Author's School

Graduate School of Arts & Sciences

Author's Department/Program

Biology and Biomedical Sciences: Molecular Cell Biology

Language

English (en)

Date of Award

1-1-2011

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Chair and Committee

Jason Weber

Abstract

The ARF tumor suppressor is upregulated upon oncogenic stress. ARF can suppress cell proliferation in both p53-dependent and -independent mechanisms. We have focused on ARF's ability to suppress protein synthesis in a p53-independent manner. Given that protein synthesis and the cell cycle are coordinately controlled, this arm of ARF tumor suppression also contributes to the restraint of cell proliferation. Our lab has recently shown that basal ARF suppresses protein synthesis in mitotic cells. The focus of my dissertation has been to determine whether basal ARF regulates cell growth in a post-mitotic setting. We used the osteoclast: OC) as a model for post-mitotic growth. We attempted to generate osteoclast-specific Arf loss using a Cre-expressing mouse, in which Cre is controlled by the Cathepsin K promoter: CtskCre/+). Surprisingly, we generated Arf -/- mice as demonstrated by genotyping, loss of ARF expression in the testis, and a phenotype comparable to traditional Arf -/- mice. Furthermore, both Cathepsin K and Cre are expressed in reproductive tissues, which results in Cre activity within gametes as confirmed by crossing CtskCre/+ mice with ROSA reporter mice. Finally, we found that Cathepsin K loss enhances serum estradiol levels. Together, this data suggests that Cathepsin K-driven Cre will not consistently result in OC-specific gene loss and may lead to misinterpretation of phenotypes generated to study the function of OC genes. In parallel, we have analyzed the role of ARF during osteoclastogenesis in vitro and found that Arf loss enhances osteoclastogenesis as demonstrated by OC number and size, protein markers of osteoclastogenesis, and increased bone resorption. Enhanced osteoclastogenesis upon Arf loss is independent of both proliferation and p53. Furthermore, we demonstrated enhanced protein synthesis and ribosome activity during osteoclastogenesis in Arf -/- cells. As an alternative approach to studying Arf loss in vivo, we generated radiation chimeras and challenged them with RANKL. We found that Arf loss results in elevated bone resorption due, at least in part, to increased osteoclastogenesis in vivo. This data collectively suggests that ARF regulates proliferation-independent cell growth, a function that is physiologically relevant both in vitro and in vivo.

Comments

Permanent URL: http://dx.doi.org/10.7936/K7KP805P

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