both activation and repression of genes. Whole genome microarray studies of fibroblasts have identified candidate genes that can serve as master regulators of fibroblast identity. A previous study showed that Prrx1 and Snai2 play important roles in activating expression of fibroblast identity, and Snai2 overexpression in hepatoma cells (Fg14) activated expression of fibroblast specific genes. Moreover, Snai2 overexpression resulted in repression of liver specific genes. This thesis addresses whether Twist1 has a similar effect as Snai2. More specifically, the objective of this work is to determine how Twist1 transfection of hepatoma cells (Fg14) affects fibroblast specific and hepatoma specific genes. qPCR analysis revealed that Twist1 was successfully over-expressed in pooled Fg14 transfectants and individual clones compared to the non-transfected cells. Following these experiments, expression of several important genes in hepatic and fibroblast function were monitored. Results show that, of the 7 fibroblast specific genes tested, Twist1 overexpression activated two genes, Prrx1 and Sema3a, in Fg14 hepatoma cells. However, the remaining 5 fibroblast genes were not affected. Twist1 overexpression was found to not affect expression of the 10 hepatoma-specific genes tested. Based on these results, unlike results obtained with Snai2 and Prrx1 overexpression studies, Twist1 appears to have limited potential to reprogram hepatoma cells toward the fibroblast phenotype

Role of p53 in Human Cancers

TP53 codes tumor protein 53-p53 that controls the cell cycle through binding DNA directly and induces reversible cell-cycle arrest. The protein activates DNA repair genes if mutated DNA will be repaired or activates apoptotosis if the damaged DNA cannot be fixed. Therefore, p53, so-called the “guardian of the genome,” promote cell survival by allowing for DNA repair. However, the tumor-suppressor function of p53 is either lost or gained through mutations in half of the human cancers. In this work, functional perturbation of the p53 mechanism is elaborated at the breast, bladder, liver, brain, lung cancers, and osteosarcoma. Mutation of wild-type p53 not only diminishes tumor suppressor activity but transforms it into an oncogenic structure. Further, malfunction of the TP53 leads accumulation of additional oncogenic mutations in the cell genome. Thus, disruption of TP53 dependent survival pathways promotes cancer progression. This oncogenic TP53 promotes cell survival, prevents cell death through apoptosis, and contributes to the proliferation and metastasis of tumor cells. The purpose of this chapter is to discuss the contribution of mutant p53 to distinct cancer types

Reprogramming of Rat Hepatoma Cells with Fibroblast-specific Protein Twist1

In mammalian development, a complex system comprised of regulatory signals causes tissue distinction with unique structures as well as functions. Gene expression controls these functions through specific combinations of transcription factors and cofactors that influence cell differentiation by both activation and repression of genes. Whole genome microarray studies of fibroblasts have identified candidate genes that can serve as master regulators of fibroblast identity. A previous study showed that Prrx1 and Snai2 play important roles in activating expression of fibroblast identity, and Snai2 overexpression in hepatoma cells (Fg14) activated expression of fibroblast specific genes. Moreover, Snai2 overexpression resulted in repression of liver specific genes. This thesis addresses whether Twist1 has a similar effect as Snai2. More specifically, the objective of this work is to determine how Twist1 transfection of hepatoma cells (Fg14) affects fibroblast specific and hepatoma specific genes. qPCR analysis revealed that Twist1 was successfully over-expressed in pooled Fg14 transfectants and individual clones compared to the non-transfected cells. Following these experiments, expression of several important genes in hepatic and fibroblast function were monitored. Results show that, of the 7 fibroblast specific genes tested, Twist1 overexpression activated two genes, Prrx1 and Sema3a, in Fg14 hepatoma cells. However, the remaining 5 fibroblast genes were not affected. Twist1 overexpression was found to not affect expression of the 10 hepatoma-specific genes tested. Based on these results, unlike results obtained with Snai2 and Prrx1 overexpression studies, Twist1 appears to have limited potential to reprogram hepatoma cells toward the fibroblast phenotype