Georg W. Otto

Genome Sequencing Reveals Loci under Artificial Selection that Underlie Disease Phenotypes in the Laboratory Rat

Summary Large numbers of inbred laboratory rat strains have been developed for a range of complex disease phenotypes. To gain insights into the evolutionary pressures underlying selection for these phenotypes, we sequenced the genomes of 27 rat strains, including 11 models of hypertension, diabetes, and insulin resistance, along with their respective control strains. Altogether, we identified more than 13 million single-nucleotide variants, indels, and structural variants across these rat strains. Analysis of strain-specific selective sweeps and gene clusters implicated genes and pathways involved in cation transport, angiotensin production, and regulators of oxidative stress in the development of cardiovascular disease phenotypes in rats. Many of the rat loci that we identified overlap with previously mapped loci for related traits in humans, indicating the presence of shared pathways underlying these phenotypes in rats and humans. These data represent a step change in resources available for evolutionary analysis of complex traits in disease models. PaperClip

Genetically distinct leukemic stem cells in human CD34- acute myeloid leukemia are arrested at a hemopoietic precursor-like stage.

Quek and colleagues identify human leukemic stem cells (LSCs) present in CD34− AML. In-depth characterization of the functional and clonal aspects of CD34− LSCs indicates that most are similar to myeloid precursors.

Causes and consequences of chromatin variation between inbred mice.

Variation at regulatory elements, identified through hypersensitivity to digestion by DNase I, is believed to contribute to variation in complex traits, but the extent and consequences of this variation are poorly characterized. Analysis of terminally differentiated erythroblasts in eight inbred strains of mice identified reproducible variation at approximately 6% of DNase I hypersensitive sites (DHS). Only 30% of such variable DHS contain a sequence variant predictive of site variation. Nevertheless, sequence variants within variable DHS are more likely to be associated with complex traits than those in non-variant DHS, and variants associated with complex traits preferentially occur in variable DHS. Changes at a small proportion (less than 10%) of variable DHS are associated with changes in nearby transcriptional activity. Our results show that whilst DNA sequence variation is not the major determinant of variation in open chromatin, where such variants exist they are likely to be causal for complex traits.

Single-cell analysis reveals the continuum of human lympho-myeloid progenitor cells.

The human hemopoietic progenitor hierarchy producing lymphoid and granulocytic-monocytic (myeloid) lineages is unclear. Multiple progenitor populations produce lymphoid and myeloid cells, but remain incompletely characterized. Here, we demonstrated cord blood lympho-myeloid containing progenitor populations - the lymphoid-primed multi-potential progenitor (LMPP), granulocyte-macrophage progenitor (GMP) and multi-lymphoid progenitor (MLP) - were functionally and transcriptionally distinct and heterogeneous at the clonal level, with progenitors of many different functional potentials present. Though most progenitors had uni-lineage myeloid or lymphoid potential, bi- and rarer multi-lineage progenitors occurred in LMPP, GMP and MLP. This, coupled with single cell expression analyses, suggested a continuum of progenitors execute lymphoid and myeloid differentiation rather than only uni-lineage progenitors being present downstream of stem cells.

Genetic Control of Differential Acetylation in Diabetic Rats

Post-translational protein modifications such as acetylation have significant regulatory roles in metabolic processes, but their relationship to both variation in gene expression and DNA sequence is unclear. We address this question in the Goto-Kakizaki (GK) rat inbred strain, a model of polygenic type 2 diabetes. Expression of the NAD-dependent deacetylase Sirtuin-3 is down-regulated in GK rats compared to normoglycemic Brown Norway (BN) rats. We show first that a promoter SNP causes down-regulation of Sirtuin-3 expression in GK rats. We then use mass-spectrometry to identify proteome-wide differential lysine acetylation of putative Sirtuin-3 protein targets in livers of GK and BN rats. These include many proteins in pathways connected to diabetes and metabolic syndrome. We finally sequence GK and BN liver transcriptomes and find that mRNA expression of these targets does not differ significantly between GK and BN rats, in contrast to other components of the same pathways. We conclude that physiological differences between GK and BN rats are mediated by a combination of differential protein acetylation and gene transcription and that genetic variation can modulate acetylation independently of expression.

Transcriptome Profiling in Rat Inbred Strains and Experimental Cross Reveals Discrepant Genetic Architecture of Genome-Wide Gene Expression

To test the impact of genetic heterogeneity on cis- and trans-mediated mechanisms of gene expression regulation, we profiled the transcriptome of adipose tissue in 20 inbred congenic strains derived from diabetic Goto–Kakizaki (GK) rats and Brown–Norway (BN) controls, which contain well-defined blocks (1–183 Mb) of genetic polymorphisms, and in 123 genetically heterogeneous rats of an (GK × BN)F2 offspring. Within each congenic we identified 73–1351 differentially expressed genes (DEGs), only 7.7% of which mapped within the congenic blocks, and which may be regulated in cis. The remainder localized outside the blocks, and therefore must be regulated in trans. Most trans-regulated genes exhibited approximately twofold expression changes, consistent with monoallelic expression. Altered biological pathways were replicated between congenic strains sharing blocks of genetic polymorphisms, but polymorphisms at different loci also had redundant effects on transcription of common distant genes and pathways. We mapped 2735 expression quantitative trait loci (eQTL) in the F2 cross, including 26% predominantly cis-regulated genes, which validated DEGs in congenic strains. A hotspot of >300 eQTL in a 10 cM region of chromosome 1 was enriched in DEGs in a congenic strain. However, many DEGs among GK, BN and congenic strains did not replicate as eQTL in F2 hybrids, demonstrating distinct mechanisms of gene expression when alleles segregate in an outbred population or are fixed homozygous across the entire genome or in short genomic regions. Our analysis provides conceptual advances in our understanding of the complex architecture of genome expression and pathway regulation, and suggests a prominent impact of epistasis and monoallelic expression on gene transcription.