Genomewide association study of liver abscess in beef cattle.

Fourteen percent of U.S. cattle slaughtered in 2011 had liver abscesses, resulting in reduced carcass weight, quality, and value. Liver abscesses can result from a common bacterial cause, , which inhabits rumen lesions caused by acidosis and subsequently escapes into the blood stream, is filtered by the liver, and causes abscesses in the liver. Our aim was to identify SNP associated with liver abscesses in beef cattle. We used lung samples as a DNA source because they have low economic value, they have abundant DNA, and we had unrestricted access to sample them. We collected 2,304 lung samples from a beef processing plant: 1,152 from animals with liver abscess and 1,152 from animals without liver abscess. Lung tissue from pairs of animals, 1 with abscesses and another without, were collected from near one another on the viscera table to ensure that pairs of phenotypically extreme animals came from the same lot. Within each phenotype (abscess or no abscess), cattle were pooled by slaughter sequence into 12 pools of 96 cattle for each phenotype for a total of 24 pools. The pools were constructed by equal volume of frozen lung tissue from each animal. The DNA needed to allelotype each pool was then extracted from pooled lung tissue and the BovineHD Bead Array (777,962 SNP) was run on all 24 pools. Total intensity (TI), an indicator of copy number variants, was the sum of intensities from red and green dyes. Pooling allele frequency (PAF) was red dye intensity divided TI. Total intensity and PAF were weighted by the inverse of their respective genomic covariance matrices computed over all SNP across the genome. A false discovery rate ≤ 5% was achieved for 15 SNP for PAF and 20 SNP for TI. Genes within 50 kbp from significant SNP were in diverse pathways including maintenance of pH homeostasis in the gastrointestinal tract, maintain immune defenses in the liver, migration of leukocytes from the blood into infected tissues, transport of glutamine into the kidney in response to acidosis to facilitate production of bicarbonate to increase pH, aggregate platelets to liver injury to facilitate liver repair, and facilitate axon guidance. Evidence from the 35 detected SNP associations combined with evidence of polygenic variation indicate that there is adequate genetic variation in incidence rate of liver abscesses, which could be exploited to select sires for reduced susceptibility to subacute acidosis and associated liver abscess.

A survey of polymorphisms detected from sequences of popular beef breeds.

The genome sequence was obtained from 270 sires used in the Germplasm Evaluation (GPE) project. These bulls included 154 purebred AI sires from GPE Cycle VII breeds (Hereford, Angus, Simmental, Limousin, Charolais, Gelbvieh, and Red Angus), 83 F crosses of those breeds, and 33 AI sires from 8 other breeds. The exome capture sequence targeting coding regions of the genome was obtained from 176 of these bulls. Sequence reads were mapped to the UMD 3.1 bovine genome assembly; a mean of 2.5-fold (x) coverage per bull was obtained from the genomic sequence, and the targeted exons were covered at a mean of 20.0x. Over 28.8 million biallelic sequence variants were detected where each allele was present in at least 3 different bulls. These included 22.0 million previously reported variants and 94.1% of the 774,660 autosomal and BTA X SNP on the BovineHD BeadChip assay (HD). More than 92% of the variants detected in targeted exons were also detected from the low-coverage genome sequence. Less than 1% of the variants detected from the combined genome and exome sequence occurred in annotated protein-coding sequences and 5' and 3' untranslated regions (UTR) surrounding the 19,994 annotated protein coding regions. Variation was detected in the coding sequence or UTR of 96.8% of the genes: loss-of-function variants were predicted for 3,298 genes, 14,973 contained nonsynonymous variants, 11,276 had variation in UTR, and 17,721 genes contained synonymous variants. Minor allele frequencies (MAF) were <0.05 for 47.8% of the coding sequence and UTR variants, and MAF distributions were skewed toward low MAF. In contrast, 11.1% of the HD SNP detected in these bulls had MAF < 0.05, and the distribution was skewed toward higher MAF. Genes involved in immune system processes and immune response were overrepresented among those genes containing high MAF loss-of-function and nonsynonymous polymorphisms. Detected variants were submitted to the National Center for Biotechnology Information genetic variation database (dbSNP) under the handle MARC, batch GPE_Bull_GenEx.

Genomewide association study of lung lesions in cattle using sample pooling.

Bovine respiratory disease complex (BRDC) is the most expensive disease in beef cattle in the United States costing the industry at least US$1 billion annually. Bovine respiratory disease complex causes damage to lung tissue resulting in persistent lung lesions observable at slaughter. Severe lung lesions at harvest have been associated with decreased preharvest ADG and increased clinical BRDC in the feedlot. Our objective was to identify SNP that are associated with severe lung lesions observed at harvest in feedlot cattle. We conducted a genomewide association study (GWAS) using a case-control design for severe lung lesions in fed cattle at slaughter using the Illumina Bovine HD array (approximately 770,000 SNP) and sample pooling. Lung samples were collected from 11,520 young cattle, a portion of which had not been treated with antibiotics (participating in a "natural" marketing program), at a large, commercial beef processing plant in central Nebraska. Lung samples with lesions (cases) and healthy lungs (controls) were collected when both phenotypes were in close proximity on the viscera (offal) table. We constructed 60 case and 60 control pools with 96 animals per pool. Pools were constructed by sampling sequence to ensure that case and control pool pairs were matched by proximity on the processing line. The Bovine HD array (770,000 SNP) was run on all pools. Fourteen SNP on BTA 2, 3, 4, 9, 11, 14, 15, 22, 24, and 25 were significant at the genomewide experiment-wise error rate of 5% ( ≤ 1.49 × 10). Eighty-five SNP on 28 chromosomes achieved a false discovery rate of 5% ( ≤ 5.38 × 10). Significant SNP were near (±100 kb) genes involved in tissue repair and regeneration, tumor suppression, cell proliferation, apoptosis, control of organ size, and immunity. Based on 85 significantly associated SNP in or near a collection of genes with diverse function on 28 chromosomes, we conclude that the genomic footprint of lung lesions is complex. A complex genomic footprint (genes and regulatory elements that affect the trait) is consistent with what is known about the cause of the disease: complex interactions among multiple viral and bacterial pathogens along with several environmental factors including dust, commingling, transportation, and stress. Characterization of sequence variation near significant SNP will enable accurate and cost effective genome-enhanced genetic evaluations for BRDC resistance in AI bulls and seed stock populations.

Evaluation of single nucleotide polymorphisms in chromosomal regions impacting pregnancy status in cattle.

Reproductive success is an important component of commercial beef cattle production, and identification of DNA markers with predictive merit for reproductive success would facilitate accurate prediction of mean daughter pregnancy rate, enabling effective selection of bulls to improve female fertility. A previous study identified SNP associated with beef cattle reproductive efficiency based on a genomewide association analysis approach using genotyping multiple-animal pools of DNA to increase the number of animals that could be genotyped with available resources. For the current study, we expand on this previous study by individually genotyping cattle from the pooling study for 89 SNP that were associated with female pregnancy rate. The aims of the study were to confirm the results of the pooling study and, more specifically, identify modes of gene action and DNA variations such as haplotypes that would not be possible with pooled genotyping. Eighty-nine SNP selected from the pooling study were evaluated using the Sequenom MassARRAY system to individually genotype animals from populations evaluated in the pooling study, including both and breeds. From this research, regions on chromosomes 5 (26.3-48.1 Mb; UMD3.1 assembly) and 9 (37,436,575 bp; UMD3.1 assembly), first identified in the previous pooling study, were shown through individual genotyping to harbor genetic variation ( < 0.05 genomewide significance) affecting reproductive efficiency in interspecific crosses ( and ) of cattle. Each of these markers exhibited additive (vs. dominant) gene action. Additionally, a haplotype block harboring an allele of origin with negative effects on reproduction was identified on chromosome 5 in interspecific composite breeds of × composites.

Genomewide association study of reproductive efficiency in female cattle.

Reproductive efficiency is of economic importance in commercial beef cattle production, as failure to achieve pregnancy reduces the number of calves marketed per cow exposed. Identification of genetic markers with predictive merit for reproductive success would facilitate early selection of sires with daughters having improved reproductive rate without increasing generation intervals. To identify regions of the genome harboring variation affecting reproductive success, we applied a genomewide association study (GWAS) approach based on the >700,000 SNP marker assay, using a procedure based on genotyping multianimal pools of DNA to increase the number of animals that could be genotyped with available resources. Cows from several populations were classified according to reproductive efficiency, and DNA was pooled within population and phenotype prior to genotyping. Populations evaluated included a research population at the U.S. Meat Animal Research Center, 2 large commercial ranch populations, and a number of smaller populations (<100 head) across the United States. We detected 2 SNP with significant genomewide association (P ≤ 1.49 × 10(-7)), on BTA21 and BTA29, 3 SNP with suggestive associations (P ≤ 2.91 × 10(-6)) on BTA5, and 1 SNP with suggestive association each on BTA1 and BTA25. In addition to our novel findings, we confirmed previously published associations for SNP on BTA-X and all autosomes except 3 (BTA21, BTA22, and BTA28) encompassing substantial breed diversity including Bos indicus and Bos taurus breeds. The study identified regions of the genome associated with reproductive efficiency, which are being targeted for further analysis to develop robust marker systems, and demonstrated that DNA pooling can be used to substantially reduce the cost of GWAS in cattle.

Deletion on chromosome 5 associated with decreased reproductive efficiency in female cattle.

Reproductive efficiency is arguably the most economically important trait in commercial beef cattle production, as failure to achieve pregnancy reduces the number of calves marketed per cow exposed to breeding. Identification of variation in the genome with predictive merit for reproductive success would facilitate accurate prediction of daughter pregnancy rate in sires enabling effective selection of bulls producing daughters with improved fertility. Copy number variation (CNV) is one form of structural variation that results in abnormal copies of DNA in the genome. Our lab previously reported a region between 25 and 70 Mb on chromosome 5 associated with reproductive efficiency. To further evaluate this region for genomic variations such as CNV, we initially applied a genomewide association approach based on the >700,000 SNP marker assay and PennCNV analysis to 68 Bos indicus cross females from a large commercial ranch in central Florida. A genomic segment located on chromosome 5, spanning the region of approximately 58.37 to 58.44 Mb (Bovine UMD3.1 assembly) was identified as containing a deletion associated with decreased reproductive efficiency. To verify this deletion, a quantitative real-time PCR test was developed to evaluate additional females in the central Florida and U.S. Meat Animal Research Center (USMARC) populations. The frequency of the homozygous deletion was 7% (23/319) in the central Florida population (Bos indicus cross) for females with 2 consecutive failed breeding seasons and 0% in the USMARC population (predominantly Bos taurus) of low reproductive females. To date, we have not identified the deletion in Bos taurus cattle, suggesting that the deletion is specific to Bos indicus cattle. From these data, we have identified a deletion on chromosome 5 associated with reproductive efficiency in Bos indicus-influenced cattle.

Identification of an ionotropic glutamate receptor AMPA1/GRIA1 polymorphism in crossbred beef cows differing in fertility.

A proposed functional polymorphism in the ionotropic glutamate receptor AMPA1 (GRIA1) has been reported to influence antral follicle numbers and fertility in cows. Repeat breeder cows that fail to produce a calf in multiple seasons have been reported to have reduced numbers of small (1 to 3 mm) antral follicles in their ovaries. Therefore, we tested the hypothesis that this GRIA1 polymorphism was affecting antral follicle numbers in repeat breeder cows. Repeat breeder cows (n = 64) and control cows (n = 72) that had always produced a calf were housed in a dry lot and observed twice daily for behavioral estrus. Blood samples were collected, and cows were genotyped for this GRIA1 polymorphism and for a polymorphism in the GnRH receptor (GnRHR) that was proposed to influence age at puberty. On d 3 to 8 after estrus cows were slaughtered, and reproductive organs were collected to determine antral follicle count, ovary size, and uterine horn diameter. Repeat breeder cows were older at first calving than control cows (P = 0.006). The length (P = 0.03) and height (P = 0.02) of the ovary contralateral to the corpus luteum (CL) were greater in control cows than repeat breeder cows. The endometrial diameter in the horn ipsilateral to the CL was greater in the control cows than the repeat breeder cows. Repeat breeder cows had fewer small (1 to 5 mm) antral follicles than control cows (P = 0.003); however, there was no association between GRIA1 genotype and antral follicle number. The GnRHR polymorphism was associated with age at first calving because cows that were homozygous for the C allele had a greater age at first calving than heterozygous cows or cows that were homozygous for the T allele (P = 0.01). In the granulosa cells from small (1 to 5 mm) antral follicles, mRNA abundances of 2 markers of oocyte quality, anti-Müllerian hormone and pentraxin 3, did not differ between fertility groups (P ≥ 0.12). We conclude that this GRIA1 polymorphism exists in beef cows but that it does not influence antral follicle numbers. The association between GnRHR genotype and age at first calving is likely not causal as this polymorphism is not functional. The utility of this polymorphism as a genetic marker for early conception in heifers will require further validation. Screening postpartum cows by ultrasonography to determine antral follicle numbers may aid in making culling decisions.

Accuracy of genomic breeding values in multibreed beef cattle populations derived from deregressed breeding values and phenotypes.

Genomic selection involves the assessment of genetic merit through prediction equations that allocate genetic variation with dense marker genotypes. It has the potential to provide accurate breeding values for selection candidates at an early age and facilitate selection for expensive or difficult to measure traits. Accurate across-breed prediction would allow genomic selection to be applied on a larger scale in the beef industry, but the limited availability of large populations for the development of prediction equations has delayed researchers from providing genomic predictions that are accurate across multiple beef breeds. In this study, the accuracy of genomic predictions for 6 growth and carcass traits were derived and evaluated using 2 multibreed beef cattle populations: 3,358 crossbred cattle of the U.S. Meat Animal Research Center Germplasm Evaluation Program (USMARC_GPE) and 1,834 high accuracy bull sires of the 2,000 Bull Project (2000_BULL) representing influential breeds in the U.S. beef cattle industry. The 2000_BULL EPD were deregressed, scaled, and weighted to adjust for between- and within-breed heterogeneous variance before use in training and validation. Molecular breeding values (MBV) trained in each multibreed population and in Angus and Hereford purebred sires of 2000_BULL were derived using the GenSel BayesCπ function (Fernando and Garrick, 2009) and cross-validated. Less than 10% of large effect loci were shared between prediction equations trained on (USMARC_GPE) relative to 2000_BULL although locus effects were moderately to highly correlated for most traits and the traits themselves were highly correlated between populations. Prediction of MBV accuracy was low and variable between populations. For growth traits, MBV accounted for up to 18% of genetic variation in a pooled, multibreed analysis and up to 28% in single breeds. For carcass traits, MBV explained up to 8% of genetic variation in a pooled, multibreed analysis and up to 42% in single breeds. Prediction equations trained in multibreed populations were more accurate for Angus and Hereford subpopulations because those were the breeds most highly represented in the training populations. Accuracies were less for prediction equations trained in a single breed due to the smaller number of records derived from a single breed in the training populations.

Associations of genetic markers in cattle receiving differing implant protocols.

The potential interaction of growth-promoting implants and genetic markers previously reported to be associated with growth, carcass traits, and tenderness was evaluated. Two implant protocols were applied to subsets of steers (n = 383) and heifers (n = 65) that were also genotyped for 47 SNP reported to be associated with variation in growth, fat thickness, LM area, marbling, or tenderness. The "mild" protocol consisted of a single terminal implant [16 mg estradiol benzoate (EB), 80 mg trenbalone acetate (TBA) or 8 mg EB, 80 mg TBA given to steers and heifers, respectively]. The "aggressive" protocol consisted of both a growing implant (8 mg EB, 40 mg TBA) for the lightest half of the animals on the aggressive protocol and 2 successive implants (28 mg EB, 200 mg TBA) given to all animals assigned to the aggressive treatment. Implant protocol had measurable impact on BW and ADG (P < 0.05), with the aggressive protocol increasing these traits before the terminal implant (relative to the mild protocol), whereas the mild protocol increased ADG after the terminal implant so that the final BW and ADG over the experimental period were similar between protocols. Animals on the aggressive protocol had significantly increased (P < 0.05) LM area (1.9 cm(2)), slice shear force (1.4 kg), and intact desmin (0.05 units), but decreased (P < 0.05) marbling score (49 units) and adjusted fat thickness (0.1 cm), and yield grade (0.15 units). Among both treatments, 8 of 9 growth-related SNP were associated with BW or ADG, and 6 of 17 tenderness-related SNP were associated with slice shear force or intact desmin. Favorable growth alleles generally were associated with increased carcass yield traits but decreased tenderness. Similarly, favorable tenderness genotypes for some markers were associated with decreased BW and ADG. Some interactions of implant protocol and genotype were noted, with some growth SNP alleles increasing the effect of the aggressive protocol. In contrast, putative beneficial effects of favorable tenderness SNP alleles were mitigated by the effects of aggressive implant. These type of antagonisms of management variables and genotypes must be accounted for in marker assisted selection (MAS) programs, and our results suggest that MAS could be used to manage, but likely will not eliminate negative impact of implants on quality.

Y are you not pregnant: identification of Y chromosome segments in female cattle with decreased reproductive efficiency.

Reproductive efficiency is of economic importance in commercial beef cattle production, since failure to achieve pregnancy reduces the number of calves marketed. Identification of genetic markers with predictive merit for reproductive success would facilitate early selection of females and avoid inefficiencies associated with sub-fertile cows. To identify regions of the genome harboring variation affecting reproductive success, we applied a genome-wide association approach based on the >700,000 SNP marker assay. To include the largest number of individuals possible under the available budget, cows from several populations were assigned to extremes for reproductive efficiency, and DNA was pooled within population and phenotype before genotyping. Surprisingly, pools prepared from DNA of low reproductive cattle returned fluorescence intensity data intermediate between fertile females and males for SNP mapped to the Y chromosome (i.e., male sex chromosome). The presence of Y-associated material in low reproductive heifers or cows was confirmed by Y-directed PCR, which revealed that 21 to 29% of females in the low reproductive category were positive by a Y chromosome PCR test normally used to sex embryos. The presence of the Y chromosome anomaly was further confirmed with application of additional Y-specific PCR amplicons, indicating the likelihood of the presence of some portion of male sex chromosome in female cattle in various beef cattle herds across the U.S. Discovery of this Y anomaly in low reproductive females may make an important contribution to management of reproductive failures in beef cattle operations.

MicroRNA expression profile in bovine cumulus-oocyte complexes: possible role of let-7 and miR-106a in the development of bovine oocytes.

The objectives of this study included: (1) identify the expression of miRNAs specific to bovine cumulus-oocyte complexes (COCs) during late oogenesis, (2) characterize the expression of candidate miRNAs as well as some miRNA processing genes, and (3) computationally identify and characterize the expression of target mRNAs for candidate miRNAs. Small RNAs in the 16-27 bp range were isolated from pooled COCs aspirated from 1- to 10-mm follicles of beef cattle ovaries and used to construct a cDNA library. A total 1798 putative miRNA sequences from the cDNA library of small RNA were compared to known miRNAs. Sixty-four miRNA clusters matched previously reported sequences in the miRBase database and 5 miRNA clusters had not been reported. TaqMan miRNA assays were used to confirm the expression of let-7b, let-7i, and miR-106a from independent collections of COCs. Real-time PCR assays were used to characterize expression of miRNA processing genes and target mRNAs (MYC and WEE1A) for the candidate miRNAs from independent collections of COCs. Expression data were analyzed using general linear model procedures for analysis of variance. The expression of let-7b and let-7i were not different between the cellular populations from various sized follicles. However, miR-106a expression was greater (P<0.01) in oocytes compared with COCs and granulosa cells. Furthermore, all the miRNA processing genes have greater expression (P<0.001) in oocytes compared with COCs and granulosa cells. The expression of potential target mRNAs for let-7 and let-7i (i.e., MYC), and miR-106a (i.e., WEE1A) were decreased (P<0.05) in oocytes compared with COCs and granulosa cells. These results demonstrate specific miRNAs within bovine COCs during late oogenesis and provide some evidence that miRNAs may play a role regulating maternal mRNAs in bovine oocytes.

Physiology and Endocrinology Symposium: How single nucleotide polymorphism chips will advance our knowledge of factors controlling puberty and aid in selecting replacement beef females.

The promise of genomic selection is accurate prediction of the genetic potential of animals from their genotypes. Simple DNA tests might replace low-accuracy predictions for expensive or lowly heritable measures of puberty and fertility based on performance and pedigree. Knowing with some certainty which DNA variants (e.g., SNP) affect puberty and fertility is the best way to fulfill the promise. Several SNP from the BovineSNP50 assay have tentatively been associated with reproductive traits including age at puberty, antral follicle count, and pregnancy observed on different sets of heifers. However, sample sizes are too small and SNP density is too sparse to definitively determine genomic regions harboring causal variants affecting reproductive success. Additionally, associations between individual SNP and similar phenotypes are inconsistent across data sets, and genomic predictions do not appear to be globally applicable to cattle of different breeds. Discrepancies may be a result of different QTL segregating in the sampled populations, differences in linkage disequilibrium (LD) patterns such that the same SNP are not correlated with the same QTL, and spurious correlations with phenotype. Several approaches can be used independently or in combination to improve detection of genomic factors affecting heifer puberty and fertility. Larger samples and denser SNP will increase power to detect real associations with SNP having more consistent LD with underlying QTL. Meta-analysis combining results from different studies can also be used to effectively increase sample size. High-density genotyping with heifers pooled by pregnancy status or early and late puberty can be a cost-effective means to sample large numbers. Networks of genes, implicated by associations with multiple traits correlated with puberty and fertility, could provide insight into the complex nature of these traits, especially if corroborated by functional annotation, established gene interaction pathways, and transcript expression. Example analyses are provided to demonstrate how integrating information about gene function and regulation with statistical associations from whole-genome SNP genotyping assays might enhance knowledge of genomic mechanisms affecting puberty and fertility, enabling reliable DNA tests to guide heifer selection decisions.

Predicting breed composition using breed frequencies of 50,000 markers from the US Meat Animal Research Center 2,000 Bull Project.

Knowledge of breed composition can be useful in multiple aspects of cattle production, and can be critical for analyzing the results of whole genome-wide association studies currently being conducted around the world. We examine the feasibility and accuracy of using genotype data from the most prevalent bovine genome-wide association studies platform, the Illumina BovineSNP50 array (Illumina Inc., San Diego, CA), to estimate breed composition for individual breeds of cattle. First, allele frequencies (of Illumina-defined allele B) of SNP on the array for each of 16 beef cattle breeds were defined by genotyping a large set of more than 2,000 bulls selected in cooperation with the respective breed associations to be representative of their breed. With these breed-specific allele frequencies, the breed compositions of approximately 2,000 two-, three-, and four-way cross (of 8 breeds) cattle produced at the US Meat Animal Research Center were predicted by using a simple multiple regression technique or Mendel (http://www.genetics.ucla.edu/software/mendel) and their genotypes from the Illumina BovineSNP50 array, and were then compared with pedigree-based estimates of breed composition. The accuracy of marker-based breed composition estimates was 89% when using either estimation method for all breeds except Angus and Red Angus (averaged 79%), based on comparing estimates with pedigree-based average breed composition. Accuracy increased to approximately 88% when these 2 breeds were combined into an aggregate Angus group. Additionally, we used a subset of these markers, approximately 3,000 that populate the Illumina Bovine3K (Illumina Inc.), to see whether breed composition could be estimated with similar accuracy when using this reduced panel of SNP makers. When breed composition was estimated using only SNP in common with the Bovine 3K array, accuracy was slightly reduced to 83%. These results suggest that SNP data from these arrays could be used to estimate breed composition in most US beef cattle in situations where pedigree is not known (e.g., multiple-sire natural service matings, non-source-verified animals in feedlots or at slaughter). This approach can aid analyses that depend on knowledge of breed composition, including identification and adjustment of breed-based population stratification, when performing genome-wide association studies on populations with incomplete pedigrees. In addition, SNP-based breed composition estimates may facilitate fitting cow germplasm to the environment, managing cattle in the feedlot, and tracing disease cases back to the geographic region or farm of origin.

Genome-wide association study of growth in crossbred beef cattle.

Chromosomal regions harboring variation affecting cattle birth weight and BW gain to 1 yr of age were identified by marker association using the highly parallel BovineSNP50 BeadChip (50K) assay composed of 54,001 individual SNP. Genotypes were obtained from progeny (F(1); 590 steers) and 2-, 3-, and 4-breed cross grandprogeny (F(1)(2) = F(1) x F(1); 1,306 steers and 707 females) of 150 AI sires representing 7 breeds (22 sires per breed; Angus, Charolais, Gelbvieh, Hereford, Limousin, Red Angus, and Simmental). Genotypes and birth, weaning, and yearling BW records were used in whole-genome association analyses to estimate effects of individual SNP on growth. Traits analyzed included growth component traits: birth weight (BWT), 205-d adjusted birth to weaning BW gain (WG), 160-d adjusted postweaning BW gain (PWG); cumulative traits: 205-d adjusted weaning weight (WW = BWT + WG) and 365-d adjusted yearling weight (YW = BWT + WG + PWG); and indexes of relative differences between postnatal growth and birth weight. Modeled fixed effects included additive effects of calf and dam SNP genotype, year-sex-management contemporary groups, and covariates for calf and dam breed composition and heterosis. Direct and maternal additive polygenic effects and maternal permanent environment effects were random. Missing genotypes, including 50K genotypes of most dams, were approximated with a single-locus BLUP procedure from pedigree relationships and known 50K genotypes. Various association criteria were applied: stringent tests to account for multiple testing but with limited power to detect associations with small effects, and relaxed nominal P that may detect SNP associated with small effects but include excessive false positive associations. Genomic locations of the 231 SNP meeting stringent criteria generally coincided with described previously QTL affecting growth traits. The 12,425 SNP satisfying relaxed tests were located throughout the genome. Most SNP associated with BWT and postnatal growth affected components in the same direction, although detection of SNP associated with one component independent of others presents a possible opportunity for SNP-assisted selection to increase postnatal growth relative to BWT.

MicroRNA: mechanism of gene regulation and application to livestock.

Posttranscriptional regulation of gene expression plays a role in multiple cellular pathways. MicroRNA (miRNA) are an emerging class of small RNA that regulate gene translation. However, the mechanisms by which miRNA regulate this process remain controversial. By altering posttranscriptional regulation, miRNA have a role in guiding developmental decisions, including cell fate, cell cycle progression, apoptosis, adipocyte differentiation, and processes that alter muscle development and growth. The role of miRNA in developmental decisions that affect animal biology is of significant interest, yet the current literature is limited in livestock models. Therefore, a review of the mechanisms by which miRNA alter gene translation and the current research evaluating miRNA in production livestock is needed.

Ankyrin repeat and suppressor of cytokine signaling (SOCS) box-containing protein (ASB) 15 alters differentiation of mouse C2C12 myoblasts and phosphorylation of mitogen-activated protein kinase and Akt.

Ankyrin repeat and suppressor of cytokine signaling box-containing protein (ASB) 15 is a novel ASB gene family member predominantly expressed in skeletal muscle. We have previously reported that overexpression of ASB15 delays differentiation and alters protein turnover in mouse C(2)C(12) myoblasts. However, the extent of ASB15 regulation of differentiation and molecular pathways underlying this activity are unknown. The extracellular signal-regulated kinase (Erk) 1/2 and phosphatidylinositol-3 kinase-Akt (PI3K/Akt; Akt is also known as protein kinase B) signaling pathways have a role in skeletal muscle growth. Activation (phosphorylation) of the Erk1/2 signaling pathway promotes proliferation, whereas activation of the PI3K/Akt signaling pathway promotes myoblast differentiation. Accordingly, we tested the hypothesis that ASB15 controls myoblast differentiation through its regulation of these kinases. Stably transfected myoblasts overexpressing ASB15 (ASB15+) demonstrated decreased differentiation, whereas attenuation of ASB15 expression (ASB15-) increased differentiation. However, ASB15+ cells had less abundance of the phosphorylated mitogen-activated protein kinase (active) form, despite decreased differentiation relative to control myoblasts (ASB15Con). The mitogen-activated protein kinase kinase inhibitor, U0126, effectively decreased mitogen-activated protein kinase phosphorylation and stimulated differentiation in ASB15- and ASB15Con cells. However, inhibition of the Erk1/2 pathway was unable to overcome the inhibitory effect of overexpressing ASB15 on differentiation (ASB15+), suggesting that the Erk1/2 pathway is likely not the predominant mediator of ASB15 activity on differentiation. Expression of ASB15 also altered phosphorylation of the PI3K/Akt pathway, as ASB15+ and ASB15- cells had decreased and increased Akt phosphorylation, respectively. These data were consistent with observed differences in differentiation. Administration of IGF-I, a PI3K/Akt activator, in ASB15+ was able to partially override the previously observed phenotype of delayed differentiation, whereas administration of the PI3K/ Akt inhibitor, LY294002, decreased phosphorylation of Akt and differentiation of all cell lines similar to the untreated ASB15+ myoblasts. These results provide initial evidence that ASB15 has a role in early myoblast differentiation and that its effects may be mediated in part by the PI3K/Akt signal transduction pathway.

Ankyrin repeat and SOCS box protein 15 regulates protein synthesis in skeletal muscle.

Ankyrin repeat and SOCS box protein 15 (ASB15) is an Asb family member expressed predominantly in skeletal muscle. We have previously reported that ASB15 mRNA abundance decreases after administration of beta-adrenergic receptor agonists. Because beta-adrenergic receptor agonists are known to stimulate muscle hypertrophy, the objective of this study was to determine whether ASB15 regulates cellular processes that contribute to muscle growth. Stable myoblast C2C12 cells expressing full-length ASB15 (ASB15-FL) and ASB15 lacking the ankyrin repeat (ASB15-Ank) or SOCS box (ASB15-SOCS) motifs were evaluated for changes in proliferation, differentiation, protein synthesis, and protein degradation. Expression of ASB15-FL caused a delay in differentiation, followed by an increase in protein synthesis of approximately 34% (P<0.05). A consistent effect of ASB15 overexpression was observed in vivo, where ectopic expression of ASB15 increased skeletal muscle fiber area (P<0.0001) after 9 days. Expression of ASB15-SOCS altered differentiation of myoblasts, resulting in detachment of cells from culture plates. Expression of ASB15-Ank increased protein degradation by 84 h of differentiation (P<0.05), and in vivo ectopic expression of an ASB15 construct lacking both the ankyrin repeat and SOCS box motifs decreased skeletal muscle fiber area (P<0.0001). Together, these results suggest ASB15 participates in the regulation of protein turnover and muscle cell development by stimulating protein synthesis and regulating differentiation of muscle cells. This is the first study to demonstrate a role for an Asb family member in skeletal muscle growth.

Uncoupling proteins and energy expenditure in mice divergently selected for heat loss.

The objective of this study was to determine whether variation in energy expenditure created by selection on heat loss is mediated by uncoupling protein-1 (UCP1) in brown adipose tissue. Divergent selection for heat loss developed lines of mice with high (MH) and low (ML) maintenance energy expenditure. Concentration of UCP1 mRNA in brown adipose tissue (BAT) was 93% greater in ML than in MH mice (P < 0.02). Two new lines of mice, KH and KL, were bred by backcrossing a UCP1 knockout gene into the MH and ML lines, respectively; KH and KL with both knock-out (-/-) and wild type (+/+) UCP1 genotypes were generated. At 13 wk of age, KH mice exhibited greater heat loss (166 kcal x kg(0.75) x d(-1)) than KL mice (126.4 kcalkg(0.75) x d(-1)) regardless of the UCP1 knockout (P < 0.0001). Concentration of UCP2 mRNA in BAT was 74% greater in UCP1 knockout mice (-/-) than in wild type (+/+; P = 0.0001). We conclude that response to selection for increased energy expenditure was not mediated by increased expression or function of UCP1.