Porcine EEF1A1 and EEF1A2 genes: genomic structure, polymorphism, mapping and expression.

Eukaryotic translation elongation factor 1 alpha (EEF1A) plays a key role in protein synthesis. In higher vertebrates EEF1A occurs in two isoforms, EEF1A1 and EEF1A2, encoded by distinct genes. The purpose of this study was to compare the two porcine genes as for the genomic sequence, gene organization and mRNA expression in different tissues, as well as to search for polymorphism and chromosomal assignment. Standard methods of DNA and mRNA analysis were used. We determined the complete genomic sequence of the porcine EEF1A1 and EEF1A2 genes. The two genes differ in the lengths of transcription units (3102 and 8588 bp, respectively), but have similar genomic organization and their coding sequences are highly similar (78% identity of coding sequences and 92.4% identity of amino acid sequences). Several polymorphisms in the two genes were detected. EEF1A1 and EEF1A2 were mapped to SSC1p11.1 and SSC17q23.3, respectively. mRNA of EEF1A1 was expressed in all studied tissues (the highest expression was in 44-day fetal muscle and low expression in adult liver and brain), while EEF1A2 was expressed only in skeletal-muscle, tongue, heart, diaphragm and brain tissues. EEF1A2 was not expressed in fetal muscle tissue (44 days). In this paper results are provided on genomic sequences, genomic organization, polymorphism, chromosomal assignment and spatial and temporal expressions of the porcine EEF1A1 and EEF1A2 genes. Novel polymorphisms were described in both genes. Porcine EEF1A2 was studied for the first time.

Porcine ubiquitin-like 5 (UBL5) gene: genomic organization, polymorphisms, mRNA cloning, splicing variants and association study.

Ubiquitin-like 5 (UBL5), which is supposed to be involved in regulation of feed intake, energy metabolism, obesity and type 2 diabetes, is located at position 62.1 cM on the pig chromosome 2 region harbouring quantitative trait loci for carcass and meat quality. The 4,354 bp genomic sequence (FR798948) of the porcine gene encompassing the promoter and entire gene was cloned by polymerase chain reaction. Comparative sequencing revealed 13 polymorphisms in noncoding regions. Synthesis of full-length cDNA sequences using rapid amplification of 5' and 3' ends showed three splice variants. Variants 1 and 2 differ in transcription length for the untranslated part of exon 1 with deduced protein of 73 amino acid (aa) residues and 100 % identities between human, mouse and other species. Variant 3, with 4 bp deletion at the 3' end of exon 2, encodes a truncated protein with 28 aa residues. In a Wild boar×Meishan F2 population (n = 334) with 47 recorded traits, loci FR798948:g.2788G>A and FR798948:g.2141T>C were associated at nominal P < 0.05 with fat deposition, growth and fattening and muscling but after adjustment for multiple testing (Benjamini and Hochberg, J R Stat Soc B 57:289-300, 1995) only eight fat deposition traits showed suggestive association with FR798948:g.2788G>A at adjusted P < 0.10. In a Meishan×Large White (MLW) cross (n = 562) with six trait records available, FR798948:g.2141T>C showed suggestive association with growth (adjusted P = 0.0690). As association mapping conducted in the outbred MLW population is more precise than in the three generation F2 population the UBL5 gene tends to be associated with growth rather than with fat accretion.

Association between polymorphism in the FTO gene and growth and carcass traits in pig crosses.

BACKGROUND: Independent studies have shown that several single nucleotide polymorphisms (SNP) in the human FTO (fat mass and obesity associated) gene are associated with obesity. SNP have also been identified in the pig FTO gene, among which some are associated with selected fat-deposition traits in F2 crosses and commercial populations. In this study, using both commercial pig populations and an experimental Meishan × Pietrain F2 population, we have investigated the association between one FTO SNP and several growth and carcass traits. Association analyses were performed with the FTO polymorphism either alone or in combination with polymorphisms in flanking loci. METHODS: SNP (FM244720:g.400C>G) in exon 3 of porcine FTO was genotyped by PCR-RFLP and tested for associations with some growth, carcass and fat-related traits. Proportions of genetic variance of four pig chromosome 6 genes (FTO, RYR1, LIPE and TGFB1) on selected traits were evaluated using single- and multi-locus models. RESULTS: Linkage analysis placed FTO on the p arm of pig chromosome 6, approximately 22 cM from RYR1. In the commercial populations, allele C of the FTO SNP was significantly associated with back fat depth and allele G with muscling traits. In the Meishan × Pietrain F2 pigs, heterozygotes with allele C from the Pietrain sows and allele G from the Meishan boar were more significantly associated with fat-related traits compared to homozygotes with allele G from the Pietrain and allele G from the Meishan breed. In single- and multi-locus models, genes RYR1, TGFB1 and FTO showed high associations. The contribution in genetic variance from the polymorphism in the FTO gene was highest for back fat depth, meat area on the musculus longissimus lumborum et thoracis tissues and metabolite glucose-6-phosphate dehydrogenase. CONCLUSIONS: Our results show that in pig, FTO influences back fat depth in the commercial populations, while in the Meishan × Pietrain F2 pigs with a CG genotype, heterosis occurs for several fat-related traits.

Porcine insulin receptor substrate 4 (IRS4) gene: cloning, polymorphism and association study.

Using PCR and inverse PCR techniques we obtained a 4,498 bp nucleotide sequence FN424076 encompassing the complete coding sequence of the porcine insulin receptor substrate 4 (IRS4) gene and its proximal promoter. The 1,269 amino acid porcine protein deduced from the nucleotide sequence shares 92% identity with the human IRS4 and possesses the same domains and the same number of tyrosine phosphorylation motifs as the human protein. We detected substitution FN424076:g.96C

Genome-wide mapping of quantitative trait loci for fatness, fat cell characteristics and fat metabolism in three porcine F2 crosses.

BACKGROUND: QTL affecting fat deposition related performance traits have been considered in several studies and mapped on numerous porcine chromosomes. However, activity of specific enzymes, protein content and cell structure in fat tissue probably depend on a smaller number of genes than traits related to fat content in carcass. Thus, in this work traits related to metabolic and cytological features of back fat tissue and fat related performance traits were investigated in a genome-wide QTL analysis. QTL similarities and differences were examined between three F2 crosses, and between male and female animals. METHODS: A total of 966 F2 animals originating from crosses between Meishan (M), Pietrain (P) and European wild boar (W) were analysed for traits related to fat performance (11), enzymatic activity (9) and number and volume of fat cells (20). Per cross, 216 (MxP), 169 (WxP) and 195 (WxM) genome-wide distributed marker loci were genotyped. QTL mapping was performed separately for each cross in steps of 1 cM and steps were reduced when the distance between loci was shorter. The additive and dominant components of QTL positions were detected stepwise by using a multiple position model. RESULTS: A total of 147 genome-wide significant QTL (76 at P<0.05 and 71 at P<0.01) were detected for the three crosses. Most of the QTL were identified on SSC1 (between 76-78 and 87-90 cM), SSC7 (predominantly in the MHC region) and SSCX (in the vicinity of the gene CAPN6). Additional genome-wide significant QTL were found on SSC8, 12, 13, 14, 16, and 18. In many cases, the QTL are mainly additive and differ between F2 crosses. Many of the QTL profiles possess multiple peaks especially in regions with a high marker density. Sex specific analyses, performed for example on SSC6, SSC7 and SSCX, show that for some traits the positions differ between male and female animals. For the selected traits, the additive and dominant components that were analysed for QTL positions on different chromosomes, explain in combination up to 23% of the total trait variance. CONCLUSIONS: Our results reveal specific and partly new QTL positions across genetically diverse pig crosses. For some of the traits associated with specific enzymes, protein content and cell structure in fat tissue, it is the first time that they are included in a QTL analysis. They provide large-scale information to analyse causative genes and useful data for the pig industry.

Polymorphic microsatellite sites in the PRNP region point to excess of homozygotes in Creutzfeldt-Jakob disease patients.

Polymorphic microsatellite sites within 148 kb of the human prion gene complex, including the genes PRNP, PRND and PRNT, were analysed together with the Codon129 variants regarding 50 CJD (Creutzfeldt-Jakob Disease) patients and 46 non-diseased control persons. Three of the sites (MM03, MM04, Codon129) differed significantly (P<0.05) for their allele frequencies between the two groups--the predominant allele being always more frequent in the CJD group. Deviations from Hardy-Weinberg Equilibrium were mainly obtained in the CJD group--in all cases with a reduction of the observed heterozygosity. The sites MM03, MM04 and Codon129 were also analysed for their haplotypes. The predominant homozygous haplotype combination was more frequently observed in the CJD group (0.875) than in the non-diseased group (0.38). Thus the different polymorphic sites indicate that high CJD disposition is associated with homozygosity in the PRNP gene.

Comparative and genetic analysis of the porcine glucocerebrosidase (GBA) gene.

The genomic sequence of the porcine (Sus scrofa) glucocerebrosidase (GBA) gene (approximately 5.7 kb), encoding glucocerebrosidase (glucosylceramidase; acid beta-glucosidase; EC 3.2.1.45), was determined and compared with human (Homo sapiens) GBA and GBAP (pseudogene). The porcine gene harbours 11 exons and 10 introns, and the genomic organization is identical with human GBA. The exon sequences, coding for signal peptide and mature protein, show 81% and 90% sequence identity, respectively, with the corresponding human GBA sequences. Short interspersed elements, SINEs (PREs), are present in introns 2, 4 and 7. There is no evidence of a pseudogene in pig. The deduced protein sequence of GBA consists of 39 amino acids of signal peptide (long form) and 497 amino acids of the mature protein; the latter shows 90% sequence identity with the human protein. Four polymorphisms were observed within the porcine gene: insertion/deletion of one of the two SINEs (PREs) in intron 2 (locus PREA); deletion of a 37- to 39-bp stretch in intron 4 (one direct repeat and 5' end of PRE); deletion of a 47-bp stretch in the middle part of PRE in intron 4 (locus PREB); and single-base transition (C-T) in intron 6 (locus HaeIII-RFLP). GBA was assigned to chromosome 4q21 by FISH and was localized to the same region by linkage analysis and RH mapping, i.e., to the chromosome 4 segment where quantitative trait loci for growth and some carcass traits are located.

A distinctive gene expression fingerprint in mentally retarded male patients reflects disease-causing defects in the histone demethylase KDM5C.

BACKGROUND: Mental retardation is a genetically heterogeneous disorder, as more than 90 genes for this disorder has been found on the X chromosome alone. In addition the majority of patients are non-syndromic in that they do not present with clinically recognisable features. This makes it difficult to determine the molecular cause of this disorder on the basis of the phenotype alone. Mutations in KDM5C (previously named SMCX or JARID1C), a gene that encodes a transcriptional regulator with histone demethylase activity specific for dimethylated and trimethylated H3K4, are a comparatively frequent cause of non-syndromic X-linked mental retardation (NS-XLMR). Specific transcriptional targets of KDM5C, however, are still unknown and the effects of KDM5C deficiency on gene expression have not yet been investigated. RESULTS: By whole-mount in situ hybridisation we showed that the mouse homologue of KDM5C is expressed in multiple tissues during mouse development.We present the results of gene expression profiling performed on lymphoblastoid cell lines as well as blood from patients with mutations in KDM5C. Using whole genome expression arrays and quantitative reverse transcriptase polymerase chain reaction (QRT-PCR) experiments, we identified several genes, including CMKOR1, KDM5B and KIAA0469 that were consistently deregulated in both tissues. CONCLUSIONS: Our findings shed light on the pathological mechanisms underlying mental retardation and have implications for future diagnostics of this heterogeneous disorder.

OLA-DRB1 microsatellite variants are associated with ovine growth and reproduction traits.

The DRB1 intron 2 (GT)(n)(GA)(m) microsatellite was genotyped in experimental flocks of seven Merinoland rams and 249 ewes as well as their offspring (381 lambs) from consecutive lambings. A total of 16 DRB1 alleles were detected, ranging between 353 and 857 bp. In comparison with carriers of other alleles, the ewes carrying the predominant 411 bp allele had higher values of all the recorded fertility traits. For ewes carrying the 394 and 857 bp alleles, the birth weight of lambs was about 400 g higher as compared to the residual group of ewes. The observed associations could be due to differences in disease resistance, cell recognition or tissue differentiation between carriers of various MHC haplotypes which can in turn affect individual fertility and growth performance.