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Title: Functional analysis of the promoters of the Y-linked genes, Ube1y and Usp9y
Author: Hall, N.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2002
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Abstract:
Ube1y and Usp9y map to the Sxrb region of the mouse Y chromosome, deletion of which results in infertility. This study aimed to investigate the basis for their testis-specific expression pattern. The 8bk coding sequence of Usp9y was determined, confirming that it encodes a potentially functional protein. There is evidence to suggest that the gene is subject to alternative splicing. The genomic structure of the gene was analysed and indicated the Usp9y covers 47 exons. Six kilobases of genomic sequence encompassing exon one of Ube1y was obtained and 1.2kb genomic sequence upstream of the transcriptional start site of Usp9y was generated. These sequences were subjected to a number of promoter prediction programmes. Neither Ube1y nor Usp9y appear to contain TATA boxes, but do have potential Initiator sites and CCAAT boxes as well as a large number of additional putative transcription factor binding sites. Two germ cell-specific cell lines (GC1 and GC2), and a somatic cell line (3T3) were selected for use in transfection studies to assess promoter activity of Usp9y and Ube1y. Expression of Usp9y and Ube1y, other testis-specific and non-testis-specific genes was investigated by RT-PCR to characterize the cell lines. Luciferase reporter constructs were generated containing the putative promoter regions for each gene and shown to be active in the germ-cell specific lines. Further deleted constructs indicated that core promoter activity for Usp9y resides within the 300bp 5' of its transcriptional start site, and for Ube1y, within 100bp. Constructs that were deleted for the first intron of Ube1y indicated a potential repressor element. All constructs also showed activity in the somatic 3T3 cell line, suggesting either that the regions conferring testis specific expression were not present or that the transformed cell lines were not representative of cells in vivo.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.599857  DOI: Not available
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