Molecular characterization of the DDX3Y gene and its homologs in cattle.

DDX3Y (also known as DBY) is a member of the DEAD box protein family, which is involved in ATP-dependent RNA unwinding, needed in a variety of cellular processes including splicing, ribosome biogenesis and RNA degradation. In the human, DDX3Y is located in the AZFa interval in the Y chromosome. Deletion of the AZFa region has been shown to disrupt spermatogenesis, causing subfertility and infertility in otherwise healthy men. Here, we report the characterization of the bovine (b) DDX3Y gene and its homologs DDX3X and PL10. We found 2 transcripts for the bDDX3Y (bDDX3Y-L and -S), which correspond to the long and short transcripts of the human DDX3Y and mouse Ddx3y gene. The 2 transcripts are identical except for a 3-bp (AGT) insertion at the position of nt 2025 and an expanded 3'UTR (nt 2155-2769) in bDDX3Y-L. The bDDX3Y-S encodes a peptide of 660 amino acids (aa), while the bDDX3Y-L encodes a peptide of 661 aa as the result of an additional serine (S) insertion at the position of aa 634. Both bDDX3Y isoforms contain the conserved DEAD-box motif. The bDDX3Y is composed of 17 exons. The homologous gene on the X chromosome, bDDX3X, is highly conserved to the Y-copy at mRNA (83%) and protein (88%) levels as well as in the genomic structure. The autosomal copy, bPL10, mapped on BTA15, is a processed pseudogene with a similarity of 88.1% to bDDX3Y and 93.7% to bDDX3X mRNA, suggesting that PL10 is a retroposon of DDX3X. RT-PCR analyses showed that bDDX3Y-L, -S, bDDX3X and bPL10 were all widely expressed with predominant expression in testis and brain. Testicular section in situ hybridization revealed that sense and anti-sense RNAs of bDDX3Y-L, -S, and bDDX3X were expressed in interstitial cells. These results together with the finding that the pseudogene bPL10 is transcriptionally active in this study provide a basis for further investigating the DDX3 gene function in spermatogenesis, male fertility and gene evolution in mammals.

High-resolution comprehensive radiation hybrid maps of the porcine chromosomes 2p and 9p compared with the human chromosome 11.

We are constructing high-resolution, chromosomal 'test' maps for the entire pig genome using a 12,000-rad WG-RH panel (IMNpRH2(12,000-rad))to provide a scaffold for the rapid assembly of the porcine genome sequence. Here we present an initial, comparative map of human chromosome (HSA) 11 with pig chromosomes (SSC) 2p and 9p. Two sets of RH mapping vectors were used to construct the RH framework (FW) maps for SSC2p and SSC9p. One set of 590 markers, including 131 microsatellites (MSs), 364 genes/ESTs, and 95 BAC end sequences (BESs) was typed on the IMNpRH2(12,000-rad) panel. A second set of 271 markers (28 MSs, 138 genes/ESTs, and 105 BESs) was typed on the IMpRH(7,000-rad) panel. The two data sets were merged into a single data-set of 655 markers of which 206 markers were typed on both panels. Two large linkage groups of 72 and 194 markers were assigned to SSC2p, and two linkage groups of 84 and 168 markers to SSC9p at a two-point LOD score of 10. A total of 126 and 114 FW markers were ordered with a likelihood ratio of 1000:1 to the SSC2p and SSC9p RH(12,000-rad) FW maps, respectively, with an accumulated map distance of 4046.5 cR(12,000 )and 1355.2 cR(7,000 )for SSC2p, and 4244.1 cR(12,000) and 1802.5 cR(7,000) for SSC9p. The kb/cR ratio in the IMNpRH2(12,000-rad) FW maps was 15.8 for SSC2p, and 15.4 for SSC9p, while the ratio in the IMpRH(7,000-rad) FW maps was 47.1 and 36.3, respectively, or an approximately 3.0-fold increase in map resolution in the IMNpRH(12,000-rad) panel over the IMpRH(7,000-rad) panel. The integrated IMNpRH(12,000-rad) andIMpRH(7,000-rad) maps as well as the genetic and BAC FPC maps provide an inclusive comparative map between SSC2p, SSC9p and HSA11 to close potential gaps between contigs prior to sequencing, and to identify regions where potential problems may arise in sequence assembly.

Mapping, expression, and association study of the bovine PSMC1 gene.

The 26S ATP-dependent protease is composed of a 20S catalytic proteasome and two PA700 regulatory modules; it plays a central role in many regulatory pathways, such as cell cycle regulation, differentiation, and apoptosis. The PA700 complex is composed of multiple subunits, including at least six related ATPases and approximately 15 non-ATPase polypeptides. PSMC1 (proteasome 26S subunit, ATPase, 1) is one of these ATPases. In this study, we amplified a fragment of 507 bp from intron 9 of the bovine PSMC1 gene and found a SNP (G/A) at position 216 in the PCR fragment. Genotyping of 138 animals from four beef breeds revealed that the average frequency for allele A (G-base) was 0.4271 (0.3269-0.5517); for allele B (A-base) it was 0.5729 (0.4483-0.6731). This SNP is significantly associated with average daily feed intake (P < 0.01), average daily gain, finishing average daily gain, body length, ratio of feed to meat, backfat thickness, and loin-muscle area (P < 0.05). Our experimental data showed that animals with an AA genotype have a significantly lower food intake, grow faster, are longer in the body, and have less backfat and bigger loin muscle; hence, their ratio of feed to meat is significantly lower. We believe that the PSMC1 SNP is a potential candidate marker for marker-assisted selection in these traits. We also found that the bovine PSMC1 gene was expressed mainly in lung, testis, and spleen. In addition, we mapped the bovine PSMC1 gene on BTA10 by an RH mapping method.