Hi-Tag: a simple and efficient method for identifying protein-mediated long-range chromatin interactions with low cell numbers.

Protein-mediated chromatin interactions can be revealed by coupling proximity-based ligation with chromatin immunoprecipitation. However, these techniques require complex experimental procedures and millions of cells per experiment, which limits their widespread application in life science research. Here, we develop a novel method, Hi-Tag, that identifies high-resolution, long-range chromatin interactions through transposase tagmentation and chromatin proximity ligation (with a phosphorothioate-modified linker). Hi-Tag can be implemented using as few as 100,000 cells, involving simple experimental procedures that can be completed within 1.5 days. Meanwhile, Hi-Tag is capable of using its own data to identify the binding sites of specific proteins, based on which, it can acquire accurate interaction information. Our results suggest that Hi-Tag has great potential for advancing chromatin interaction studies, particularly in the context of limited cell availability.

Single cell multiomic analysis reveals diabetes-associated ß-cell heterogeneity driven by HNF1A.

Broad heterogeneity in pancreatic ß-cell function and morphology has been widely reported. However, determining which components of this cellular heterogeneity serve a diabetes-relevant function remains challenging. Here, we integrate single-cell transcriptome, single-nuclei chromatin accessibility, and cell-type specific 3D genome profiles from human islets and identify Type II Diabetes (T2D)-associated ß-cell heterogeneity at both transcriptomic and epigenomic levels. We develop a computational method to explicitly dissect the intra-donor and inter-donor heterogeneity between single ß-cells, which reflect distinct mechanisms of T2D pathogenesis. Integrative transcriptomic and epigenomic analysis identifies HNF1A as a principal driver of intra-donor heterogeneity between ß-cells from the same donors; HNF1A expression is also reduced in ß-cells from T2D donors. Interestingly, HNF1A activity in single ß-cells is significantly associated with lower Na+ currents and we nominate a HNF1A target, FXYD2, as the primary mitigator. Our study demonstrates the value of investigating disease-associated single-cell heterogeneity and provides new insights into the pathogenesis of T2D.

A compendium and comparative epigenomics analysis of cis-regulatory elements in the pig genome.

Although major advances in genomics have initiated an exciting new era of research, a lack of information regarding cis-regulatory elements has limited the genetic improvement or manipulation of pigs as a meat source and biomedical model. Here, we systematically characterize cis-regulatory elements and their functions in 12 diverse tissues from four pig breeds by adopting similar strategies as the ENCODE and Roadmap Epigenomics projects, which include RNA-seq, ATAC-seq, and ChIP-seq. In total, we generate 199 datasets and identify more than 220,000 cis-regulatory elements in the pig genome. Surprisingly, we find higher conservation of cis-regulatory elements between human and pig genomes than those between human and mouse genomes. Furthermore, the differences of topologically associating domains between the pig and human genomes are associated with morphological evolution of the head and face. Beyond generating a major new benchmark resource for pig epigenetics, our study provides basic comparative epigenetic data relevant to using pigs as models in human biomedical research.

Selective sweep analysis reveals extensive parallel selection traits between large white and Duroc pigs.

In the process of pig genetic improvement, different commercial breeds have been bred for the same purpose, improving meat production. Most of the economic traits, such as growth and fertility, have been selected similarly despite the discrepant selection pressure, which is known as parallel selection. Here, 28 whole-genome sequencing data of Danish large white pigs with an approximately 25-fold depth each were generated, resulting in about 12 million high-quality SNPs for each individual. Combined with the sequencing data of 27 Duroc and 23 European wild boars, we investigated the parallel selection of Danish large white and Duroc pigs using two complementary methods, Fst and iHS. In total, 67 candidate regions were identified as the signatures of parallel selection. The genes in candidate regions of parallel selection were mainly associated with sensory perception, growth rate, and body size. Further functional annotation suggested that the striking consistency of the terms may be caused by the polygenetic basis of quantitative traits, and revealing the complex genetic basis of parallel selection. Besides, some unique terms were enriched in population-specific selection regions, such as the limb development-related terms enriched in Duroc-specific selection regions, suggesting unique selections of breed-specific selected traits. These results will help us better understand the parallel selection process of different breeds. Moreover, we identified several potential causal SNPs that may contribute to the pig genetic breeding process.

Genome-Wide Patterns of Homozygosity and Relevant Characterizations on the Population Structure in Piétrain Pigs.

Investigating the patterns of homozygosity, linkage disequilibrium, effective population size and inbreeding coefficients in livestock contributes to our understanding of the genetic diversity and evolutionary history. Here we used Illumina PorcineSNP50 Bead Chip to identify the runs of homozygosity (ROH) and estimate the linkage disequilibrium (LD) across the whole genome, and then predict the effective population size. In addition, we calculated the inbreeding coefficients based on ROH in 305 Piétrain pigs and compared its effect with the other two types of inbreeding coefficients obtained by different calculation methods. A total of 23,434 ROHs were detected, and the average length of ROH per individual was about 507.27 Mb. There was no regularity on how those runs of homozygosity distributed in genome. The comparisons of different categories suggested that the formation of long ROH was probably related with recent inbreeding events. Although the density of genes located in ROH core regions is lower than that in the other genomic regions, most of them are related with Piétrain commercial traits like meat qualities. Overall, the results provide insight into the way in which ROH is produced and the identified ROH core regions can be used to map the genes associated with commercial traits in domestic animals.

The DNA Methylation Status of Wnt and Tgfß Signals Is a Key Factor on Functional Regulation of Skeletal Muscle Satellite Cell Development.

DNA methylation is an important form of epigenetic regulation that can regulate the expression of genes and the development of tissues. Muscle satellite cells play an important role in skeletal muscle development and regeneration. Therefore, the DNA methylation status of genes in satellite cells is important in the regulation of the development of skeletal muscle. This study systematically investigated the changes of genome-wide DNA methylation in satellite cells during skeletal muscle development. According to the MeDIP-Seq data, 52,809-123,317 peaks were obtained for each sample, covering 0.70-1.79% of the genome. The number of reads and peaks was highest in the intron regions followed by the CDS regions. A total of 96,609 DMRs were identified between any two time points. Among them 6198 DMRs were annotated into the gene promoter regions, corresponding to 4726 DMGs. By combining the MeDIP-Seq and RNA-Seq data, a total of 202 overlap genes were obtained between DMGs and DEGs. GO and Pathway analysis revealed that the overlap genes were mainly involved in 128 biological processes and 23 pathways. Among the biological processes, terms related to regulation of cell proliferation and Wnt signaling pathway were significantly different. Gene-gene interaction analysis showed that Wnt5a, Wnt9a, and Tgfß1 were the key nodes in the network. Furthermore, the expression level of Wnt5a, Wnt9a, and Tgfß1 genes could be influenced by the methylation status of promoter region during skeletal muscle development. These results indicated that the Wnt and Tgfß signaling pathways may play an important role in functional regulation of satellite cells, and the DNA methylation status of Wnt and Tgfß signals is a key regulatory factor during skeletal muscle development. This study provided new insights into the effects of genome-wide methylation on the function of satellite cells.

Identifying Selection Signatures for Backfat Thickness in Yorkshire Pigs Highlights New Regions Affecting Fat Metabolism.

Identifying the genetic basis of improvement in pigs contributes to our understanding of the role of artificial selection in shaping the genome. Here we employed the Cross Population Extended Haplotype Homozogysity (XPEHH) and the Wright's fixation index (FST) methods to detect trait-specific selection signatures by making phenotypic gradient differential population pairs, and then attempted to map functional genes of six backfat thickness traits in Yorkshire pigs. The results indicate that a total of 283 and 466 single nucleotide polymorphisms (SNPs) were identified as trait-specific selection signatures using FST and XPEHH, respectively. Functional annotation suggested that the genes overlapping with the trait-specific selection signatures such as OSBPL8, ASAH2, SMCO2, GBE1, and ABL1 are responsible for the phenotypes including fat metabolism, lean body mass and fat deposition, and transport in mouse. Overall, the study developed the methods of gene mapping on the basis of identification of selection signatures. The candidate genes putatively associated with backfat thickness traits can provide important references and fundamental information for future pig-breeding programs.

Genomic Analysis To Identify Signatures of Artificial Selection and Loci Associated with Important Economic Traits in Duroc Pigs.

Identifying genetic basis of domestication and improvement in livestock contributes to our understanding of the role of artificial selection in shaping the genome. Here we used whole-genome sequencing and the genotyping by sequencing approach to detect artificial selection signatures and identify the associated SNPs of two economic traits in Duroc pigs. A total of 38 candidate selection regions were detected by combining the fixation index and the Composite Likelihood Ratio methods. Further genome-wide association study revealed seven associated SNPs that were related with intramuscular fat content and feed conversion ratio traits, respectively. Enrichment analysis suggested that the artificial selection regions harbored genes, such as MSTN, SOD2, MC5R and CD83, which are responsible for economic traits including lean muscle mass, fertility and immunization. Overall, this study found a series of candidate genes putatively associated with the breeding improvement of Duroc pigs and the polygenic basis of adaptive evolution, which can provide important references and fundamental information for future breeding programs.