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Thyroid hormones play a critical role in mammalian development and metabolism. Their activity is regulated in a complex, tissue-specific manner by three isoforms of deiodinases. The goal of this study was to sequence the full-length bovine type II deiodinase (DIO2) and type III deiodinase (DIO3) cDNAs and characterize mRNA expression levels of each of the three deiodinase isoforms in several bovine tissues. Sequencing of bovine DIO2 and DIO3 cDNAs revealed a high degree of predicted amino acid sequence identity with their orthologs in other mammalian species and demonstrated the conservation of selenocysteine residues within the catalytic domains of both bovine proteins. Bovine DIO2 and DIO3 were positioned on chromosomes 10 and 21, respectively, by radiation hybrid mapping. Expression patterns of the three deiodinase isoforms were similar for deiodinase type I (DIO1) and DIO2 to those observed in other species. Expression level of DIO3 transcripts was greatest in mammary gland and kidney, although low-level expression was detected in most tissues sampled. Results of this work will aid in the study of deiodinase gene expression and thyroid hormone regulation in cattle.

The FAS-associated death domain (FADD) protein is an adapter/signaling molecule that has been shown to function in human cells to promote apoptosis and to inhibit NF-kappaB activation. Because of the critical role that apoptosis and NF-kappaB play in a variety of disease states, we mapped the bovine FADD gene, sequenced bovine FADD cDNA, and characterized its expression in endothelial cells (EC). Sequencing of bovine FADD revealed approximately 65 and 58% amino acid sequence identity to its human and murine homologues, respectively. Bovine FADD was mapped to chromosome 29 by radiation hybrid mapping. In addition, the functionality of bovine FADD was studied. Expression of a bovine FADD dominant-negative construct blocked bacterial lipopolysaccharide (LPS)- and TNF-alpha-induced apoptosis in bovine EC consistent with previous studies of human FADD. In contrast to human FADD, elevated expression of bovine FADD had no effect on LPS- or TNF-alpha-induced upregulation of NF-kappaB-dependent gene products as assayed by E-selectin expression. Thus, while the role of FADD in mediating apoptosis is conserved across species, its role in regulating NF-kappaB-dependent gene expression is not.

At present, the density of genes on the bovine maps is extremely limited and current resolution of the human-bovine comparative map is insufficient for selection of candidate genes controlling many economic traits of interest in dairy cattle. This study describes the chromosomal mapping of 10 selected gene-associated markers to bovine linkage and radiation hybrid maps to improve the breakpoint resolution in the human-bovine comparative map near two previously identified quantitative trait loci for the linear type trait, dairy form. Two regions of conserved synteny not previously described are reported between the telomeric region of bovine chromosome 27 (BTA27) and human chromosome 3 (HSA3) p24 region and between the HSA4q34.1 region and BTA8. These data increase the number of genes positioned on the bovine gene maps, refine the human-bovine comparative map, and should improve the efficiency of candidate gene selection for the dairy form trait in cattle.

Sequence-based gene expression data are used to interpret results from functional genomic and proteomics studies. Although more than 300 000 bovine-expressed sequence tags (ESTs) are available in public databases, a more thorough and directed sampling of the expressed genome is needed to identify new transcripts and improve assembly and annotation of existing transcript sequences. Accordingly, we examined the utility of constructing cDNA libraries synthesized by arbitrarily primed RT-PCR of mRNA from tissues not well represented in the publicly available bovine EST database. A total of 33 cDNA libraries were constructed from healthy and infected mammary gland tissues of Brazilian Gir and Holstein cattle. This series of libraries was used to generate 6481 open reading frame-expressed sequence tags (ORESTES) that assembled into 1798 unique sequence elements of which, 1157 did not significantly match sequence assemblies available in the Bos taurus gene index. However, a total of 264 of these 1157 sequence elements aligned with mouse and human expressed sequences demonstrating that ORESTES is an effective resource for discovery of novel expressed sequences in cattle. Furthermore, comparison of the alignment position of bovine ORESTES-derived sequence elements to human gene reference sequences suggested that the priming events for cDNA synthesis more often occurred at the central portion of a transcript, which may have contributed to the relatively high rate of novel sequence discovery.