Characterisation of LVI-1 (WDR76) as a candidate tumour suppressor gene

Miranda, Alexandra de Sousa Montenegro (2006) Characterisation of LVI-1 (WDR76) as a candidate tumour suppressor gene. PhD thesis, University of Glasgow.

Abstract

The central aim of this study was to characterise the expression of the candidate
tumour suppressor gene, LV/-J (lentivirus integration-I] and its products. The LVI-J
gene (WDR76) was discovered as a target for disruption by proviral insertional
mutagenesis in a case of pre-B cell lymphoma in a FIV infected cat {Beatty, 1998 5
lid; Beatty, 2002 7 lid}. As LV/-J is highly conserved, my work focused on the
human and murine orthologues to take advantage of the superior resources available
for these species. My work showed potentially important differences in expression of
the human and mouse genes with respect to promoter use and length of 3'
untranslated sequences. The murine gene is transcribed mainly from the distal PI
promoter, which appears to be a bi-directional element shared with the adjacent
MJapJ gene, while the human gene transcripts are derived exclusively from the
proximal P2 promoter. Direct analysis by RT-PCR showed that the murine gene
could also be expressed from the P2 promoter. These findings have significant
implications for Lvi-l protein expression as the P2 transcripts are predicted to express
larger proteins with a distinct N-terminal sequence. To characterise the LV/-J gene
products, rabbit polyclonal antisera were raised to GST fusion proteins expressed in
bacteria. Use of the murine anti-ml.vi-I antiserum in Western blotting identified a
protein of the expected size (58 kDa) based on translation of the major mRNA species
from the PI promoter. Immunofluorescence and confocal microscopy suggested that
this protein is localised mainly in the cytoplasm. Although the function of LVI-I is
unknown, its closest relative in the human genome is DDB2, a protein involved in
repair of UV-induced DNA damage. Regulation of LVI-I expression was examined
after UV irradiation, providing preliminary evidence of responses at transcriptional
and post-transcriptional levels. Further leads were followed by analogy with the yeast
orthologue of LVI-I, YDLI56W, but no evidence of a complex between LVI-I and
MSH6 was found. In conclusion, while function of the LVI-I gene remains to be
establishes, it provides the basis for future characterisation of this highly conserved
and potentially important eukaryotic gene.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: Q Science > QR Microbiology; R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
• Colleges/Schools: College of Medical Veterinary and Life Sciences > Institute of Biodiversity Animal Health and Comparative Medicine
• Supervisor's Name: Neil, Professor James and Stewart, Dr. Monica
• Date of Award: 2006
• Depositing User: Ms Dawn Pike
• Unique ID: glathesis:2006-4967
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 18 Feb 2014 09:54
• Last Modified: 19 Feb 2014 08:32
• URI: https://theses.gla.ac.uk/id/eprint/4967