Epigenome-wide association data implicates DNA methylation-mediated genetic risk in psoriasis

Abstract Background Psoriasis is a chronic inflammatory skin disease characterized by epidermal hyperproliferation and altered keratinocyte differentiation and inflammation and is caused by the interplay of genetic and environmental factors. Previous studies have revealed that DNA methylation (DNAm) and genetic makers are closely associated with psoriasis, and strong evidences have shown that DNAm can be controlled by genetic factors, which attracted us to evaluate the relationship among DNAm, genetic makers, and disease status. Methods We utilized the genome-wide methylation data of psoriatic skin (PP, N = 114) and unaffected control skin (NN, N = 62) tissue samples in our previous study, and we performed whole-genome genotyping with peripheral blood of the same samples to evaluate the underlying genetic effect on skin DNA methylation. Causal inference test (CIT) was used to assess whether DNAm regulate genetic variation and gain a better understanding of the epigenetic basis of psoriasis susceptibility. Results We identified 129 SNP-CpG pairs achieving the significant association threshold, which constituted 28 unique methylation quantitative trait loci (MethQTL) and 34 unique CpGs. There are 18 SNPs were associated with psoriasis at a Bonferoni-corrected P < 0.05, and these 18 SNPs formed 93 SNP-CpG pairs with 17 unique CpG sites. We found that 11 of 93 SNP-CpG pairs, composed of 5 unique SNPs and 3 CpG sites, presented a methylation-mediated relationship between SNPs and psoriasis. The 3 CpG sites were located on the body of C1orf106, the TSS1500 promoter region of DMBX1 and the body of SIK3. Conclusions This study revealed that DNAm of some genes can be controlled by genetic factors and also mediate risk variation for psoriasis in Chinese Han population and provided novel molecular insights into the pathogenesis of psoriasis

Infrared Spectroscopic Study on the Modified Mechanism of Aluminum-Impregnated Bone Charcoal

Fluoride contamination in drinking water is a prominent and widespread problem in many parts of the world. Excessive ingestion of fluoride through water can lead to the high risk of fluorosis in human body. Bone charcoal, with the principal active component of hydroxyapatite, is a frequently used adsorbent for fluoride removal. Many laboratory experiments suggest that the aluminum-impregnated bone charcoal is an effective adsorbent in defluoridation. However, the mechanisms underlying this modification process are still not well understood, which in turn greatly impedes the further studies on other different modified adsorbents. To address this issue, we used the infrared spectroscopy to examine the bone charcoal and the aluminum-impregnated bone charcoal, respectively. The comparative results show that the −OH peak of infrared spectroscopy has been intensified after modification. This significant change helped speculate the modified mechanism of the aluminum-impregnated bone charcoal. In addition, it is found that the hydroxide ion dissociates from hydroxyapatite in the modification process. Such finding implies that the tetrahydroxoaluminate can be combined with the hydroxyapatite and the aluminum ion can be impregnated onto the bone char surface

Phosphorylation modulates AfsR transcriptional activation.

(A) LC-MS/MS analysis showing that S22, T337, S391, T506, and S953 are phosphorylated in AfsR by AfsKΔC. The lowercase letter in the peptide sequence indicates phosphorylated residue. “b” and “y” denote peptide fragment ions retaining charges at the N and C terminus, respectively. The subscript numbers indicate their positions in the identified peptide. (B) Sequence alignment of Streptomyces AfsR family members highlighting the consensuses sequences neighboring phosphorylation sites S22, T337, S391, T506, and S953. Orange boxes represent phosphorylation sites. (C) Transcription assays with increasing concentrations of phosphorylated AfsR by AfsKΔC (phos-AfsR) and untreated AfsR. (D) Transcription assays of 500 nM AfsR mutants mimicking dephosphorylation (T337A) and phosphorylation (S22E, T337E, T506E, S953E, and S391E/T337E/T506E/S953E (4E)). Data are presented as mean ± SEM from 3 independent assays. n.s. means no significance; *P P T337A and AfsR4E. (F) Fluorescence polarization assay of AfsR4E and dephosphorylated AfsR (dephos-AfsR) with afs box. The concentration of afs box was 10 nM. Error bars represent mean ± SEM of n = 3 experiments. The data underlying A, C, D, E, and F are provided in S1 Data. (TIF)</p

SARP interacts with σ^HrdB R4, β FTH, β′ ZBD.

(A) The SARP protomers interact with σHrdB, β, and β′. (B) The upstream SARP protomer contacts σHrdB R4 by its ODB domain. Salt bridges are shown as red dashed lines. (C) The H499 of σHrdBR4 is enfolded in an amphiphilic pocket of the BTA domain. The K496 of σHrdBR4 makes a salt bridge with the E246 of the BTA domain. (D) The downstream SARP protomer make extensive interactions with the β FTH, the preceding loop (TPL) and the following loop (TFL) of the β flap. SARP is colored orange and β flap is colored blue. Hydrogen bonds, salt-bridges, and van der Waals interactions are shown as yellow, red, and gray dashed lines, respectively. (E) Interactions between the β′ ZBD and the ODB of downstream SARP. The positively charged R67 and R69 of β′ ZBD contact the negatively charged E76 and E77 of the HTH loop of the ODB domain. (F) Mutating interfacial residues of SARP impaired transcription activation. The data underlying this figure can be found in S1 Data; error bars, SEM; n = 3; *P P Streptomyces strains, highlighting the residues interacting with β (blue), σ R4 (cyan), and αCTD (purple). The black boxes highlight the positions conserved. BTA, bacterial transcriptional activation; FTH, flap tip helix; ODB, OmpR-type DNA-binding; SARP, Streptomyces antibiotic regulatory protein; ZBD, zinc-binding domain.</p

ATPγS activates phosphorylated AfsR and AfsR_4E.

Transcription assays involving 500 nM phosphorylated AfsR (phos-AfsR) or AfsR4E, with and without preincubation with 1 mM ATP or ATPγS. CK represents the control group without the addition of AfsR. The data underlying this figure can be found in S1 Data; error bars, SEM; n = 3. (TIF)</p

Scheme model summarizing the ATP-dependent and phosphorylation-dependent regulation of AfsR.

Unmodified full-length AfsR is autoinhibited by its C-terminal domains. AfsR is activated by ATP binding and significantly stimulates the production of transcripts. The phosphorylation of AfsR results in increased ATPase activity, potentially counteracting the effects of ATP binding. Phosphorylated AfsR forms more oligomers and shows slightly higher transcriptional activation compared to the unmodified form. (TIF)</p

Comparison of SARP with EmbR (PDB ID: 2FEZ) comprising an additional C-terminal FHA domain.

Comparison of SARP with EmbR (PDB ID: 2FEZ) comprising an additional C-terminal FHA domain.</p

List of primer sequences used in this study.

Streptomyces antibiotic regulatory proteins (SARPs) are widely distributed activators of antibiotic biosynthesis. Streptomyces coelicolor AfsR is an SARP regulator with an additional nucleotide-binding oligomerization domain (NOD) and a tetratricopeptide repeat (TPR) domain. Here, we present cryo-electron microscopy (cryo-EM) structures and in vitro assays to demonstrate how the SARP domain activates transcription and how it is modulated by NOD and TPR domains. The structures of transcription initiation complexes (TICs) show that the SARP domain forms a side-by-side dimer to simultaneously engage the afs box overlapping the −35 element and the σHrdB region 4 (R4), resembling a sigma adaptation mechanism. The SARP extensively interacts with the subunits of the RNA polymerase (RNAP) core enzyme including the β-flap tip helix (FTH), the β′ zinc-binding domain (ZBD), and the highly flexible C-terminal domain of the α subunit (αCTD). Transcription assays of full-length AfsR and truncated proteins reveal the inhibitory effect of NOD and TPR on SARP transcription activation, which can be eliminated by ATP binding. In vitro phosphorylation hardly affects transcription activation of AfsR, but counteracts the disinhibition of ATP binding. Overall, our results present a detailed molecular view of how AfsR serves to activate transcription.</div

In vitro assays of the <i>Streptomyces coelicolor</i> AfsR SARP.

(A) Core promoter sequences used for in vitro assays. The mutations introduced into the afs box are highlighted. The mutated target sites contained mutations in upstream repeat (M1), the downstream repeat (M2), or both repeats (M1M2). The actII-4 promoter was used as a control. (B) Fluorescence polarization assays of the SARP with mutant afs box (M1, M2, and M1M2). Error bars represent mean ± SEM of n = 3 experiments. (C) In vitro MangoIII-based transcription assays with or without 500 nM SARP in the absence of RbpA and CarD. Error bars represent mean ± SEM of n = 3 experiments. The data underlying B and C can be found in S1 Data. (TIF)</p