Mendelian randomization analyses explore the relationship between cathepsins and lung cancer.

Lung cancer, a major contributor to cancer-related fatalities worldwide, involves a complex pathogenesis. Cathepsins, lysosomal cysteine proteases, play roles in various physiological and pathological processes, including tumorigenesis. Observational studies have suggested an association between cathepsins and lung cancer. However, the causal link between the cathepsin family and lung cancer remains undetermined. This study employed Mendelian randomization analyses to investigate this causal association. The univariable Mendelian randomization analysis results indicate that elevated cathepsin H levels increase the overall risk of lung cancer, adenocarcinoma, and lung cancer among smokers. Conversely, reverse Mendelian randomization analyses suggest that squamous carcinoma may lead to increased cathepsin B levels. A multivariable analysis using nine cathepsins as covariates reveals that elevated cathepsin H levels lead to an increased overall risk of lung cancer, adenocarcinoma, and lung cancer in smokers. In conclusion, cathepsin H may serve as a marker for lung cancer, potentially inspiring directions in lung cancer diagnosis and treatment.

Candidate pathway analysis of surfactant proteins identifies CTSH and SFTA2 that influences lung cancer risk.

Pulmonary surfactant is a lipoprotein synthesized and secreted by alveolar type II cells in lung. We evaluated the associations between 200,139 single nucleotide polymorphisms (SNPs) of 40 surfactant-related genes and lung cancer risk using genotyped data from two independent lung cancer genome-wide association studies. Discovery data included 18,082 cases and 13,780 controls of European ancestry. Replication data included 1,914 cases and 3,065 controls of European descent. Using multivariate logistic regression, we found novel SNPs in surfactant-related genes CTSH [rs34577742 C > T, odds ratio (OR) = 0.90, 95% confidence interval (CI) = 0.89-0.93, P = 7.64 × 10-9] and SFTA2 (rs3095153 G > A, OR = 1.16, 95% CI = 1.10-1.21, P = 1.27 × 10-9) associated with overall lung cancer in the discovery data and validated in an independent replication data-CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.80-0.96, P = 5.76 × 10-3) and SFTA2 (rs3095153 G > A, OR = 1.14, 95% CI = 1.01-1.28, P = 3.25 × 10-2). Among ever smokers, we found SNPs in CTSH (rs34577742 C > T, OR = 0.89, 95% CI = 0.85-0.92, P = 1.94 × 10-7) and SFTA2 (rs3095152 G > A, OR = 1.20, 95% CI = 1.14-1.27, P = 4.25 × 10-11) associated with overall lung cancer in the discovery data and validated in the replication data-CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.79-0.97, P = 1.64 × 10-2) and SFTA2 (rs3095152 G > A, OR = 1.15, 95% CI = 1.01-1.30, P = 3.81 × 10-2). Subsequent transcriptome-wide association study using expression weights from a lung expression quantitative trait loci study revealed genes most strongly associated with lung cancer are CTSH (PTWAS = 2.44 × 10-4) and SFTA2 (PTWAS = 2.32 × 10-6).

Cathepsin H: Molecular characteristics and clues to function and mechanism.

Cathepsin H (CatH) is a lysosomal cysteine protease with a unique aminopeptidase activity that is extensively expressed in the lung, pancreas, thymus, kidney, liver, skin, and brain. Owing to its specific enzymatic activity, CatH has critical effects on the regulation of biological behaviours of cancer cells and pathological processes in brain diseases. Moreover, a neutral pH level is optimal for CatH activity, so it is expected to be active in the extra-lysosomal and extracellular space. In the present review, we describe the expression, maturation, and enzymatic properties of CatH, and summarize the available experimental evidence that mechanistically links CatH to various physiological and pathological processes. Finally, we discuss the challenges and potentials of CatH inhibitors in CatH-induced disease therapy.

Cancer-Associated Fibroblasts Are Key Determinants of Cancer Cell Invasion in the Earliest Stage of Colorectal Cancer.

BACKGROUND & AIMS: Improving clinical management of early stage colorectal cancers (T1CRCs) requires a better understanding of their underlying biology. Accumulating evidence shows that cancer-associated fibroblasts (CAFs) are important determinants of tumor progression in advanced colorectal cancer (CRC), but their role in the initial stages of CRC tumorigenesis is unknown. Therefore, we investigated the contribution of T1CAFs to early CRC progression. METHODS: Primary T1CAFs and patient-matched normal fibroblasts (NFs) were isolated from endoscopic biopsy specimens of histologically confirmed T1CRCs and normal mucosa, respectively. The impact of T1CAFs and NFs on tumor behavior was studied using 3-dimensional co-culture systems with primary T1CRC organoids and extracellular matrix (ECM) remodeling assays. Whole-transcriptome sequencing and gene silencing were used to pinpoint mediators of T1CAF functions. RESULTS: In 3-dimensional multicellular cultures, matrix invasion of T1CRC organoids was induced by T1CAFs, but not by matched NFs. Enhanced T1CRC invasion was accompanied by T1CAF-induced ECM remodeling and up-regulation of CD44 in epithelial cells. RNA sequencing of 10 NF-T1CAF pairs revealed 404 differentially expressed genes, with significant enrichment for ECM-related pathways in T1CAFs. Cathepsin H, a cysteine-type protease that was specifically up-regulated in T1CAFs but not in fibroblasts from premalignant lesions or advanced CRCs, was identified as a key factor driving matrix remodeling by T1CAFs. Finally, we showed high abundance of cathepsin H-expressing T1CAFs at the invasive front of primary T1CRC sections. CONCLUSIONS: Already in the earliest stage of CRC, cancer cell invasion is promoted by CAFs via direct interactions with epithelial cancer cells and stage-specific, cathepsin H-dependent ECM remodeling. RNA sequencing data of the 10 NF-T1CAF pairs can be found under GEO accession number GSE200660.

Cathepsin H Knockdown Reverses Radioresistance of Hepatocellular Carcinoma via Metabolic Switch Followed by Apoptosis.

Despite the wide application of radiotherapy in HCC, radiotherapy efficacy is sometimes limited due to radioresistance. Although radioresistance is reported with high glycolysis, the underlying mechanism between radioresistance and cancer metabolism, as well as the role of cathepsin H (CTSH) within it, remain unclear. In this study, tumor-bearing models and HCC cell lines were used to observe the effect of CTSH on radioresistance. Proteome mass spectrometry, followed by enrichment analysis, were used to investigate the cascades and targets regulated by CTSH. Technologies such as immunofluorescence co-localization flow cytometry and Western blot were used for further detection and verification. Through these methods, we originally found CTSH knockdown (KD) perturbed aerobic glycolysis and enhanced aerobic respiration, and thus promoted apoptosis through up-regulation and the release of proapoptotic factors such as AIFM1, HTRA2, and DIABLO, consequently reducing radioresistance. We also found that CTSH, together with its regulatory targets (such as PFKL, HK2, LDH, and AIFM1), was correlated with tumorigenesis and poor prognosis. In summary, our study found that the cancer metabolic switch and apoptosis were regulated by CTSH signaling, leading to the occurrence of radioresistance in HCC cells and suggesting the potential value of HCC diagnosis and therapy.

Cathepsin H and cathepsin B of Cynoglossus semilaevis are involved in anti-bacterial immunity against Edwardsiella tarda.

Cathepsin H and Cathepsin B are two lysosomal cysteine proteases participating in various physiological processes including immune responses. In fish, the functional roles of Cathepsin H and Cathepsin B during bacterial infection are less understood. In a previous work, we characterized a Cathepsin B homologue (CsCatB) of half-smooth tongue sole (Cynoglossus semilaevis), an economically valuable fish species in China. In this report, we identified a Cathepsin H homologue (CsCatH) from C. semilaevis. In healthy tongue sole, the transcriptional expression of CsCatH was detected in nine different tissues. Laser scanning confocal microscopic analysis showed that ectopically expressed CsCatH and CsCatB were co-localized with the lysosome. Upon infection by Edwardsiella tarda, a significant fish pathogen which caused a severe fish disease termed edwardsiellosis, the expressions of CsCatH and CsCatB were remarkedly upregulated. The knockdown of CsCatH and CsCatB significantly increased the replication of E. tarda and mitigated E. tarda-induced apoptosis in tongue sole tissues. These findings revealed the importance of CsCatH and CsCatB in anti-bacterial immunity of tongue sole.

Functional genomics identify causal variant underlying the protective CTSH locus for Alzheimer's disease.

Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disease, which has a high heritability of up to 79%. Exploring the genetic basis is essential for understanding the pathogenic mechanisms underlying AD development. Recent genome-wide association studies (GWASs) reported an AD-associated signal in the Cathepsin H (CTSH) gene in European populations. However, the exact functional/causal variant(s), and the genetic regulating mechanism of CTSH in AD remain to be determined. In this study, we carried out a comprehensive study to characterize the role of CTSH variants in the pathogenesis of AD. We identified rs2289702 in CTSH as the most significant functional variant that is associated with a protective effect against AD. The genetic association between rs2289702 and AD was validated in independent cohorts of the Han Chinese population. The CTSH mRNA expression level was significantly increased in AD patients and AD animal models, and the protective allele T of rs2289702 was associated with a decreased expression level of CTSH through the disruption of the binding affinity of transcription factors. Human microglia cells with CTSH knockout showed a significantly increased phagocytosis of Aß peptides. Our study identified CTSH as being involved in AD genetic susceptibility and uncovered the genetic regulating mechanism of CTSH in pathogenesis of AD.

A five-gene methylation signature predicts overall survival of patients with clear cell renal cell carcinoma.

BACKGROUND: In this study, we aimed to screen methylation signatures associated with the prognosis of patients with clear cell renal cell carcinoma (ccRCC). METHODS: Gene expression and methylation profiles of ccRCC patients were downloaded from publicly available databases, and differentially expressed genes (DEGs)-differentially methylated genes (DMGs) were obtained. Subsequently, gene set enrichment and transcription factor (TF) regulatory network analyses were performed. In addition, a prognostic model was constructed and the relationship between disease progression and immunity was analyzed. RESULTS: A total of 23 common DEGs-DMGs were analyzed, among which 14 DEGs-DMGs were obtained with a cutoff value of PCC < 0 and p < 0.05. The enrichment analysis showed that the 14 DEGs-DMGs were enriched in three GO terms and three KEGG pathways. In addition, a total of six TFs were shown to be associated with the 14 DEGs-DMGs, including RP58, SOX9, NF-κB65, ATF6, OCT, and IK2. A prognostic model using five optimized DEGs-DMGs which efficiently predicted survival was constructed and validated using the GSE105288 dataset. Additionally, four types of immune cells (NK cells, macrophages, neutrophils, and cancer-associated fibroblasts), as well as ESTIMATE, immune, and stromal scores were found to be significantly correlated with ccRCC progression (normal, primary, and metastasis) in addition to the five optimized DEGs-DMGs. CONCLUSION: A five-gene methylation signature with the predictive ability for ccRCC prognosis was investigated in this study, consisting of CCNB2, CDKN1C, CTSH, E2F2, and ERMP1. In addition, potential targets for methylation-mediated immunotherapy were highlighted.

Cathepsin H deficiency decreases hypoxia-ischemia-induced hippocampal atrophy in neonatal mice through attenuated TLR3/IFN-ß signaling.

BACKGROUND: Cathepsin H (CatH) is a lysosomal cysteine protease with a unique aminopeptidase activity. Its expression level is increased in activated immune cells including dendritic cells, macrophages, and microglia. We have previously reported that CatH deficiency impairs toll-like receptor 3 (TLR3)-mediated activation of interferon regulatory factor 3 (IRF3), and the subsequent secretion of interferon (IFN)-ß from dendritic cells. Furthermore, there is increasing evidence that IFN-ß secreted from microglia/macrophages has neuroprotective effects. These observations prompted further investigation into the effects of CatH deficiency on neuropathological changes. METHODS: In this study, neuropathological changes were examined using histochemical staining (both hematoxylin-eosin (H&E) and Nissl) of the hippocampus of wild-type (WT) and CatH-deficient (CatH-/-) mice after hypoxia-ischemia (HI). The density and the localization of CatH and TLR3 were examined by immunofluorescent staining. CatH processing in microglia was assayed by pulse-chase experiments, while immunoblotting was used to examine TLR3 expression and IRF3 activation in microglia/macrophages in the presence of poly(I:C). Microglial cell death was examined by fluorescence-activated cell sorting (FACS), and primary astrocyte proliferation in the presence of IFN-ß was examined using scratch wound assay. RESULTS: WT mice displayed severe atrophy in association with neuronal death and moderate astrogliosis in the hippocampus following neonatal HI. Somewhat surprisingly, CatH-/- mice showed marked neuronal death without severe atrophy in the hippocampus following HI. Furthermore, there was notable microglia/macrophages cell death and strong astrogliosis in the hippocampus. The TLR3 and phosphorylated IRF3 expression level in the hippocampus or splenocytes (mainly splenic macrophages); from CatH-/- mice was lower than in WT mice. In vitro experiments demonstrated that recombinant IFN-ß suppressed HI-induced microglial cell death and astrocyte proliferation. CONCLUSION: These observations suggest that CatH plays a critical role in the proteolytic maturation and stabilization of TLR3, which is necessary for IFN-ß production. Therefore, impaired TLR3/IFN-ß signaling resulting from CatH deficiency may induce microglial cell death after activation and astrogliosis/glial scar formation in the hippocampus following HI injury, leading to suppression of hippocampal atrophy.

Genetic and environmental factors regulate the type 1 diabetes gene CTSH via differential DNA methylation.

Cathepsin H (CTSH) is a type 1 diabetes (T1D) risk gene; large-scale genetic and epidemiological studies found that T1D genetic risk correlates with high CTSH expression, rapid decline of beta-cell function, and early onset T1D. Counterintuitively, transcriptional downregulation of CTSH by proinflammatory cytokines has been shown to promote beta-cell apoptosis. Here, we potentially explain these observed contrasting effects, describing a new mechanism where proinflammatory cytokines and T1D genetic risk variants regulate CTSH transcription via differential DNA methylation. We show that, in human islets, CTSH downregulation by the proinflammatory cytokine cocktail interleukin 1ß + tumor necrosis factor α + interferon γ was coupled with DNA hypermethylation in an open chromatin region in CTSH intron 1. A luciferase assay in human embryonic kidney 293 cells revealed that methylation of three key cytosine-phosphate-guanine dinucleotide (CpG) residues in intron 1 was responsible for the reduction of promoter activity. We further found that cytokine-induced intron 1 hypermethylation is caused by lowered Tet1/3 activities, suggesting that attenuated active demethylation lowered CTSH transcription. Importantly, individuals who carry the T1D risk variant showed lower methylation variability at the intron 1 CpG residues, presumably making them less sensitive to cytokines, whereas individuals who carry the protective variant showed higher methylation variability, presumably making them more sensitive to cytokines and implying differential responses to environment between the two patient populations. These findings suggest that genetic and environmental influences on a T1D locus are mediated by differential variability and mean of DNA methylation.

Some thiocarbamoyl based novel anticathepsin agents.

Cathepsins have emerged as important targets in various tissues degenerative disorders due to their involvement in degradation of extracellular matrices and endogenous protein turnover. Elevated cathepsins levels vis-à-vis decreased concentration of endogenous inhibitors has been reported at different diseased sites. The design and synthesis of specific potential anti-cathepsin agents is therefore of great significance. Most of potential anti-cathepsin agents developed have peptide based structures with an active warhead. Due to oral instability and immunogenic problems related to peptidyl inhibitors drift the synthesis and evaluation of non-peptide cathepsin inhibitors in last two decades. The present work provides a detailed structure activity relationship for developing potential non-peptide anticathepsin agents based on in-vitro inhibition studies of a library of synthesized thiocarbamoyl- non-peptide inhibitors.

Study on enhanced serum protein protecting and anti-cathepsin activities of various curcumin formulations containing traditional excipients and bio-enhancers.

Cathepsins have emerged out as significant targets in variety of tissue degenerative disorders such as inflammation, alzeimers, tumerogenesis including metastasis and invasion. Elevated levels of cathepsins and reduced cellular inhibitors at the site of these diseased conditions suggest the exploration of novel inhibitors of cathepsins. In the search of effective novel inhibitors as anti-cathepsin agents different natural products are also screened. One such molecule, curcumin has been reported as potential anti-cathepsin agent in recent past. Low solubility of curcumin makes it an important subject for screening effect of different pharmaceutical excipients toward enhanced solubility. In the present work we report serum protein protecting and anti-cathepsin activities of 28 different formulations of curcumin. The formulations have been prepared using four ingredients used in traditional medicinal system. Milk has been found to enhance solubility to a significant level. Cow milk fat, sucrose and piperine exhibited positive cooperation. The results have been explained on the basis of chemical behavior of different ingredients.

The Rac2 GTPase contributes to cathepsin H-mediated protection against cytokine-induced apoptosis in insulin-secreting cells.

The type 1 diabetes (T1D) risk locus on chromosome 15q25.1 harbors the candidate gene CTSH (cathepsin H). We previously demonstrated that CTSH regulates ß-cell function in vitro and in vivo. CTSH overexpression protected insulin-secreting INS-1 cells against cytokine-induced apoptosis. The purpose of the present study was to identify the genes through which CTSH mediates its protective effects. Microarray analysis identified 63 annotated genes differentially expressed between CTSH-overexpressing INS-1 cells and control cells treated with interleukin-1ß and interferon-γ for up to 16h. Permutation test identified 10 significant genes across all time-points: Elmod1, Fam49a, Gas7, Gna15, Msrb3, Nox1, Ptgs1, Rac2, Scn7a and Ttn. Pathway analysis identified the "Inflammation mediated by chemokine and cytokine signaling pathway" with Gna15, Ptgs1 and Rac2 as significant. Knockdown of Rac2 abolished the protective effect of CTSH overexpression on cytokine-induced apoptosis, suggesting that the small GTPase and T1D candidate gene Rac2 contributes to the anti-apoptotic effect of CTSH.

Neonatal genetics of gene expression reveal potential origins of autoimmune and allergic disease risk.

Chronic immune-mediated diseases of adulthood often originate in early childhood. To investigate genetic associations between neonatal immunity and disease, we map expression quantitative trait loci (eQTLs) in resting myeloid cells and CD4+ T cells from cord blood samples, as well as in response to lipopolysaccharide (LPS) or phytohemagglutinin (PHA) stimulation, respectively. Cis-eQTLs are largely specific to cell type or stimulation, and 31% and 52% of genes with cis-eQTLs have response eQTLs (reQTLs) in myeloid cells and T cells, respectively. We identified cis regulatory factors acting as mediators of trans effects. There is extensive colocalisation between condition-specific neonatal cis-eQTLs and variants associated with immune-mediated diseases, in particular CTSH had widespread colocalisation across diseases. Mendelian randomisation shows causal neonatal gene expression effects on disease risk for BTN3A2, HLA-C and others. Our study elucidates the genetics of gene expression in neonatal immune cells, and aetiological origins of autoimmune and allergic diseases.

Cathepsin regulation on microglial function.

Microglia, the resident mononuclear phagocyte population in the brain, have long been implicated in the pathology of neurodegenerative age-associated disorders. However, activated microglia have now been identified as homeostatic keepers in the brain, because they are involved in the initiation and resolution of neuropathology. The complex roles of activated microglia appear to be linked to change from inflammatory and neurotoxic to anti-inflammatory and neuroprotective phenotypes. Increased expression and secretion of various cathepsins support roles of activated microglia in chronic neuroinflammation, the neurotoxic M1-like polarization and neuronal death. Moreover, changes in expression and localization of microglial cathepsin B play a critical role in the acceleration of the brain aging. Beyond the role as brain-resident macrophages, many lines of evidence have shown that microglia have essential roles in the maturation and maintenance of neuronal circuits in the developing and adult brain. Cathepsin S secreted from microglia induces the diurnal variation of spine density of cortical neurons though proteolytic modification of peri-synaptic extracellular matrix molecules. In this review, I highlight the emerging roles of cathepsins that support the roles of microglia in both normal healthy and pathological brains. In addition, I discuss cathepsin inhibitors as potential therapeutic targets for brain disorders.

Characterizing the Causal Pathway for Genetic Variants Associated with Neurological Phenotypes Using Human Brain-Derived Proteome Data.

Leveraging high-dimensional molecular datasets can help us develop mechanistic insight into associations between genetic variants and complex traits. In this study, we integrated human proteome data derived from brain tissue to evaluate whether targeted proteins putatively mediate the effects of genetic variants on seven neurological phenotypes (Alzheimer disease, amyotrophic lateral sclerosis, depression, insomnia, intelligence, neuroticism, and schizophrenia). Applying the principles of Mendelian randomization (MR) systematically across the genome highlighted 43 effects between genetically predicted proteins derived from the dorsolateral prefrontal cortex and these outcomes. Furthermore, genetic colocalization provided evidence that the same causal variant at 12 of these loci was responsible for variation in both protein and neurological phenotype. This included genes such as DCC, which encodes the netrin-1 receptor and has an important role in the development of the nervous system (p = 4.29 × 10-11 with neuroticism), as well as SARM1, which has been previously implicated in axonal degeneration (p = 1.76 × 10-08 with amyotrophic lateral sclerosis). We additionally conducted a phenome-wide MR study for each of these 12 genes to assess potential pleiotropic effects on 700 complex traits and diseases. Our findings suggest that genes such as SNX32, which was initially associated with increased risk of Alzheimer disease, may potentially influence other complex traits in the opposite direction. In contrast, genes such as CTSH (which was also associated with Alzheimer disease) and SARM1 may make worthwhile therapeutic targets because they did not have genetically predicted effects on any of the other phenotypes after correcting for multiple testing.

Effects of Five Substances with Different Modes of Action on Cathepsin H, C and L Activities in Zebrafish Embryos.

Cathepsins have been proposed as biomarkers of chemical exposure in the zebrafish embryo model but it is unclear whether they can also be used to detect sublethal stress. The present study evaluates three cathepsin types as candidate biomarkers in zebrafish embryos. In addition to other functions, cathepsins are also involved in yolk lysosomal processes for the internal nutrition of embryos of oviparous animals until external feeding starts. The baseline enzyme activity of cathepsin types H, C and L during the embryonic development of zebrafish in the first 96 h post fertilisation was studied. Secondly, the effect of leupeptin, a known cathepsin inhibitor, and four embryotoxic xenobiotic compounds with different modes of action (phenanthrene-baseline toxicity; rotenone-an inhibitor of electron transport chain in mitochondria; DNOC (Dinitro-ortho-cresol)-an inhibitor of ATP synthesis; and tebuconazole-a sterol biosynthesis inhibitor) on in vivo cathepsin H, C and L total activities have been tested. The positive control leupeptin showed effects on cathepsin L at a 20-fold lower concentration compared to the respective LC50 (0.4 mM) of the zebrafish embryo assay (FET). The observed effects on the enzyme activity of the four other xenobiotics were not or just slightly more sensitive (factor of 1.5 to 3), but the differences did not reach statistical significance. Results of this study indicate that the analysed cathepsins are not susceptible to toxins other than the known peptide-like inhibitors. However, specific cathepsin inhibitors might be identified using the zebrafish embryo.

Increased cystatin F levels correlate with decreased cytotoxicity of cytotoxic T cells.

Background Cystatin F is a protein inhibitor of cysteine peptidases, expressed predominantly in immune cells and localised in endosomal/lysosomal compartments. In cytotoxic immune cells cystatin F inhibits both the major pro-granzyme convertases, cathepsins C and H that activate granzymes, and cathepsin L, that acts as perforin activator. Since perforin and granzymes are crucial molecules for target cell killing by cytotoxic lymphocytes, defects in the activation of either granzymes or perforin can affect their cytotoxic potential. Materials and methods Levels of cystatin F were assessed by western blot and interactions of cystatin F with cathepsins C, H and L were analysed by immunoprecipitation and confocal microscopy. In TALL-104 cells specific activities of the cathepsins and granzyme B were determined using peptide substrates. Results Two models of reduced T cell cytotoxicity of TALL-104 cell line were established, either by treatment by ionomycin or by immunosuppressive transforming growth factor beta. Reduced cytotoxicity correlated with increased levels of cystatin F and with attenuated activities of cathepsins C, H and L and of granzyme B. Co-localisation of cystatin F and cathepsins C, H and L and interactions between cystatin F and cathepsins C and H were demonstrated. Conclusions Cystatin F is designated as a possible regulator of T cell cytotoxicity, similar to its role in natural killer cells.

Identification of loci where DNA methylation potentially mediates genetic risk of type 1 diabetes.

The risk of Type 1 Diabetes (T1D) comprises both genetic and environmental components. We investigated whether genetic susceptibility to T1D could be mediated by changes in DNA methylation, an epigenetic mechanism that potentially plays a role in autoimmune diabetes. From enrichment analysis, we found that there was a common genetic influence for both DNA methylation and T1D across the genome, implying that methylation could be either on the causal pathway to T1D or a non-causal biomarker of T1D genetic risk. Using data from a general population comprising blood samples taken at birth (n = 844), childhood (n = 846) and adolescence (n = 907), we then evaluated the associations between 64 top GWAS single nucleotide polymorphisms (SNPs) and DNA methylation levels at 55 non-HLA loci. We identified 95 proximal SNP-cytosine phosphate guanine (CpG) pairs (cis) and 1 distal SNP-CpG association (trans) consistently at birth, childhood, and adolescence. Combining genetic co-localization and Mendelian Randomization analysis, we provided evidence that at 5 loci, ITGB3BP, AFF3, PTPN2, CTSH and CTLA4, DNA methylation is potentially mediating the genetic risk of T1D mainly by influencing local gene expression.

Crystal structures of human procathepsin H.

Cathepsin H is a member of the papain superfamily of lysosomal cysteine proteases. It is the only known aminopeptidase in the family and is reported to be involved in cancer and other major diseases. Like many other proteases, it is synthesized as an inactive proenzyme. Although the crystal structure of mature porcine cathepsin H revealed the binding of the mini-chain and provided structural basis for the aminopeptidase activity, detailed structural and functional information on the inhibition and activation of procathepsin H has been elusive. Here we present the crystal structures of human procathepsin H at 2.00 Å and 1.66 Å resolution. These structures allow us to explore in detail the molecular basis for the inhibition of the mature domain by the prodomain. Comparison with cathepsin H structure reveals how mini-chain reorients upon activation. We further demonstrate that procathepsin H is not auto-activated but can be trans-activated by cathepsin L.