Dosage compensation and the global re-balancing of aneuploid genomes

Diploid genomes are exquisitely balanced systems of gene expression. The dosage-compensation systems that evolved along with monosomic sex chromosomes exemplify the intricacies of compensating for differences in gene copy number by transcriptional regulation

The MOF-containing NSL complex associates globally with housekeeping genes, but activates only a defined subset

The MOF (males absent on the first)-containing NSL (non-specific lethal) complex binds to a subset of active promoters in Drosophila melanogaster and is thought to contribute to proper gene expression. The determinants that target NSL to specific promoters and the circumstances in which the complex engages in regulating transcription are currently unknown. Here, we show that the NSL complex primarily targets active promoters and in particular housekeeping genes, at which it colocalizes with the chromatin remodeler NURF (nucleosome remodeling factor) and the histone methyltransferase Trithorax. However, only a subset of housekeeping genes associated with NSL are actually activated by it. Our analyses reveal that these NSL-activated promoters are depleted of certain insulator binding proteins and are enriched for the core promoter motif ‘Ohler 5’. Based on these results, it is possible to predict whether the NSL complex is likely to regulate a particular promoter. We conclude that the regulatory capacity of the NSL complex is highly context-dependent. Activation by the NSL complex requires a particular promoter architecture defined by combinations of chromatin regulators and core promoter motifs

Fibrillar Aβ (beta) triggers microglial proteome alterations and dysfunction in Alzheimer mouse models

Microglial dysfunction is a key pathological feature of Alzheimer's disease (AD), but little is known about proteome-wide changes in microglia during the course of AD and their functional consequences. Here, we performed an in-depth and time-resolved proteomic characterization of microglia in two mouse models of amyloid beta (A beta) pathology, the overexpression APPPS1 and the knock-in APP-NL-G-F (APP-KI) model. We identified a large panel of Microglial A beta Response Proteins (MARPs) that reflect heterogeneity of microglial alterations during early, middle and advanced stages of A beta deposition and occur earlier in the APPPS1 mice. Strikingly, the kinetic differences in proteomic profiles correlated with the presence of fibrillar A beta, rather than dystrophic neurites, suggesting that fibrillar A beta may trigger the AD-associated microglial phenotype and the observed functional decline. The identified microglial proteomic fingerprints of AD provide a valuable resource for functional studies of novel molecular targets and potential biomarkers for monitoring AD progression or therapeutic efficacy

Transcription through Intergenic Chromosomal Memory Elements of the Drosophila Bithorax Complex Correlates with an Epigenetic Switch

The proteins of the trithorax and Polycomb groups maintain the differential expression pattern of homeotic genes established by the early embryonic patterning system during development. These proteins generate stable and heritable chromatin structures by acting via particular chromosomal memory elements. We established a transgenic assay system showing that the Polycomb group response elements bxd and Mcp confer epigenetic inheritance throughout development. With previously published data for the Fab7 cellular memory module, we confirmed the cellular memory function of Polycomb group response elements. In Drosophila melanogaster, several of these memory elements are located in the large intergenic regulatory regions of the homeotic bithorax complex. Using a transgene assay, we showed that transcription through a memory element correlated with the relief of silencing imposed by the Polycomb group proteins and established an epigenetically heritable active chromatin mode. A memory element remodeled by the process of transcription was able to maintain active expression of a reporter gene throughout development. Thus, transcription appears to reset and change epigenetic marks at chromosomal memory elements regulated by the Polycomb and trithorax proteins. Interestingly, in the bithorax complex of D. melanogaster, the segment-specific expression of noncoding intergenic transcripts during embryogenesis seems to fulfill this switching role for memory elements regulating the homeotic genes

Intergenic transcription through a Polycomb group response element counteracts silencing

Polycomb group response elements (PREs) mediate the mitotic inheritance of gene expression programs and thus maintain determined cell fates. By default, PREs silence associated genes via the targeting of Polycomb group (PcG) complexes. Upon an activating signal, however, PREs recruit counteracting trithorax group (trxG) proteins, which in turn maintain target genes in a transcriptionally active state. Using a transgenic reporter system, we show that the switch from the silenced to the activated state of a PRE requires noncoding transcription. Continuous transcription through the PRE induced by an actin promoter prevents the establishment of PcG-mediated silencing. The maintenance of epigenetic activation requires transcription through the PRE to proceed at least until embryogenesis is completed. At the homeotic bithorax complex of Drosophila, intergenic PRE transcripts can be detected not only during embryogenesis, but also at late larval stages, suggesting that transcription through endogenous PREs is required continuously as an anti-silencing mechanism to prevent the access of repressive PcG complexes to the chromatin. Furthermore, all other PREs outside the homeotic complex we tested were found to be transcribed in the same tissue as the mRNA of the corresponding target gene, suggesting that anti-silencing by transcription is a fundamental aspect of the cellular memory system

Transcription of histone gene cluster by differential core-promoter factors

The 100 copies of tandemly arrayed Drosophila linker (H1) and core (H2A/B and H3/H4) histone gene cluster are coordinately regulated during the cell cycle. However, the molecular mechanisms that must allow differential transcription of linker versus core histones prevalent during development remain elusive. Here, we used fluorescence imaging, biochemistry, and genetics to show that TBP (TATA-box-binding protein)-related factor 2 (TRF2) selectively regulates the TATA-less Histone H1 gene promoter, while TBP/TFIID targets core histone transcription. Importantly, TRF2-depleted polytene chromosomes display severe chromosomal structural defects. This selective usage of TRF2 and TBP provides a novel mechanism to differentially direct transcription within the histone cluster. Moreover, genome-wide chromatin immunoprecipitation (ChIP)-on-chip analyses coupled with RNA interference (RNAi)-mediated functional studies revealed that TRF2 targets several classes of TATA-less promoters of >1000 genes including those driving transcription of essential chromatin organization and protein synthesis genes. Our studies establish that TRF2 promoter recognition complexes play a significantly more central role in governing metazoan transcription than previously appreciated

Life span extension by targeting a link between metabolism and histone acetylation in <em>Drosophila</em>.

Old age is associated with a progressive decline of mitochondrial function and changes in nuclear chromatin. However, little is known about how metabolic activity and epigenetic modifications change as organisms reach their midlife. Here, we assessed how cellular metabolism and protein acetylation change during early aging in Drosophila melanogaster. Contrary to common assumptions, we find that flies increase oxygen consumption and become less sensitive to histone deacetylase inhibitors as they reach midlife. Further, midlife flies show changes in the metabolome, elevated acetyl-CoA levels, alterations in protein-notably histone-acetylation, as well as associated transcriptome changes. Based on these observations, we decreased the activity of the acetyl-CoA-synthesizing enzyme ATP citrate lyase (ATPCL) or the levels of the histone H4 K12-specific acetyltransferase Chameau. We find that these targeted interventions both alleviate the observed aging-associated changes and promote longevity. Our findings reveal a pathway that couples changes of intermediate metabolism during aging with the chromatin-mediated regulation of transcription and changes in the activity of associated enzymes that modulate organismal life span

The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1

Background and Purpose- Genome-wide association studies have identified the HDAC9 (histone deacetylase 9) gene region as a major risk locus for atherosclerotic stroke and coronary artery disease in humans. Previous results suggest a role of altered HDAC9 expression levels as the underlying disease mechanism. rs2107595, the lead single nucleotide polymorphism for stroke and coronary artery disease resides in noncoding DNA and colocalizes with histone modification marks suggestive of enhancer elements. Methods- To determine the mechanisms by which genetic variation at rs2107595 regulates HDAC9 expression and thus vascular risk we employed targeted resequencing, proteome-wide search for allele-specific nuclear binding partners, chromatin immunoprecipitation, genome-editing, reporter assays, circularized chromosome conformation capture, and gain- and loss-of-function experiments in cultured human cell lines and primary immune cells. Results- Targeted resequencing of the HDAC9 locus in patients with atherosclerotic stroke and controls supported candidacy of rs2107595 as the causative single nucleotide polymorphism. A proteomic search for nuclear binding partners revealed preferential binding of the E2F3/TFDP1/Rb1 complex (E2F transcription factor 3/transcription factor Dp-1/Retinoblastoma 1) to the rs2107595 common allele, consistent with the disruption of an E2F3 consensus site by the risk allele. Gain- and loss-of-function studies showed a regulatory effect of E2F/Rb proteins on HDAC9 expression. Compared with the common allele, the rs2107595 risk allele exhibited higher transcriptional capacity in luciferase assays and was associated with higher HDAC9 mRNA levels in primary macrophages and genome-edited Jurkat cells. Circularized chromosome conformation capture revealed a genomic interaction of the rs2107595 region with the HDAC9 promoter, which was stronger for the common allele as was the in vivo interaction with E2F3 and Rb1 determined by chromatin immunoprecipitation. Gain-of-function experiments in isogenic Jurkat cells demonstrated a key role of E2F3 in mediating rs2107595-dependent transcriptional regulation of HDAC9. Conclusions- Collectively, our findings imply allele-specific transcriptional regulation of HDAC9 via E2F3 and Rb1 as a major mechanism mediating vascular risk at rs2107595