scholarly journals Harnessing natural variation to identify cis regulators of sex-biased gene expression in a multi-strain mouse liver model

2021 ◽  
Author(s):  
Bryan J. Matthews ◽  
David J Waxman
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Sex Differences ◽  
Genetic Variants ◽  
Mouse Liver ◽  
Regulatory Elements ◽  
Individual Variability ◽  
Sex Bias ◽  
Mouse Strains ◽  
Strain Dependent

Sex differences in gene expression are widespread in the liver, where a large number of autosomal factors act in tandem with growth hormone signaling to regulate individual variability of sex differences in liver metabolism and disease. Here, we compare hepatic transcriptomic and epigenetic profiles of mouse strains C57Bl/6J and CAST/EiJ, representing two subspecies separated by 0.5-1 million years of evolution, to elucidate the actions of genetic factors regulating liver sex differences. We identify 144 protein coding genes and 78 lncRNAs showing strain-conserved sex bias; many have gene ontologies relevant to liver function, are more highly liver-specific and show greater sex bias, and are more proximally regulated than genes whose sex bias is strain-dependent. The strain-conserved genes include key growth hormone-dependent transcriptional regulators of liver sex bias; however, three other transcription factors, Trim24 , Tox , and Zfp809, lose sex-biased expression in CAST/EiJ mouse liver. To elucidate these strain specificities in expression, we characterized the strain-dependence of sex-biased chromatin opening and enhancer marks at cis regulatory elements (CREs) within expression quantitative trait loci (eQTL) regulating liver sex-biased genes. Strikingly, 208 of 286 eQTLs with strain-specific, sex-differential effects on expression were associated with a complete gain, loss, or reversal of expression sex differences between strains. Moreover, 166 of the 286 eQTLs were linked to the strain-specific gain or loss of localized sex-biased CREs. Remarkably, a subset of these CREs lacked strain-specific genetic variants yet showed coordinated, strain-dependent sex-biased epigenetic regulation. Thus, we directly link hundreds of strain-specific genetic variants to the high variability in CRE activity and expression of sex-biased genes, and uncover underlying genetically-determined epigenetic states controlling liver sex bias in genetically diverse mouse populations.

scholarly journals Harnessing natural variation to identify cis regulators of sex-biased gene expression in a multi-strain mouse liver model

PLoS Genetics ◽  
2021 ◽  
Vol 17 (11) ◽  
pp. e1009588
Author(s):  
Bryan J. Matthews ◽  
Tisha Melia ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Sex Differences ◽  
Genetic Variants ◽  
Mouse Liver ◽  
Regulatory Elements ◽  
Individual Variability ◽  
Sex Bias ◽  
Mouse Strains ◽  
Strain Dependent

Sex differences in gene expression are widespread in the liver, where many autosomal factors act in tandem with growth hormone signaling to regulate individual variability of sex differences in liver metabolism and disease. Here, we compare hepatic transcriptomic and epigenetic profiles of mouse strains C57BL/6J and CAST/EiJ, representing two subspecies separated by 0.5–1 million years of evolution, to elucidate the actions of genetic factors regulating liver sex differences. We identify 144 protein coding genes and 78 lncRNAs showing strain-conserved sex bias; many have gene ontologies relevant to liver function, are more highly liver-specific and show greater sex bias, and are more proximally regulated than genes whose sex bias is strain-dependent. The strain-conserved genes include key growth hormone-dependent transcriptional regulators of liver sex bias; however, three other transcription factors, Trim24, Tox, and Zfp809, lose their sex-biased expression in CAST/EiJ mouse liver. To elucidate the observed strain specificities in expression, we characterized the strain-dependence of sex-biased chromatin opening and enhancer marks at cis regulatory elements (CREs) within expression quantitative trait loci (eQTL) regulating liver sex-biased genes. Strikingly, 208 of 286 eQTLs with strain-specific, sex-differential effects on expression were associated with a complete gain, loss, or reversal of the sex differences in expression between strains. Moreover, 166 of the 286 eQTLs were linked to the strain-dependent gain or loss of localized sex-biased CREs. Remarkably, a subset of these CREs apparently lacked strain-specific genetic variants yet showed coordinated, strain-dependent sex-biased epigenetic regulation. Thus, we directly link hundreds of strain-specific genetic variants to the high variability in CRE activity and expression of sex-biased genes and uncover underlying genetically-determined epigenetic states controlling liver sex bias in genetically diverse mouse populations.

scholarly journals STAT5 regulation of sex-dependent hepatic CpG methylation at distal regulatory elements mapping to sex-biased genes

2020 ◽  
Author(s):  
Pengying Hao ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Sex Differences ◽  
Mouse Liver ◽  
Regulatory Elements ◽  
Cpg Methylation ◽  
Sex Bias ◽  
Liver Chromatin ◽  
Differential Gene ◽  
Genomic Regions ◽  
Distal Regulatory Elements

Growth hormone-activated STAT5b is an essential regulator of sex-differential gene expression in mouse liver, however, its impact on hepatic gene expression and epigenetic responses is poorly understood. Here, we found a substantial, albeit incomplete loss of liver sex bias in hepatocyte-specific STAT5a/STAT5b (collectively, STAT5)-deficient mouse liver. In male liver, many male-biased genes were down regulated in direct association with the loss of STAT5 binding; many female-biased genes, which show low STAT5 binding, were de-repressed, indicating an indirect mechanism for repression by STAT5. Extensive changes in CpG-methylation were seen in STAT5-deficient liver, where sex differences were abolished at 88% of ∼1,500 sex-differentially methylated regions, largely due to increased DNA methylation upon STAT5 loss. STAT5-dependent CpG-hypomethylation was rarely found at proximal promoters of STAT5-dependent genes. Rather, STAT5 primarily regulated the methylation of distal enhancers, where STAT5 deficiency induced widespread hypermethylation at genomic regions enriched for accessible chromatin, enhancer histone marks (H3K4me1, H3K27ac), STAT5 binding, and DNA motifs for STAT5 and other transcription factors implicated in liver sex differences. Thus, the sex-dependent binding of STAT5 to liver chromatin is closely linked to the sex-dependent demethylation of distal regulatory elements linked to STAT5-dependent genes important for liver sex bias.

scholarly journals STAT5 regulation of sex-dependent hepatic CpG methylation at distal regulatory elements mapping to sex-biased genes

2020 ◽  
Author(s):  
Pengying Hao ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Sex Differences ◽  
Mouse Liver ◽  
Regulatory Elements ◽  
Cpg Methylation ◽  
Sex Bias ◽  
Liver Chromatin ◽  
Differential Gene ◽  
Genomic Regions ◽  
Distal Regulatory Elements

AbstractGrowth hormone-activated STAT5b is an essential regulator of sex-differential gene expression in mouse liver, however, its impact on hepatic gene expression and epigenetic responses is poorly understood. Here, we found a substantial, albeit incomplete loss of liver sex bias in hepatocyte-specific STAT5a/STAT5b (collectively, STAT5)-deficient mouse liver. In male liver, many male-biased genes were down regulated in direct association with the loss of STAT5 binding; many female-biased genes, which show low STAT5 binding, were de-repressed, indicating an indirect mechanism for repression by STAT5. Extensive changes in CpG-methylation were seen in STAT5-deficient liver, where sex differences in DNA methylation were abolished at 88% of ~1,500 differentially-methylated regions, largely due to an increase in methylation at the hypomethylated sites. STAT5-dependent CpG-hypomethylation was rarely found at proximal promoters of STAT5-dependent genes. Rather, STAT5 primarily regulated the methylation of distal enhancers, where STAT5 deficiency induced widespread hypermethylation at genomic regions enriched for accessible chromatin, enhancer histone marks (H3K4me1, H3K27ac), STAT5 binding, and DNA motifs for STAT5 and other transcription factors implicated in liver sex differences. In conclusion, the sex-dependent binding of STAT5 to liver chromatin is closely linked to sex-dependent demethylation of distal regulatory elements mapping to STAT5-dependent genes important for liver sex bias.

scholarly journals Genetic factors contributing to extensive variability of sex-specific hepatic gene expression in Diversity Outbred mice

2020 ◽  
Author(s):  
Tisha Melia ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Sex Differences ◽  
Genetic Variants ◽  
Genetic Factors ◽  
Disease Susceptibility ◽  
Gene Clusters ◽  
Individual Variability ◽  
Diversity Outbred ◽  
The Individual

AbstractSex-specific transcription characterizes hundreds of genes in mouse liver, many implicated in sex-differential drug and lipid metabolism and disease susceptibility. While the regulation of liver sex differences by growth hormone-activated STAT5 is well established, little is known about autosomal genetic factors regulating the sex-specific liver transcriptome. Here we show, using genotyping and expression data from a large population of Diversity Outbred mice, that genetic factors work in tandem with growth hormone to control the individual variability of hundreds of sex-biased genes, including many lncRNA genes. Significant associations between single nucleotide polymorphisms and sex-specific gene expression were identified as expression quantitative trait loci (eQTLs), many of which showed strong sex-dependent associations. Remarkably, autosomal genetic modifiers of sex-specific genes were found to account for more than 200 instances of gain or loss of sex-specificity across eight Diversity Outbred mouse founder strains. Sex-biased STAT5 binding sites and open chromatin regions with strain-specific variants were significantly enriched at eQTL regions regulating correspondingly sex-specific genes, supporting the proposed functional regulatory nature of the eQTL regions identified. Binding of the male-biased, growth hormone-regulated repressor BCL6 was most highly enriched at trans-eQTL regions controlling female-specific genes. Co-regulated gene clusters defined by overlapping eQTLs included sets of highly correlated genes from different chromosomes, further supporting trans-eQTL action. These findings elucidate how an unexpectedly large number of autosomal factors work in tandem with growth hormone signaling pathways to regulate the individual variability associated with sex differences in liver metabolism and disease.Author summaryMale-female differences in liver gene expression confer sex differences in many biological processes relevant to health and disease, including lipid and drug metabolism and liver disease susceptibility. While the role of hormonal factors, most notably growth hormone, in regulating hepatic sex differences is well established, little is known about how autosomal genetic factors impact sex differences on an individual basis. Here, we harness the power of mouse genetics provided by the Diversity Outbred mouse model to discover significant genome-wide associations between genetic variants and sex-specific liver gene expression. Remarkably, we found that autosomal expression quantitative trait loci with a strong sex-bias account for the loss or gain of sex-specific expression of more than 200 autosomal genes seen across eight founder mice strains. Genetic associations with sex-specific genes were enriched for sex-biased and growth hormone-dependent regulatory regions harboring strain-specific genetic variants. Co-regulated gene clusters identified by overlapping regulatory regions included highly correlated genes from different chromosomes. These findings reveal the extensive regulatory role played by autosomal genetic variants, working in tandem with growth hormone signaling pathways, in the transcriptional control of sex-biased genes, many of which have been implicated in sex differential outcomes in liver metabolism and disease susceptibility.

scholarly journals Sex-biased long non-coding RNAs negatively correlated with sex-opposite protein coding gene co-expression networks in Diversity Outbred mouse liver

2018 ◽  
Author(s):  
Tisha Melia ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Sex Differences ◽  
Mouse Liver ◽  
Sex Bias ◽  
Response Class ◽  
Protein Coding ◽  
Diversity Outbred ◽  
Outbred Mouse ◽  
Gene Modules ◽  
Liver Gene

AbstractSex differences in liver gene expression and disease susceptibility are regulated by pituitary growth hormone secretion patterns, which activate sex-dependent liver transcription factors and establish sex-specific chromatin states. Ablation of pituitary hormone by hypophysectomy (hypox) has identified two major classes of liver sex-biased genes, defined by their sex-dependent positive or negative responses to hypox, respectively; however, the mechanisms that determine the hypox responsiveness of each gene class are unknown. Here, we sought to discover candidate regulatory long noncoding RNAs (lncRNAs) that control hypox responsiveness. First, we used mouse liver RNA-seq data for 30 different biological conditions to discover gene structures and expression patterns for ~15,500 liver-expressed lncRNAs, including antisense and intragenic lncRNAs, as well as lncRNAs that overlap active enhancers, marked by enhancer RNAs. We identified >200 robust sex-specific liver lncRNAs, including 157 whose expression is regulated during postnatal liver development or is subject to circadian oscillations. Next, we utilized the high natural allelic variance of Diversity Outbred (DO) mice, a multi-parental outbred population, to discover tightly co-expressed clusters of sex-specific protein-coding genes (gene modules) in male liver, and separately, in female liver. Sex differences in the gene modules identified were extensive. Remarkably, many gene modules were strongly enriched for male-specific or female-specific genes belonging to a single hypox-response classes, indicating that the genetic heterogeneity of DO mice captures responsiveness to hypox. Hypox-responsiveness was shown to be facilitated by multiple, distinct gene regulatory mechanisms, indicating its complex nature. Further, we identified 16 sex-specific lncRNAs whose expression across DO mouse livers showed an unexpected significant negative correlation with protein-coding gene modules enriched for genes of the opposite-sex bias and inverse hypox response class, indicating strong negative regulatory potential for these lncRNAs. Thus, we used a genetically diverse outbred mouse population to discover tightly co-expressed sex-specific gene modules that reveal broad characteristics of gene regulation related to responsiveness to hypox, and generated testable hypotheses for regulatory roles of sex-biased liver lncRNAs that control the sex-bias in liver gene expression.

scholarly journals Genetic factors contributing to extensive variability of sex-specific hepatic gene expression in Diversity Outbred mice

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0242665
Author(s):  
Tisha Melia ◽  
David J. Waxman
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Sex Differences ◽  
Genetic Factors ◽  
Individual Variability ◽  
Specific Gene ◽  
Open Chromatin ◽  
Diversity Outbred ◽  
Outbred Mice ◽  
The Individual

Sex-specific transcription characterizes hundreds of genes in mouse liver, many implicated in sex-differential drug and lipid metabolism and disease susceptibility. While the regulation of liver sex differences by growth hormone-activated STAT5 is well established, little is known about autosomal genetic factors regulating the sex-specific liver transcriptome. Here we show, using genotyping and expression data from a large population of Diversity Outbred mice, that genetic factors work in tandem with growth hormone to control the individual variability of hundreds of sex-biased genes, including many long non-coding RNA genes. Significant associations between single nucleotide polymorphisms and sex-specific gene expression were identified as expression quantitative trait loci (eQTLs), many of which showed strong sex-dependent associations. Remarkably, autosomal genetic modifiers of sex-specific genes were found to account for more than 200 instances of gain or loss of sex-specificity across eight Diversity Outbred mouse founder strains. Sex-biased STAT5 binding sites and open chromatin regions with strain-specific variants were significantly enriched at eQTL regions regulating correspondingly sex-specific genes, supporting the proposed functional regulatory nature of the eQTL regions identified. Binding of the male-biased, growth hormone-regulated repressor BCL6 was most highly enriched at trans-eQTL regions controlling female-specific genes. Co-regulated gene clusters defined by overlapping eQTLs included sets of highly correlated genes from different chromosomes, further supporting trans-eQTL action. These findings elucidate how an unexpectedly large number of autosomal factors work in tandem with growth hormone signaling pathways to regulate the individual variability associated with sex differences in liver metabolism and disease.

scholarly journals Evolution of mouse circadian enhancers from transposable elements

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Julius Judd ◽  
Hayley Sanderson ◽  
Cédric Feschotte
Keyword(s):  
Gene Expression ◽  
Transposable Elements ◽  
Binding Sites ◽  
Mouse Liver ◽  
Integration Site ◽  
Point Mutations ◽  
Regulatory Elements ◽  
Circadian Gene ◽  
Binding Motifs ◽  
Reporter Assays

Abstract Background Transposable elements are increasingly recognized as a source of cis-regulatory variation. Previous studies have revealed that transposons are often bound by transcription factors and some have been co-opted into functional enhancers regulating host gene expression. However, the process by which transposons mature into complex regulatory elements, like enhancers, remains poorly understood. To investigate this process, we examined the contribution of transposons to the cis-regulatory network controlling circadian gene expression in the mouse liver, a well-characterized network serving an important physiological function. Results ChIP-seq analyses reveal that transposons and other repeats contribute ~ 14% of the binding sites for core circadian regulators (CRs) including BMAL1, CLOCK, PER1/2, and CRY1/2, in the mouse liver. RSINE1, an abundant murine-specific SINE, is the only transposon family enriched for CR binding sites across all datasets. Sequence analyses and reporter assays reveal that the circadian regulatory activity of RSINE1 stems from the presence of imperfect CR binding motifs in the ancestral RSINE1 sequence. These motifs matured into canonical motifs through point mutations after transposition. Furthermore, maturation occurred preferentially within elements inserted in the proximity of ancestral CR binding sites. RSINE1 also acquired motifs that recruit nuclear receptors known to cooperate with CRs to regulate circadian gene expression specifically in the liver. Conclusions Our results suggest that the birth of enhancers from transposons is predicated both by the sequence of the transposon and by the cis-regulatory landscape surrounding their genomic integration site.

scholarly journals Tissue-specific sex differences in human gene expression

2019 ◽  
Vol 28 (17) ◽  
pp. 2976-2986 ◽  
Author(s):  
Irfahan Kassam ◽  
Yang Wu ◽  
Jian Yang ◽  
Peter M Visscher ◽  
Allan F McRae
Keyword(s):  
Gene Expression ◽  
Sex Differences ◽  
Complex Traits ◽  
Tissue Expression ◽  
Regulatory Elements ◽  
Tissue Specific ◽  
Significance Threshold ◽  
Expression Of Genes ◽  
Causal Variants ◽  
Similar Distance

Abstract Despite extensive sex differences in human complex traits and disease, the male and female genomes differ only in the sex chromosomes. This implies that most sex-differentiated traits are the result of differences in the expression of genes that are common to both sexes. While sex differences in gene expression have been observed in a range of different tissues, the biological mechanisms for tissue-specific sex differences (TSSDs) in gene expression are not well understood. A total of 30 640 autosomal and 1021 X-linked transcripts were tested for heterogeneity in sex difference effect sizes in n = 617 individuals across 40 tissue types in Genotype–Tissue Expression (GTEx). This identified 65 autosomal and 66 X-linked TSSD transcripts (corresponding to unique genes) at a stringent significance threshold. Results for X-linked TSSD transcripts showed mainly concordant direction of sex differences across tissues and replicate previous findings. Autosomal TSSD transcripts had mainly discordant direction of sex differences across tissues. The top cis-expression quantitative trait loci (eQTLs) across tissues for autosomal TSSD transcripts are located a similar distance away from the nearest androgen and estrogen binding motifs and the nearest enhancer, as compared to cis-eQTLs for transcripts with stable sex differences in gene expression across tissue types. Enhancer regions that overlap top cis-eQTLs for TSSD transcripts, however, were found to be more dispersed across tissues. These observations suggest that androgen and estrogen regulatory elements in a cis region may play a common role in sex differences in gene expression, but TSSD in gene expression may additionally be due to causal variants located in tissue-specific enhancer regions.

scholarly journals Sex Differences in the Epigenome: A Cause or Consequence of Sexual Differentiation of the Brain?

Genes ◽  
2019 ◽  
Vol 10 (6) ◽  
pp. 432 ◽  
Author(s):  
Bruno Gegenhuber ◽  
Jessica Tollkuhn
Keyword(s):  
Gene Expression ◽  
Sex Differences ◽  
Sexual Differentiation ◽  
Hormone Receptors ◽  
Neurological Diseases ◽  
Activity Patterns ◽  
Regulatory Elements ◽  
The Body ◽  
Epigenetic Mechanism ◽  
The Brain

Females and males display differences in neural activity patterns, behavioral responses, and incidence of psychiatric and neurological diseases. Sex differences in the brain appear throughout the animal kingdom and are largely a consequence of the physiological requirements necessary for the distinct roles of the two sexes in reproduction. As with the rest of the body, gonadal steroid hormones act to specify and regulate many of these differences. It is thought that transient hormonal signaling during brain development gives rise to persistent sex differences in gene expression via an epigenetic mechanism, leading to divergent neurodevelopmental trajectories that may underlie sex differences in disease susceptibility. However, few genes with a persistent sex difference in expression have been identified, and only a handful of studies have employed genome-wide approaches to assess sex differences in epigenomic modifications. To date, there are no confirmed examples of gene regulatory elements that direct sex differences in gene expression in the brain. Here, we review foundational studies in this field, describe transcriptional mechanisms that could act downstream of hormone receptors in the brain, and suggest future approaches for identification and validation of sex-typical gene programs. We propose that sexual differentiation of the brain involves self-perpetuating transcriptional states that canalize sex-specific development.

scholarly journals Site-specific methylation of the rat prolactin and growth hormone promoters correlates with gene expression.

1996 ◽  
Vol 16 (7) ◽  
pp. 3245-3254 ◽  
Author(s):  
V Ngô ◽  
D Gourdji ◽  
J N Laverrière
Keyword(s):  
Gene Expression ◽  
Growth Hormone ◽  
Cell Lines ◽  
Anterior Pituitary ◽  
Regulatory Elements ◽  
Specific Expression ◽  
Site Specific ◽  
Cpg Dinucleotides ◽  
Cpg Sites

The methylation patterns of the rat prolactin (rPRL) (positions -440 to -20) and growth hormone (rGH) (positions -360 to -110) promoters were analyzed by bisulfite genomic sequencing. Two normal tissues, the anterior pituitary and the liver, and three rat pituitary GH3 cell lines that differ considerably in their abilities to express both genes were tested. High levels of rPRL gene expression were correlated with hypomethylation of the CpG dinucleotides located at positions -277 and -97, near or within positive cis-acting regulatory elements. For the nine CpG sites analyzed in the rGH promoter, an overall hypomethylation-expression coupling was also observed for the anterior pituitary, the liver, and two of the cell lines. The effect of DNA methylation was tested by measuring the transient expression of the chloramphenicol acetyltransferase reporter gene driven by a regionally methylated rPRL promoter. CpG methylation resulted in a decrease in the activity of the rPRL promoter which was proportional to the number of modified CpG sites. The extent of the inhibition was also found to be dependent on the position of methylated sites. Taken together, these data suggest that site-specific methylation may modulate the action of transcription factors that dictate the tissue-specific expression of the rPRL and rGH genes in vivo.

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