Lukianov, Stefan
(2013)
A Transgenic Mouse Model of Merkel Cell Virus Small Tumor Antigen.
Master's Thesis, University of Pittsburgh.
(Unpublished)
Abstract
Merkel cell carcinoma (MCC), a primary cutaneous neoplasm, originates in the mechanoreceptor Merkel cells in the basal layer of the epidermis. Risk factors include UV exposure, advanced age and immunosuppression, suggesting an infectious etiology. MCC
incidence in the US is rising, with approximately 1500 cases per year. The non-enveloped, double-stranded DNA Merkel cell polyomavirus (MCV) is responsible for approximately 80% of MCC cases. The virus was discovered by subjecting MCC tissue samples to digital transcriptome subtraction, in which mRNA is isolated, the human transcripts subtracted in silico and the remaining transcripts compared to viral sequences.
MCV expresses differentially spliced Large (LT), Small (sT) and 57 kT tumor antigens from the T antigen early locus, similar to other polyomaviruses such as SV40. Both LT and sT are critical for transformation. LT is a helicase responsible for replication of the viral genome, however in integrated viral genomes it is either truncated or mutated to eliminate its replicative functions. sT contributes to transformation via hyperphosphorylation and inhibition of the cap-dependent translation inhibitor 4E-BP1. The function of 57 kT remains unknown. Knockdown of LT induces necroptosis of MCVpositive MCC cells, whereas sT expression in rodent Rat-1 cells is transformative.
Being that sT is the transformative agent in rodent cells, it would be of interest to develop a mouse model expressing sT in a tissue-specific manner to determine whether tumor formation occurs. Indeed, several mouse models of SV40 T antigen have been developed over the past decades, each resulting in tissue-specific tumor formation. We developed a MCV sT transgenic mouse model, in which a lox-stop-lox sT is expressed via an ER-inducible Cre gene under the control of the ubiquitin promoter. Upon tamoxifen induced MCV sT expression, ER-Cre-positive mice demonstrate severe weight loss, ruffled fur and a hunched posture, necessitating euthanasia. Western blotting reveals sT expression in several tissues, whereas TUNEL staining shows significant cell death. While
we were unable to observe transformation, we believe this drastic phenotype demonstrates the validity of our MCV sT transgenic mouse model and warrants further investigation into the mechanism of death.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
2 December 2013 |
| Date Type: |
Publication |
| Defense Date: |
1 October 2013 |
| Approval Date: |
2 December 2013 |
| Submission Date: |
25 November 2013 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
53 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Integrative Molecular Biology |
| Degree: |
MS - Master of Science |
| Thesis Type: |
Master's Thesis |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Transgenic
Merkel
Tumor |
| Date Deposited: |
02 Dec 2013 16:13 |
| Last Modified: |
14 Nov 2024 18:18 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/20100 |
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