Disrupted Hypothalamic Transcriptomics and Proteomics in a Mouse Model of Type 2 Diabetes Exposed to Recurrent Hypoglycaemia

Aims/hypothesis: Repeated exposures to insulin-induced hypoglycaemia in people with diabetes progressively impairs the counterregulatory response (CRR) that restores normoglycaemia. This defect is characterised by reduced secretion of glucagon and other counterregulatory hormones. Evidence indicates that glucose-responsive neurons located in the hypothalamus orchestrate the CRR. Here, we aimed to identify the changes in hypothalamic gene and protein expression that underlie impaired CRR in a mouse model of defective CRR.Methods: High-fat-diet fed and low-dose streptozocin-treated C57BL/6N mice were exposed to one (acute hypoglycaemia [AH]) or multiple (recurrent hypoglycaemia [RH]) insulin-induced hypoglycaemic episodes and plasma glucagon levels were measured. Single-nuclei RNA-seq (snRNA-seq) data were obtained from the hypothalamus and cortex of mice exposed to AH and RH. Proteomic data were obtained from hypothalamic synaptosomal fractions.Results: The final insulin injection resulted in similar plasma glucose levels in the RH group and AH groups, but glucagon secretion was significantly lower in the RH group (AH: 94.5±9.2 ng/l [n=33]; RH: 59.0±4.8 ng/l [n=37]; p&lt;0.001). Analysis of snRNA-seq data revealed similar proportions of hypothalamic cell subpopulations in the AH- and RH-exposed mice. Changes in transcriptional profiles were found in all cell types analysed. In neurons from RH-exposed mice, we observed a significant decrease in expression of Avp, Pmch and Pcsk1n, and the most overexpressed gene was Kcnq1ot1, as compared with AH-exposed mice. Gene ontology analysis of differentially expressed genes (DEGs) indicated a coordinated decrease in many oxidative phosphorylation genes and reduced expression of vacuolar H +- and Na +/K +-ATPases; these observations were in large part confirmed in the proteomic analysis of synaptosomal fractions. Compared with AH-exposed mice, oligodendrocytes from RH-exposed mice had major changes in gene expression that suggested reduced myelin formation. In astrocytes from RH-exposed mice, DEGs indicated reduced capacity for neurotransmitters scavenging in tripartite synapses as compared with astrocytes from AH-exposed mice. In addition, in neurons and astrocytes, multiple changes in gene expression suggested increased amyloid beta (Aβ) production and stability. The snRNA-seq analysis of the cortex showed that the adaptation to RH involved different biological processes from those seen in the hypothalamus.Conclusions/interpretation: The present study provides a model of defective counterregulation in a mouse model of type 2 diabetes. It shows that repeated hypoglycaemic episodes induce multiple defects affecting all hypothalamic cell types and their interactions, indicative of impaired neuronal network signalling and dysegulated hypoglycaemia sensing, and displaying features of neurodegenerative diseases. It also shows that repeated hypoglycaemia leads to specific molecular adaptation in the hypothalamus when compared with the cortexData availability: The transcriptomic dataset is available via the GEO (http://www.ncbi.nlm.nih.gov/geo/), using the accession no. GSE226277. The proteomic dataset is available via the ProteomeXchange data repository (http://www.proteomexchange.org), using the accession no. PXD040183. Graphical Abstract: [Figure not available: see fulltext.].</p

Correction to:Disrupted hypothalamic transcriptomics and proteomics in a mouse model of type 2 diabetes exposed to recurrent hypoglycaemia

The affiliations for Mark Evans, Alison McNeilly and Rory J. McCrimmon have been corrected. The original article has been corrected.</p

Disrupted Hypothalamic Transcriptomics and Proteomics in a Mouse Model of Type 2 Diabetes Exposed to Recurrent Hypoglycaemia

Aims/hypothesis: Repeated exposures to insulin-induced hypoglycaemia in people with diabetes progressively impairs the counterregulatory response (CRR) that restores normoglycaemia. This defect is characterised by reduced secretion of glucagon and other counterregulatory hormones. Evidence indicates that glucose-responsive neurons located in the hypothalamus orchestrate the CRR. Here, we aimed to identify the changes in hypothalamic gene and protein expression that underlie impaired CRR in a mouse model of defective CRR.Methods: High-fat-diet fed and low-dose streptozocin-treated C57BL/6N mice were exposed to one (acute hypoglycaemia [AH]) or multiple (recurrent hypoglycaemia [RH]) insulin-induced hypoglycaemic episodes and plasma glucagon levels were measured. Single-nuclei RNA-seq (snRNA-seq) data were obtained from the hypothalamus and cortex of mice exposed to AH and RH. Proteomic data were obtained from hypothalamic synaptosomal fractions.Results: The final insulin injection resulted in similar plasma glucose levels in the RH group and AH groups, but glucagon secretion was significantly lower in the RH group (AH: 94.5±9.2 ng/l [n=33]; RH: 59.0±4.8 ng/l [n=37]; p&lt;0.001). Analysis of snRNA-seq data revealed similar proportions of hypothalamic cell subpopulations in the AH- and RH-exposed mice. Changes in transcriptional profiles were found in all cell types analysed. In neurons from RH-exposed mice, we observed a significant decrease in expression of Avp, Pmch and Pcsk1n, and the most overexpressed gene was Kcnq1ot1, as compared with AH-exposed mice. Gene ontology analysis of differentially expressed genes (DEGs) indicated a coordinated decrease in many oxidative phosphorylation genes and reduced expression of vacuolar H +- and Na +/K +-ATPases; these observations were in large part confirmed in the proteomic analysis of synaptosomal fractions. Compared with AH-exposed mice, oligodendrocytes from RH-exposed mice had major changes in gene expression that suggested reduced myelin formation. In astrocytes from RH-exposed mice, DEGs indicated reduced capacity for neurotransmitters scavenging in tripartite synapses as compared with astrocytes from AH-exposed mice. In addition, in neurons and astrocytes, multiple changes in gene expression suggested increased amyloid beta (Aβ) production and stability. The snRNA-seq analysis of the cortex showed that the adaptation to RH involved different biological processes from those seen in the hypothalamus.Conclusions/interpretation: The present study provides a model of defective counterregulation in a mouse model of type 2 diabetes. It shows that repeated hypoglycaemic episodes induce multiple defects affecting all hypothalamic cell types and their interactions, indicative of impaired neuronal network signalling and dysegulated hypoglycaemia sensing, and displaying features of neurodegenerative diseases. It also shows that repeated hypoglycaemia leads to specific molecular adaptation in the hypothalamus when compared with the cortexData availability: The transcriptomic dataset is available via the GEO (http://www.ncbi.nlm.nih.gov/geo/), using the accession no. GSE226277. The proteomic dataset is available via the ProteomeXchange data repository (http://www.proteomexchange.org), using the accession no. PXD040183. Graphical Abstract: [Figure not available: see fulltext.].</p

Correction to:Disrupted hypothalamic transcriptomics and proteomics in a mouse model of type 2 diabetes exposed to recurrent hypoglycaemia

The affiliations for Mark Evans, Alison McNeilly and Rory J. McCrimmon have been corrected. The original article has been corrected.</p

Three-dimensional chromatin interactions remain stable upon CAG/CTG repeat expansion

Expanded CAG/CTG repeats underlie 13 neurological disorders, including myotonic dystrophy type 1 (DM1) and Huntington's disease (HD). Upon expansion, disease loci acquire heterochromatic characteristics, which may provoke changes to chromatin conformation and thereby affect both gene expression and repeat instability. Here, we tested this hypothesis by performing 4C sequencing at the DMPK and HTT loci from DM1 and HD-derived cells. We find that allele sizes ranging from 15 to 1700 repeats displayed similar chromatin interaction profiles. This was true for both loci and for alleles with different DNA methylation levels and CTCF binding. Moreover, the ectopic insertion of an expanded CAG repeat tract did not change the conformation of the surrounding chromatin. We conclude that CAG/CTG repeat expansions are not enough to alter chromatin conformation in cis. Therefore, it is unlikely that changes in chromatin interactions drive repeat instability or changes in gene expression in these disorders

Conformational adaptation of Asian macaque TRIMCyp directs lineage specific antiviral activity

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Transposable elements and their KRAB/KAP1 controllers broadly regulate transcription in adult human cells

KAP1 is a universal corepressor for the large family of KRAB-ZFP proteins that coordinates epigenetic silencing of endogenous retroelements (EREs) during early embryonic development. This process is essential not just to prevent replication of EREs, but also to control their regulatory influence on host genes. Although in differentiated cells the control of EREs is believed to be mainly DNA methylation-dependent, emerging evidence suggests that KAP1-dependent, histone-based repression might remain a key regulator of ERE expression and its influence on cellular gene networks. Using a combination of transcriptional and histone modification profiling, we have investigated the impact of KAP1 epigenetic regulation on the transcriptional dynamics of human CD4+ T cells. We found that KAP1 binding on subsets of EREs is maintained in differentiated CD4+ T cells, where its locus-specific recruitment is dynamically modulated along with the T cell activation status. KAP1 depletion by RNA interference leads to a global decrease in repressive histone modifications that is accompanied by broad transcriptional alterations. Indeed, not only a large fraction of EREs become derepressed, but their deregulation correlates with illegittimate activation of nearby genes. Correspondingly, loss of KAP1 unleashes the intrinsic regulatory activities that some EREs are endowed with, suggesting a functional link between control of mobile genetic elements and protection of lineage-specific transcriptional networks. These findings establish the KRAB/KAP1 system as an important safeguard of CD4+ T cells transcriptional identity and suggest a critical role in maintaining epigenetic silencing of ERE-based regulatory sequences in adult tissues. Furthermore, our study shed light onto a previously unappreciated contribution of retrotransposon-derived sequences to the complexity of the human T cell transcriptome. A better knowledge of the physiological transcriptional activity of these mobile genetic elements, as well as their control mechanisms throughout development, would greatly help the debate on their pathologic implications

Interplay of TRIM28 and DNA methylation in controlling human endogenous retroelements

Reverse transcription-derived sequences account for at least half of the human genome. Although these retroelements are formidable motors of evolution, they can occasionally cause disease, and accordingly are inactivated during early embryogenesis through epigenetic mechanisms. In the mouse, at least for endogenous retroviruses, important mediators of this process are the tetrapod-specific KRAB-containing zinc finger proteins (KRAB-ZFPs) and their cofactor TRIM28. The present study demonstrates that KRAB/TRIM28-mediated regulation is responsible for controlling a very broad range of human-specific endogenous retroelements (EREs) in human embryonic stem (ES) cells and that it exerts, as a consequence, a marked effect on the transcriptional dynamics of these cells. It further reveals reciprocal dependence between TRIM28 recruitment at specific families of EREs and DNA methylation. It finally points to the importance of persistent TRIM28-mediated control of ERE transcriptional impact beyond their presumed inactivation by DNA methylation

HIV-1 Vpr and p21 restrict LINE-1 mobility

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