The association of Taq 1 and Fok 1 restriction fragment length polymorphisms of the vitamin D receptor with occurrence and outcome of malignant melanoma (MM), as predicted by tumour (Breslow) thickness, has been reported previously. We now report a novel adenine -guanine substitution À1012 bp relative to the exon 1a transcription start site (A-1012G), found following screening by single-stranded conformational polymorphism of this promoter region. There was a total of 191 MM cases , which were stratified according to conventional Breslow thickness groups, cases being randomly selected from each group to form a distribution corresponding to the known distribution of Breslow thickness in our area, and this population (n ¼ 176) was compared to 80 controls. The A allele was over-represented in MM patients and, with GG as reference, odds ratio (OR) for AG was 2.5, 95% confidence interval (CI) 1.1 -5.7, (P ¼ 0.03) and AA 3.3, CI 1.4 -8.1, (P ¼ 0.007). The outcome was known in 171 of 191 patients and the A allele was related to the development of metastasis, the Kaplan -Meier estimates of the probability of metastasis at 5 years being: GG 0%; AG 9%, CI 4 -16%; AA 21%, CI 12 -36%; (P ¼ 0.008), and to thicker Breslow thickness groups (P ¼ 0.04). The effect on metastasis was independent of tumour thickness and A-1012G may have predictive potential, additional to Breslow thickness. Neither the Fok 1 nor Taq 1 variants (f and t) were significantly related to the development of metastasis, although there was a strong relationship of fftt with the thickest Breslow thickness group (P ¼ 0.005). There was an interaction between the A-1012G and Fok 1 polymorphisms (P ¼ 0.025) and the Fok 1 variant enhanced the effect of the A allele of the A-1012G polymorphism on metastasis, the probability of metastasis for AAff at 5 years follow-up being 57%, CI 24 -92%.

Psoriasis is a genetically determined disease characterized by hyperproliferation and disordered maturation of the epidermis. Th1 lymphocytes are implicated in its pathogenesis. The vitamin D receptor (VDR) is a candidate modifying gene, having immunosuppressive effects and being involved in anti-proliferative and pro-differentiation pathways in keratinocytes. There is suggestive evidence that the A allele of the A-1012G polymorphism is associated with down-regulation of the Th1 response, via GATA-3. The F and T alleles of Fok1 and Taq1 have been associated with increased VDR activity. The present study aimed to test the hypothesis that the A allele of A-1012G is protective for occurrence and severity of psoriasis and enhances therapeutic response to vitamin D analogues and that these effects would be additive to those of Fok1 and Taq1. The study group comprised 206 psoriasis patients who had received topical calcipotriol treatment and 80 controls. There was no significant linkage disequilibrium between any pair of the three polymorphic sites (P=0.3-0.8). The A, F and T alleles were positively associated with calcipotriol response: AA genotype (compared to AG/GG), odds ratio (OR)=2.18 (P=0.04); TT, OR=1.97 (P=0.03); AAFF genotype combination, OR=4.11 (P=0.03); AATT, OR=5.64 (P=0.005); and FFTT, OR=3.22 (P=0.01). Comparing patients without, to patients with, a family history of psoriasis, the A allele was under represented (P=0.01) and the AAFF genotype combination even more so (compared to residual genotypes) (OR=0.24; P=0.005). AAFF was also under-represented in patients without a family history compared to controls (OR=0.31; P=0.04). There were no associations of family history with Fok1 and Taq1. There were no associations of severity of psoriasis with any polymorphism. In conclusion, the A-1012G, Fok1 and Taq1 VDR polymorphisms were associated with response to calcipotriol. A-1012G and Fok1 were associated with susceptibility to non-familial psoriasis.

BackgroundImmunolabeling of metaphase chromosome spreads can map components of the human epigenome at the single cell level. Previously, there has been no systematic attempt to explore the potential of this approach for epigenomic mapping and thereby to complement approaches based on chromatin immunoprecipitation (ChIP) and sequencing technologies.ResultsBy immunostaining and immunofluorescence microscopy, we have defined the distribution of selected histone modifications across metaphase chromosomes from normal human lymphoblastoid cells and constructed immunostained karyotypes. Histone modifications H3K9ac, H3K27ac and H3K4me3 are all located in the same set of sharply defined immunofluorescent bands, corresponding to 10- to 50-Mb genomic segments. Primary fibroblasts gave broadly the same banding pattern. Bands co-localize with regions relatively rich in genes and CpG islands. Staining intensity usually correlates with gene/CpG island content, but occasional exceptions suggest that other factors, such as transcription or SINE density, also contribute. H3K27me3, a mark associated with gene silencing, defines a set of bands that only occasionally overlap with gene-rich regions. Comparison of metaphase bands with histone modification levels across the interphase genome (ENCODE, ChIP-seq) shows a close correspondence for H3K4me3 and H3K27ac, but major differences for H3K27me3.ConclusionsAt metaphase the human genome is packaged as chromatin in which combinations of histone modifications distinguish distinct regions along the euchromatic chromosome arms. These regions reflect the high-level interphase distributions of some histone modifications, and may be involved in heritability of epigenetic states, but we also find evidence for extensive remodeling of the epigenome at mitosis.

No abstract

Histone deacetylase inhibitors (HDACi) are in clinical trials against a variety of cancers. Despite early successes, results against the more common solid tumors have been mixed. How is it that so many cancers, and most normal cells, tolerate the disruption caused by HDACi‐induced protein hyperacetylation? And why are a few cancers so sensitive? Here we discuss recent results showing that human cells mount a coordinated transcriptional response to HDACi that mitigates their toxic effects. We present a hypothetical signaling system that could trigger and mediate this response. To account for the existence of such a response, we note that HDACi of various chemical types are made by a variety of organisms to kill or suppress competitors. We suggest that the resistance response in human cells is a necessary evolutionary consequence of exposure to environmental HDACi. We speculate that cancers sensitive to HDACi are those in which the resistance response has been compromised by mutation. Identifying such mutations will allow targeting of HDACi therapy to potentially susceptible cancers.Also see the video abstract here.

BackgroundThe genome-wide hyperacetylation of chromatin caused by histone deacetylase inhibitors (HDACi) is surprisingly well tolerated by most eukaryotic cells. The homeostatic mechanisms that underlie this tolerance are unknown. Here we identify the transcriptional and epigenomic changes that constitute the earliest response of human lymphoblastoid cells to two HDACi, valproic acid and suberoylanilide hydroxamic acid (Vorinostat), both in widespread clinical use.ResultsDynamic changes in transcript levels over the first 2 h of exposure to HDACi were assayed on High Density microarrays. There was a consistent response to the two different inhibitors at several concentrations. Strikingly, components of all known lysine acetyltransferase (KAT) complexes were down-regulated, as were genes required for growth and maintenance of the lymphoid phenotype. Up-regulated gene clusters were enriched in regulators of transcription, development and phenotypic change. In untreated cells, HDACi-responsive genes, whether up- or down-regulated, were packaged in highly acetylated chromatin. This was essentially unaffected by HDACi. In contrast, HDACi induced a strong increase in H3K27me3 at transcription start sites, irrespective of their transcriptional response. Inhibition of the H3K27 methylating enzymes, EZH1/2, altered the transcriptional response to HDACi, confirming the functional significance of H3K27 methylation for specific genes.ConclusionsWe propose that the observed transcriptional changes constitute an inbuilt adaptive response to HDACi that promotes cell survival by minimising protein hyperacetylation, slowing growth and re-balancing patterns of gene expression. The transcriptional response to HDACi is mediated by a precisely timed increase in H3K27me3 at transcription start sites. In contrast, histone acetylation, at least at the three lysine residues tested, seems to play no direct role. Instead, it may provide a stable chromatin environment that allows transcriptional change to be induced by other factors, possibly acetylated non-histone proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-015-0021-9) contains supplementary material, which is available to authorized users.

Histone deacetylase inhibitors (HDACi) are increasingly used as therapeutic agents, but the mechanisms by which they alter cell behaviour remain unclear. Here we use microarray expression analysis to show that only a small proportion of genes (∼9%) have altered transcript levels after treating HL60 cells with different HDACi (valproic acid, Trichostatin A, suberoylanilide hydroxamic acid). Different gene populations respond to each inhibitor, with as many genes down- as up-regulated. Surprisingly, HDACi rarely induced increased histone acetylation at gene promoters, with most genes examined showing minimal change, irrespective of whether genes were up- or down-regulated. Many genes seem to be sheltered from the global histone hyperacetyation induced by HDACi.

The A allele of the A-1012G (rs4516035) vitamin D receptor (VDR) promoter polymorphism is associated with increased susceptibility and worsened outcome in malignant melanoma (MM). The A allele contains a GATA-3 binding site. There is a second polymorphism in the same promoter region, G-1520C (rs7139166), and there is potential for another GATA binding site in the G allele. Here, we tested the hypothesis that the G -1520 A -1012 haplotype might be a greater risk factor for MM than A-1012 alone. The A allele of A-1012G was preferentially linked to G of G-1520C and was more frequent in MM patients (p = 0.011) but G of G-1520C was not (p = 0.756). The CA haplotype was a very significant risk factor for MM (p = 0.0001) while the CG haplotype was protective (p = 0.014, combined model p = 0.00002). There was no effect of GA haplotype (p = 0.931), suggesting that that the difference in frequencies of the A allele between patients and controls was accounted for by the differences in frequencies of the CA haplotype. The A allele of A-1012G was more frequent in patients with metastasis (p = 0.054) than MM patients without metastasis, as was the G allele of G-1520C (p = 0.028). The GA haplotype was more frequent in patients with metastasis (p = 0.015), while frequencies of CA were similar. We suggest that the different roles of the A allele of A-1012G in susceptibility and metastasis risk may be a function of the availability of transcription factors in the differing cellular backgrounds related to susceptibility and progression of MM.