Characterization, annotation and expression patterns of bovine X-degenerate Y-chromosome genes UTY, ZFY, USP9Y, PRKY, RPS4Y, TBL1Y and their isoforms

Abstract

The X degenerate region of the bovine male specific region of the Y chromosome (bMSY) harbors several single copy genes, these genes share high similarity with homologous genes on the X chromosome (Chang et.al. 2012). Y chromosome genes were assumed to play an important role in maleness and spermatogenesis through the whole life of males. Understanding the function of MSY genes would provide an indication of their role in maleness and spermatogenesis and contribute to effect early age male selection in breeding programs. The purpose of this research was to obtain full sequence of these genes, identify their isoforms and expression patterns in male and female animals, through single molecule RNA sequencing and PCR amplification of gene sequences across 15 different tissues samples. Comparison to their human and mouse orthologues helped assess the structure and level of conservation of these genes. Our hypothesis is that the X-degenerate Y-encoded and X-encoded genes and their isoforms will exhibit expression variation between male and female across different tissues, and that the Y-encoded genes will have specific expression patterns in male tissues. The mRNA isoform sequence (Iso-Seq) (Pacific Bioscience, CA) information was used to identify transcripts from Domino in liver and testis and from his daughter Dominette in liver, muscle, thalamus, lung and adipose. This analysis identified one isoform for bZFY, six isoforms for bZFX; two isoforms for bUTY, five isoforms for bUTX; three isoforms for bUSP9Y, fourteen isoforms for bUSP9X; eight isoforms for bRPS4Y1, five isoforms for bRPS4X; four potential isoforms for bTBL1Y, three potential isoforms for bTBL1X and three isoforms for bPRKX. We identified sequence regions of low homology to develop specific priming sites targeting the PCR amplification of fragments for ZFY and ZFX, UTY and UTX, USP9Y and USP9X, and successfully separated their expression pattern across 15 different tissues in four new born Holstein males and females. Furthermore, the orthologous comparison of bovine genes/genomes to the human and mouse genomes were also analyzed. Results from this dissertation identified possible sex specific isoforms of bovine X-degenerate Y-encoded genes and its homolog X-chromosome genes, and it refined the database information for the mRNA sequence of those bovine genes. Moreover, it provided a comprehensive list of orthologous bovine X-degenerate Y-chromosome genes in other mammals, which provides a knowledge background to promote functional research for those X-degenerate gene in maleness. Further research needs to be focused on how expression of these genes at different developmental stages of males will regulate their male specific biological functions.