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Genetics, Vol. 175, 843-853, February 2007, Copyright © 2007
doi:10.1534/genetics.106.064535
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CSIRO Livestock Industries, Queensland Bioscience Precinct, Saint Lucia, Queensland 4067, Australia and CRC for Cattle and Beef Quality, Armidale, New South Wales 2351, Australia
1 Corresponding author: CSIRO, Queensland Bioscience Precinct, 306 Carmody Rd., St. Lucia, QL 4067, Australia.
E-mail: bill.barendse{at}csiro.au
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ABSTRACT |
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 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONACKNOWLEDGEMENTSLITERATURE CITED |
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p of marbling and 0.28 p of rump fat, explaining the previously reported QTL effect. cDNA for RORC were sequenced and 2 new alternative transcripts were found. Fetal tissue shows 40 times greater transcription of RORC than adult tissue. The highest expression in fetal tissue was found in liver and kidney, but in adults the longissimus muscle had the greatest expression of the tissues tested.
Bovine fat depots develop in a regular sequence during the life of the animal, and the development of each depot follows a well-known pathway (PETHICK et al. 2004). Although marbling appears relatively late in the life of the animal, it is early events that are of most importance (see Figure 3 of PETHICK et al. 2004), and animals that have high starting values of intramuscular fat, before marbling appears, reach much higher final values of marbling than those with low starting values. Fatness is heavily influenced by energy intake, with significant differences between cattle fed in feedlots vs. those in pasture. The degree of fatness is also influenced by circulating hormones, vitamins, and intracellular signaling molecules and includes genetic variation of growth hormone, leptin, thyroid hormone, and their receptors (SCHLEE et al. 1994; SNEYERS et al. 1994; TORII et al. 1996; BARENDSE 1997; MEARS et al. 2001; CHIKUNI and MITSUHASHI 2002; BARENDSE et al. 2004, 2005, 2006). Vitamin A is of interest because circulating levels of retinol and retinoic acid can have significant effects on marbling fatness (TORII et al. 1996) and on adipocytes in culture (BRANDEBOURG and HU 2005).
QTL affecting fatness in cattle were located at several overlapping regions of bovine chromosome 3 (ANDERSSON-EKLUND et al. 1990; CASAS et al. 2001, 2003, 2004; BARENDSE 2003), suggesting that there are several genes affecting fatness on the chromosome, although the confidence intervals of linkage mapping studies prevent certainty (DRINKWATER et al. 2006). Several of the genes in the confidence interval of the QTL were possible candidates. These include phosphatidylinositol 4-kinase, catalytic, ß (PIK4CB) (NAKANISHI et al. 1995), protein kinase, AMP activated, ß-2 subunit (PRKAB2) (THORNTON et al. 1998), and a vitamin A receptor, the retinoic acid receptor-related orphan receptor C (RORC) gene. RORC is highly expressed in skeletal muscle (HIROSE et al. 1994). RORC is a member of the steroid and thyroid hormone receptor superfamily (PETKOVICH et al. 1987; EVANS 1988) and binds retinoic acid as well as thyroid hormone. However, most of the previous research on this gene has focused on its role in the mouse immune system (KUREBAYASHI et al. 2000; SUN et al. 2000), and single nucleotide polymorphisms (SNPs) in RORC have recently been studied in human type 2 diabetes (WANG et al. 2003).
SNPs in the genes PIK4CB, PRKAB2, and RORC, in that order, were tested on animals with extreme marbling scores to determine if any were significantly associated with marbling. For RORC, the initial SNP search was performed a priori in the ligand-binding domain, partly due to the effects of vitamin A on fatness and partly due to that part of the DNA being known to have alternatively spliced transcripts (GenBank Gene ID 6097 Evidence Viewer). The significance levels were confirmed in an independent sample of cattle. Then, a further nine SNPs in and near RORC were genotyped to try to refine the position within the RORC gene, in particular to determine the limits of the SNP association, and these and their inferred haplotypes were tested for associations to fatness in feedlot cattle. This included SNPs in the region 5' to the adjacent leucine-rich repeat neuronal 6D (LRRN6D) gene, which was identified as part of a search for genes involved in axon regeneration but whose function is otherwise unknown (MI et al. 2004). A high density of SNPs was used to locate the QTL, although high SNP densities have been used only for QTL of much larger effect in livestock due to the high level of linkage disequilibrium resulting from domestication and breed formation (FARNIR et al. 2000; MCRAE et al. 2002; TENESA et al. 2003; NSENGIMANA et al. 2004). RORC gene expression was quantified in a range of tissues to elucidate the possible mode of action of the gene.
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MATERIALS AND METHODS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONACKNOWLEDGEMENTSLITERATURE CITED |
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SNP discovery:
Initially, SNPs were obtained by sequencing part of the 3'-untranslated region (UTR) of the PIK4CB gene (NM_174783.2), the third and fourth introns and fourth exon of the PRKAB2 gene, and the eighth and ninth intron and ninth exon of the RORC gene. These genes were positional candidate genes falling within the confidence interval of the QTL.
Genomic DNA for SNP discovery was obtained by direct sequencing of PCR products. A continuous section of the RORC genomic DNA sequence from exon 3 to exon 11 was obtained using overlapping PCR fragments with primers located in introns. In addition, the 5'-UTR, exons 1 and 2, part of intron 1, the 3'-UTR of RORC, and a section of the 5' region of the LRRN6D gene were also sequenced. The first primers were designed using human and mouse exons of the gene due to lack of a bovine genome sequence at the time and the DNA sequences were compared to GenBank using BLAST (ALTSCHUL et al. 1990) to confirm the identity. However, once bovine intronic material was obtained, it was used for further primer design. Each fragment was sequenced in both directions in 10 of a total pool of 20 animals that were of either the Angus or the Shorthorn breed. Exons were identified in the genomic sequence after comparison to the cDNA sequence using BLAST. DNA sequences were obtained using the Big-Dye 3.1 terminator kit from Applied Biosystems (ABI, Foster City, CA) using the manufacturer's instructions. These sequences were assessed, assembled into contigs using Phred and Phrap (EWING et al. 1998), and viewed using Consed (GORDON et al. 1998). PolyPhred (NICKERSON et al. 1997) was used to identify variable bases with a threshold Phred quality score of 20. SNPs were described using standard nomenclature (DEN DUNNEN and ANTONARAKIS 2000).
A full-length cDNA sequence was obtained for the RORC gene using RACE. The cDNA sequence was obtained in two steps: (1) primers were synthesized for sequences in human exons 3 and 8 and, using cDNA translated from muscle mRNA with the Invitrogen (San Diego) Superscript Rnase H-Reverse transcriptase kit following the manufacturer's instructions, a cDNA fragment of the RORC gene was amplified, cloned, and sequenced, and (2) this exon 3–8 sequence was used in 5' and 3' RACE to obtain the additional exons as well as the flanking 5'-UTR and 3'-UTR. All different-sized bands were cloned and sequenced. The identities of the cDNA sequences were checked using BLAST.
SNP genotyping:
SNPs were genotyped using the Taqman MGB allele discrimination method (ABI, Foster City, CA) as before (BARENDSE et al. 2005, 2006) by two individuals. Scoring was always performed without knowing the phenotypes. For ease of analysis and compact reporting of the data in the tables, genotypes were coded as 0, 1, 2, and 5 where 5 is unknown, 1 is always the heterozygote, 0 is the homozygote higher up in the alphabet, and 2 is the homozygote lower down in the alphabet: therefore, CC is 2 when AA is the alternative homozygote but 0 when GG is the alternative homozygote.
Gene expression:
Tissues for mRNA extraction were obtained from adult and day 194 fetal tissue of cattle. RNA was extracted from skin, kidney, brain, lung, liver, longissimus dorsi (LD), semitendinosus and supraspinatus muscle; subcutaneous, cardiac, omental, and kidney fat; and mammary gland, testis, heart, spleen, pancreas, and thymus based on a previously published method (CHOMCZYNSKI and SACCHI 1987; LEHNERT et al. 2004). The tissue was dissected, immediately frozen in liquid nitrogen, wrapped in aluminium foil, disrupted with a hammer, and homogenized in TRIzol (Invitrogen, Carlsbad, CA) using an ultrasonic homogenizer (IKA-Ikasonic, Staufen, Germany), and RNA was extracted from 
1 g of tissue using the manufacturer's instructions. All tissues were not available from all animals or life stages and some tissues or organs cannot be recognized easily in a fetus. All RNA was proofed on an agarose gel and quantitated using UV spectrophotometry. Five micrograms of total RNA was translated to cDNA by the oligo(dT) method using the same Invitrogen kit as above, and the cDNA was stored at –80° in a 50-µl volume.
Gene expression was measured using quantitative RT–PCR where the primers gave different-sized products on cDNA and genomic DNA. The RORC alternative transcripts were amplified in separate reactions. The gene expression was measured by quantitative PCR in a 5-µl reaction volume using 1 µl of a 10x dilution of the cDNA with 900 nmol of each primer at 1x final concentration of the ABI SYBR Green mastermix on the ABI 7900HT. Gene expression was measured on all adult and then all fetal tissues for both transcripts and both housekeeping genes in the same run. Four replicates of each cDNA were used. A melting point analysis was performed to check that each sample gave a single amplification product. Gene expression was calibrated against the two housekeeping genes glyceraldehyde dehydrogenase GAPDH-F 5'-CCTGGAGAAACCTGCCAAGT-3' and GAPDH-R 5'-GCCAAATTCATTGTCGTACCA-3' and 18S ribosomal protein 18SrRNA-F 5'-GTAACCCGTTGAACCCCATT-3' and 18SrRNA-R 5'-CCATCCAATCGGTAGTAGCG-3' (VUOCOLO et al. 2005). The RORC transcript 1 was measured using the primer sequences RORCABU1 5'-CCACAGAGACATCACCGAGCC-3' and RORCABD2 5'-GTGGATCCCAGATGACTTGTCC-3' and transcript 2 was measured using RORCAinsU1 5'-GAGGAAGCTGTCCTGCCTCTA-3' and RORCAinsD1 5'-TAGAGGCAGGACAGCTTCCTC-3'. These primer sequences use differences in the splicing of the mRNA to ensure that only a single product is produced in each reaction. One of the primers for the alternative transcript is specific for intron 1 of RORC, so the alternative transcript is unlikely to be a pseudogene.
Analysis:
The statistical methods used were described previously (BARENDSE et al. 2004, 2006). Type I errors due to multiple testing were controlled by confirming positive associations in an independent sample. Marbling score is an ordinal trait split into several categories, and scores of 1–5 are seen with reasonable frequency. The random extreme sample consists of animals of marbling score 1 (low) and marbling score 4 (high); these scores do not overlap in their underlying intramuscular fat percentages. Differences in the distribution of marbling scores were tested using the log-likelihood test with the Williams correction (SOKAL and ROHLF 1981). All marbling scores are available for the feedlot test sample, which were analyzed using a generalized linear model with a Poisson distribution (VENABLES and RIPLEY 2000) as before, taking day of slaughter and breed into consideration. No other factor was significant in the analysis after those two were taken into account. The genetic variance was estimated by comparing the model with genotypes to the model without genotypes.
Once the high-density SNP mapping began, it was more convenient to analyze marbling as a pseudoquantitative trait using general linear mixed models. In brief, general linear mixed models were fitted to the marbling and rump fat trait measures using fixed and random effects and covariates in ASREML (GILMOUR et al. 1995, 2002). For each of the traits, the slaughter day (random), the breed (fixed), and the producer (random) were always fitted, and the significant covariates among the following were also included: dof, inwt, hdwt, p8fat, marbling, and ema. Selection among these linear models was made using Aikake's Information Criterion (VENABLES and RIPLEY 2000). The models were marbling N(µ + day + breed + p8fat + dof + producer, 
) and p8fat N(µ + day + breed + ema + marbling + dof + inwt + producer, ).
The residual trait value from the model was extracted for each individual and then matched to its genotype, and the mean trait values for each genotype were then compared using t-tests. These models were thus adjusted for systematic differences in mean value between breeds. The differences between trait means and the allele frequencies were used to calculate 
, the average effect of allele substitution of the polymorphism (LYNCH and WALSH 1998). Consistency in the a, i.e., half the distance between the homozygotes; in the k, i.e., the standardized dominance deviation; and in between breeds and the combined sample was used to help determine whether (1) a useful diagnostic marker had been obtained and (2) how close to a causative mutation the SNP was located. Summary statistics such as variances and correlation coefficients were calculated using S-Plus.
SNPs were ordered using the DNA sequence and then the genotypes were examined using PHASE v2.1 (STEPHENS et al. 2001; STEPHENS and DONNELLY 2003) to infer haplotypes. The inferred haplotype frequencies were used to calculate the Lewontin D' and the square of the correlation r2 for each pair of SNP using published formulas (DEVLIN and RISCH 1995).
Haploblock (GREENSPAN and GEIGER 2004) was used to locate the causative mutations within the RORC gene. Raw marbling scores are an ordinal trait with at least five levels in the data so it was appropriate to use this method of analysis. However, rump fat thickness, as it is a continuous trait, could not be analyzed this way.
Gene expression differences were calculated using geNorm (VANDESOMPELE et al. 2002; HUGGETT et al. 2005) and a geometric mean of the expression of both housekeeping genes was used to normalize the expression of the transcripts of interest using all four replicates.
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RESULTS |
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25 kbp. For a tag SNP (CHAPMAN et al. 2003) threshold of r2 = 0.85, a minimum of seven of these SNPs would be necessary to extract most of the genotypic information. These SNPs are RORCa:g.283G>A, RORC:g.592A>G, RORC:g.1826A>G, RORC:g.3290T>G, RORC:g.3984A>G, RORC:g.8067T>C, and LRRN6Du5:g.243T>C.
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(Table 5). The marbling scores show slightly lower values for the heterozygote, separately in Angus, in Shorthorn, and in the combined sample. The SNPs for PIK4CB and PRKAB2 were again not significantly associated with marbling.
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RORC:g.3290T>G had the strongest association with marbling (see Table 6 in which the four most significant SNPs are shown) and was the only SNP to show such an association in the total sample and in the Angus and the Shorthorn breeds separately. RORC:g.2415T>C and RORC:g.2883C>T, within 1 kb of RORC:g.3290T>G, also showed associations with marbling in the Shorthorn breed and the full sample, but not in the Angus. Of these three, RORC:g.3290T>G was the only SNP to show a consistent a, k, and for genotype comparisons in both breeds separately and in the total sample, indicating (1) that it is a useful diagnostic SNP for the trait and (2) that the SNP is located very close to causative mutations and may be one itself. The RORC:g.3290TT genotype had the highest marbling score and the lowest rump fat thickness. Although the association is significant, the combined marbling was small due to the GT heterozygotes showing the largest reduction in marbling score when compared to either homozygote.
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for rump fat thickness is 0.5 cm, which is 0.09 p.
The six most common inferred haplotypes were then examined to determine whether there were subdivisions within the alleles and to improve the estimate of the size of effect. The haplotypes show that genetic subdivisions in some of the alleles act in opposite directions (Table 7). There was no breed or breed x haplotype effect on marbling score or rump fat. The overall ANOVA was significant for the effect of haplotypes on marbling (F5,2919= 3.54, P = 0.004) but not on rump fat. The RORC:g.3290T and the RORC:g.3290G alleles were each associated with three common haplotypes. While all of the RORC:g.3290T haplotypes had positive effects on marbling, one of the RORC:g.3290G haplotypes had a positive effect on marbling, one had a negative effect, and one was zero. The maximum displacement due to these common haplotypes for RORC:g.3290T>G was 0.26 marbling scores, a difference of 0.35 p. For rump fat, of the six common haplotypes, four are associated with RORC:g.3984A and two with RORC:g.3984G. One RORC:g.3984G haplotype had a positive effect on rump fat while the other had a negative effect, while one of the RORC:g.3984A haplotypes showed no effect on rump fat and three increased rump fat thickness. The maximum difference between these common haplotypes was 1.6 cm of rump fat, a difference of 0.28 p. Haplotype 14 had the strongest negative effect on marbling and the strongest positive effect on rump fat. Haplotype 20 had no effect on either trait. Haplotype 27 had the strongest positive effect on marbling but no effect on rump fat. However, haplotype 12, which had the strongest negative effect on rump fat, had as strong a positive effect on marbling as haplotype 27; the minor difference is not significant. So a different but overlapping set of haplotypes (Table 7) shows extreme effects for rump fat compared to the haplotypes showing extreme effects for marbling.
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Gene expression:
Two alternative transcripts of RORC were found, one in which a part of intron 1 was spliced between exons 1 and 2 and one in which part of exon 8 appears spliced out. The gene expression analysis applies only to the alternative transcripts that involve the splicing between exons 1 and 2, transcript 1 being the one without the insertion and transcript 2 being the one with the insertion. Transcript 1 (DQ667051) was similar to bovine isoform A transcript variants 1 and 3 as well as human and mouse sequences (XM_605862.2, XM_871532.1, NM_06050.2, NM_011281.1). RORC transcript 2 (DQ667052) contains a 38-bp part of intron 1 inserted between exons 1 and 2 and was also in GenBank as unknown transcript BC103029.1. The translation of BC103029.1 reports that the amino acid sequence after the insertion was the same as the normal RORC sequence and that it was the initial amino acid sequence that differed; from amino acid 14 of transcript 1 and amino acid 26 of BC103029.1, and also transcript 2, the amino acid sequences were identical. The translation of BC103029.1 in GenBank appeared to start from a different site. The area of similarity between the transcripts included both the DNA-binding and the ligand-binding domains, so transcript 2 should be functional.
An alternative splicing involving exon 8 was found when sequencing the cDNA. Different-sized bands were cloned and when they were sequenced some of these were found to be missing 36 bp of exon 8. This gap was found both in the 3'-UTR fragment identified by RACE and in the exon 3–exon 8 fragment amplified by PCR. There was RORC sequence on both sides of the gap in both fragments, and the missing sequence was confirmed by directly sequencing the cDNA of the individual. The missing exon 8 RORC transcripts were found in a crossbred Bos taurus taurus x B. t. indicus animal, unlike the line-bred Hereford (B. t. taurus) animal in GenBank. The genomic DNA of the individual did not have the gap, indicating that the gap is not a deletion of part of exon 8. The closest SNP to this gap was RORC:g.3984A>G, and the individual did not possess any additional DNA sequence variants in the region.
The BLAST sequence similarity was 91% to humans and 85% to mice, not including the differences due to possible splicing events. Finally, there was a major dissimilarity of the cow sequence to the mouse sequence between bases 672 and 825 of NM_011281.1.
The highest transcription of either RORC transcript 1 or RORC transcript 2 was in fetal tissues, where the highest fetal expression (fetal kidney) was 40 times greater than the highest adult expression (LD muscle) (Figure 1). RORC transcript 1 was usually expressed at least twofold higher than transcript 2 in six of the eight fetal tissues and in 7 of the 15 adult tissues. In those tissues in which the expression was approximately equal, the level of expression was generally low. Transcript 2 was more highly expressed than transcript 1 in fetal mammary gland and in adult LD fat. In LD muscle, the site of the marbling measurement, the LD muscle had higher expression of transcript 1 while the intramuscular fat tissue had higher expression of transcript 2. However, tissues rich in fat did not all have equal or similar expression of the two transcripts, and two fat tissues, adult cardiac and subcutaneous fat, had higher expression of transcript 1 than transcript 2. All skeletal muscles did not show high expression of RORC transcripts.
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DISCUSSION |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONACKNOWLEDGEMENTSLITERATURE CITED |
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0.36 p for marbling and 0.31 p for fat depth (CASAS et al. 2003). The inferred RORC haplotypes reported here show a maximum difference of 0.35 p for marbling and 0.28 p for rump fat thickness, suggesting that the haplotypes explain most of the QTL effect. The rump fat thickness explained by RORC is only 92% of the fat thickness for the QTL (CASAS et al. 2003). This discrepancy may be due to the two subcutaneous fat depots being in different locations in the animal or it may merely be due to random differences in the two experiments. These effects of the RORC SNPs are slightly larger than those due to the TG:g.-537C>T (TG5) or GH1:c.457C>G SNPs measured in the same population (BARENDSE et al. 2004, 2006). Nevertheless, since SNPs were not sampled in the promoter of the gene, the presence of additional causative mutations cannot be excluded in this analysis. The additive effect of this gene on marbling is small, and most of the effect is due to dominant and even overdominant effects. All of the SNPs show strong dominant or overdominant effects on marbling, and this is so strong that it can be seen by inspection in Table 5, where the raw distribution shows a decrease in the higher marbling scores for the heterozygote genotypes. However, the effect on rump fat thickness is additive, and neither RORC:g.3290T>G nor RORC:g.3984A>G have k-values above an absolute value of 0.5. This suggests that if the SNPs in this gene affect the distribution of fat in different fat depots, then the shift is not specifically from intramuscular to subcutaneous, but must involve other fat depots in the animal.
These effects could be due to more than one causative mutation. First, the RORC:g.3290T>G SNP has the strongest association to marbling, while its neighbor RORC:g.3984A>G SNP has the strongest association to rump fat. Second, while all the common haplotypes for one allele show an increase in marbling, the haplotypes for the other allele show contradictory differences in marbling score. It could be that not all the mutations affecting fatness have been identified in this region, such as in the promoter that could not be sampled, or it could be that different mutations affect relative intramuscular compared to rump fatness. It is unlikely to be due to background genetic effects in response to genetic variation at RORC since there is no breed x haplotype interaction.
The QTL for fatness appear to be located near the ligand-binding domain of RORC. Single-point associations between the RORC SNP and marbling or rump fat show that only some of these SNPs are associated with the trait, that only those near the ligand-binding domain show consistent associations, and that the sublocalization of the QTL effect to the ligand-binding domain is confirmed by the Haploblock analysis. Allele substitution at these SNPs increases marbling and decreases rump fat, reshaping fat depots. Although none of the SNPs obtained to date would appear to cause an obvious functional change, the splicing of exons 7–10 of the RORC ligand-binding domain is complex. Results presented here show that some of the transcripts differ for part of exon 8, but in humans, transcripts have been reported in which all of the intronic material between exons 7 and 10 has been transcribed, which would expose these SNPs as possible amino acid changes (see GenBank Gene ID 6097 Evidence Viewer and cDNA sequence AK128522). The SNPs that would be most affected by such alterations in transcription are RORC:g.3984A>G and RORC:g.3290T>G, which are also those that show the strongest associations to marbling and rump fat thickness. Clearly, then, it will be necessary to search for other transcripts in a wide range of different life stages of the animal to determine the full range of alternative transcripts in cattle.
In livestock, the examples where fine mapping has been used to map QTL have been for QTL with effects much >0.5 p (GRISART et al. 2002; VAN LAERE et al. 2003). Such fine mapping has not been attempted for smaller QTL as the high linkage disequilibrium (FARNIR et al. 2000; MCRAE et al. 2002; TENESA et al. 2003; NSENGIMANA et al. 2004) in livestock was thought to act as a barrier to genetic resolution (BARENDSE 2005). However, instead of working with family groups in experimental populations, the cattle in this study were chosen at random from a large potential genetic base in feedlots, thereby allowing a much higher genetic resolution to be achieved (BARENDSE and FRIES 1999). The D' values showed high levels of linkage disequilibrium, probably due to bottlenecks from domestication and breed formation. Crucially, however, the r2 values, which provide information on the ability to generate high-resolution maps in a particular sequence (DEVLIN and RISCH 1995; DE BAKKER et al. 2005; TERWILLIGER and HIEKKALINNA 2006), were low. The QTL map resolution reported here was comparable to that found in undomesticated species.
The high levels of expression of RORC in cattle skeletal muscle are consistent with that previously reported (HIROSE et al. 1994) for a commercial panel of human tissues and the strong expression in fetal cattle for liver and kidney is consistent with that reported for wild-type fetal mice (KUREBAYASHI et al. 2000). Nevertheless, this is the first time that both fetal and adult tissues were compared in the same experiment and where a range of fetal tissues was reported. These similarities suggest that this switchover from fetal to adult patterns of gene expression is a generic feature of the gene. Clearly, the high levels of expression during development are consistent with an important role of the gene in fetal development, but its altered pattern and continued expression in adult animals indicate that it may affect different cellular pathways or have a different function later in life. Retinoic acid receptors are well known to be involved in the formation of tissue during fetal development, and the role of this gene in the fetal development of lymph nodes is well established. However, its role later in life is less clear. One possible role is in fatness in cattle as reported here, a role that is consistent with the impact of two of its ligands, vitamin A and thyroid hormone, on fatness in cattle (cf. above).
One hypothesis for further testing is that RORC may be involved in the maintenance of adipocytes and the ability to process glucose. First, the pattern of gene expression is consistent with the known development of marbling, namely that starting values of intramuscular fat levels after birth correlate well with final marbling scores (PETHICK et al. 2004, 2005), which implies that it is gene expression early in life that sets up an animal to have high marbling scores. Second, the development of intramuscular fat depots appears to be driven by excess glucose in the diet and marbling appears late in adulthood (PETHICK et al. 2004). Third, QTL for blood glucose levels in humans have been located to the homologous chromosome (MEIGS et al. 2002) as well as QTL for fatness (PERUSSE et al. 2001; NORRIS et al. 2005) and diabetes, but this gene has not itself been tested for association or function in either human obesity or glucose levels; it does not show an association to type 2 diabetes in a case control study of diabetic individuals (WANG et al. 2003). Fourth, RORC appears to play a role in the modulation of the later stages of adipocyte differentiation (AUSTIN et al. 1998). Finally, two important tissues that have a metabolic role to play in what happens to excess glucose are the liver and skeletal muscle (KAHN and ROSSETTI 1998), tissues that show strong RORC gene expression. These findings are consistent with the hypothesis of RORC affecting the development of adipocytes and the ability to process glucose to fat, which could be tested by further functional studies.
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ACKNOWLEDGEMENTS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONACKNOWLEDGEMENTSLITERATURE CITED |
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FOOTNOTES |
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LITERATURE CITED |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONACKNOWLEDGEMENTSLITERATURE CITED |
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