P values less than 0.05 were considered statistically significant. | Mutations in Borealin and clinical data. MNW and TWL performed bioinformatics analysis for genome sequencing and targeted next-generation sequencing data. Kalender Stage 7 islet-like aggregates differentiated from WT, KO, and YIPF5Ile98Ser H1 stem cells were implanted under the kidney capsule of immunocompromised NOD/SCID-γ mice, and their function was monitored by measurement of the serum levels of human C-peptide. | Insulin gene mutations as a cause of permanent neonatal diabetes. Functional studies show that YIPF5 deficiency affects β cell function by enhancing ER stress and sensitizing human β cells to ER stress–induced apoptosis. JCI JCI 13Institute of Interdisciplinary Research (IRIBHM), ULB Neuroscience Institute, Université Libre de Bruxelles, Brussels, Belgium. It is therefore possible that this phenotypic variability is directly linked to the severity of the mutation; however, our cohort of patients with homozygous YIPF5 mutations is currently too small to allow accurate estimation of any genotype-phenotype relationship. Balboa D, et al. In conclusion, we report homozygous mutations in YIPF5 as the genetic cause of an autosomal recessive syndrome characterized by microcephaly, epilepsy, and neonatal/early-onset diabetes. Recessive Mutations in RTN4IP1 Cause Isolated and Syndromic Optic Neuropathies. Paired 2-way ANOVA or mixed-model analysis (in case of missing values) followed by Bonferroni post hoc test. JCI |, Find articles by Scale bars: 25 μm. JCI PBMCs from patients IIIa and IIIb were reprogrammed into iPSCs using Sendai virus. iPSCs were cultured in Matrigel-coated plates (Corning) in E8 medium as previously described (61, 62). PubMed Google Scholar, Find articles by |, Find articles by |, Find articles by SavvyVcfHomozygosity was used to identify large (>3 Mb) homozygous regions in the genome sequencing data (https://github.com/rdemolgen/SavvySuite). Cosentino C, et al. Demarez, C. Improved genetic testing for monogenic diabetes using targeted next-generation sequencing. Google Scholar, Find articles by The bioinformatics tools SIFT, PolyPhen-2, MutationTaster, and Align GVGD were accessed through the Alamut software (Interactive Biosoftware) to predict the effect of variants on the YIPF5 protein. The KO cell line was characterized for pluripotency using immunocytochemistry for OCT4, TRA1-60, and SSEA4, and for gene expression levels of OCT4, SOX2, and NANOG by qPCR. These data strongly suggest that while β cells are highly dependent on YIPF5 function, α cells appear to be able to survive without YIPF5. Aggregates were lysed by sonication and acid-ethanol for determination of insulin and DNA contents. Conflict of interest: The authors have declared that no conflict of interest exists. Tavtigian SV, et al. | Recently, pathogenic variants in 3 genes (TRMT10A [ref. Abdulkarim B, et al. YIPF5 is expressed in human pancreatic tissue and brain. Variant confirmation and cosegregation in family members were performed by Sanger sequencing (see Supplemental Table 4 for primer sequence). (Ile98Ser)-derived islet-like aggregates showed a significant decrease in human C-peptide levels associated with increased proinsulin accumulation and signs of ER stress after implantation, demonstrating the evolution of cellular pathology with further maturation of the stem cell–derived β cells. | The cerebrospinal fluid provides a proliferative niche for neural progenitor cells. | Functional studies in human β cell models highlight the importance of ER-to-Golgi trafficking in β cells and neurons. PubMed 7Yeditepe University Hospital, Istanbul, Turkey. Yip1A, a novel host factor for the activation of the IRE1 pathway of the unfolded protein response during Brucella infection. (A) Partial pedigrees and summary of clinical features of the 6 patients with homozygous YIPF5 mutations. In order to examine whether the induction of CHOP, a proapoptotic transcription factor in the PERK branch of the ER stress response, sensitizes YIPF5-deficient β cells to apoptosis, we double-knocked-down CHOP and YIPF5 (Figure 3I and Supplemental Figure 2H). Google Scholar In addition, the human YIPF5 gene appears to be intolerant to loss-of-function variants based on gnomAD gene constraints (pLI = 0.99), further supporting that loss-of-function variants are likely strongly deleterious in humans. | |, Find articles by This is an open access article published under the terms of the Creative Commons Attribution 4.0 International License. In addition to the KO, we generated an isogenic YIPF5Ile98Ser mutation using CRISPR/Cpf1–mediated homology-directed repair (HDR) (Supplemental Figure 5). Further studies will be needed in order to elucidate which pathways regulate YIPF5 expression in response to ER stressors such as brefeldin A and whether Golgi stress response transcription factors such as CREB3 (36) and TFE3 (37) are involved. Testing for the mutations in family members confirmed that the parents were all heterozygous for the mutations and that patient III’s affected sister was also homozygous for the p.(Ile98Ser) variant (Figure 1A). sequenced the genome of two probands with a rare neonatal diabetes subtype that also associated with microcephaly and epilepsy. toutes les réponses pour lettre: changement poste cause maladie professionelle. Google Scholar, Find articles by PubMed in: The p.(Ile98Ser) mutation did not affect proinsulin and insulin content (Supplemental Figure 12, A and B) nor glucose- and/or forskolin-stimulated insulin secretion (Supplemental Figure 12C). hESC culturing, differentiation into β cells, and chemical ER-stress induction. in: Understanding the genetic basis of rare diabetes subtypes highlights fundamental biological processes in β cells. in: 9Dicle University, Faculty of Medicine, Department of Pediatric Endocrinology, Diyarbakır, Turkey. iPSC culturing and differentiation into β cells. Functional characterisation of the YIPF protein family in mammalian cells. Jokitalo, E. De Franco E, et al. Partial YIPF5 silencing in EndoC-βH1 cells and a patient mutation in stem cells increased the β cell sensitivity to ER stress–induced apoptosis. 18Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy. Dominant ER stress-inducing. To further assess the impact of one of the YIPF5 missense mutations in a directly patient-relevant model, we generated patients’ induced pluripotent stem cells (iPSCs) and differentiated them into pancreatic endocrine cells. Reduced endoplasmic reticulum (ER)-to-Golgi protein trafficking contributes to ER stress in lipotoxic mouse beta cells by promoting protein overload. (C) Insulin secretion expressed as percentage of total insulin content. Among the residues investigated were p.Gly97 and p.Ile98, which are mutated in families III and V. Substitutions at these positions were not found to result in whorl formation in the study by Dykstra et al. While ubiquitously expressed in human tissues, (partial) YIPF5 loss of function caused by the mutations results in a β cell– and brain-specific phenotype in the patients, possibly pointing to YIPF5 cargo specificity; it will be of interest to examine this in future studies. Clinical evaluation. Patient II had genome-wide homozygosity of 6.9%, and rare homozygous coding variants were identified in 10 genes (see Supplemental Tables 1 and 2 and Supplemental Figure 1A; supplemental material available online with this article; https://doi.org/10.1172/JCI141455DS1). pour la maladie professionnel mon médecin en avait fait la demande,et sa été refusé par le. Address correspondence to: Andrew T. Hattersley, University of Exeter Medical School, RILD Building, Barrack Road, Exeter EX2 5DW, United Kingdom. in: 6Electron Microscopy Unit, Institute of Biotechnology, University of Helsinki, Helsinki, Finland. RNA in situ hybridization. JCI |, Find articles by JCI YIPF5 and YIF1A recycle between the ER and the Golgi apparatus and are involved in the maintenance of the Golgi structure. Scale bars: 1 μm. For the embryoid body assay, iPSCs reaching 60%–70% confluence were detached with 0.5 mM EDTA (Life Technologies), resuspended in E8 medium containing 10 μM ROCK inhibitor (Stemcell Technologies), and transferred to super-low-attachment plate (Corning) on a rotating platform to form aggregates. Differentiation was started 24 hours later and proceeded through a 7-stage differentiation protocol (stages 1–4 in adherent culture, stage 5 in AggreWell [34421, Stemcell Technologies], and stages 6 and 7 in suspension culture). Identification of homozygous YIPF5 mutations in 6 patients with neonatal diabetes, severe microcephaly, and epilepsy. Google Scholar in: Fifty hESC-derived stage 7 aggregates were washed twice with glucose-free Krebs buffer and preincubated on a rotating platform with 1 mL 3.3 mM glucose-containing Krebs buffer for 1 hour. These KO β cells showed a strong increase in the proinsulin/insulin ratio, consistent with proinsulin not being transported into the Golgi, and triggered the ER stress response. De l'influence des causes morales dans les maladies: thèse pour le doctorat en médecine: HAMAIDE-L-A: Amazon.com.au: Books JCI We identified 3 homozygous YIPF5 missense mutations, p.(Ile98Ser), p.(Trp218Arg), and p.(Gly97Val), in 3 cases. (E) Percentage of INS+BiPhi cells per total number of INS+ cells (n = 3–6). Learn about genetic conditions, genes, chromosomes, and more. (D) Percentage of cytoplasmic area covered by proinsulin or insulin in insulin-positive cells (n = 3–4). PubMed Embryoid bodies were plated on Matrigel-coated ICC chambers (15–20 per well) for 2 weeks, with medium refreshed every second day, and fixed in PFA 4% for immunocytochemistry. iPSCs from patients IIIa and IIIb differentiated into β cells are sensitive to ER stress–induced apoptosis. A CREB3-ARF4 signalling pathway mediates the response to Golgi stress and susceptibility to pathogens. Taken together, our results demonstrate that the p.(Ile98Ser) YIPF5 mutation does not compromise differentiation and function of β cells but affects cell survival by sensitizing them to ER stress–induced apoptosis. Pachera, N. (G) Insulin content of stage 7 differentiated cells normalized to micrograms DNA of β cells (n = 3–8). In EndoC-βH1 cells, YIPF5 expression was strongly upregulated by brefeldin A, which blocks ER-to-Golgi transport, suggesting that transcriptional induction of YIPF5 could be part of a compensatory mechanism to overcome the inhibition of trafficking. Further information can be found in Supplemental Methods. YIPF5 was silenced using 2 siRNAs targeting different sequences of YIPF5 (si1 SI04182745 and si2 SI04344984, Qiagen). Activating germline mutations in STAT3 cause early-onset multi-organ autoimmune disease. YIPF5 is expressed in human islets and brain. McKenzie MD, et al. JCI Find articles by Comprehensive statistical study of 452 BRCA1 missense substitutions with classification of eight recurrent substitutions as neutral. Gladwyn-Ng I, et al. Proinsulin accumulation, increased ER stress signaling, and reduced insulin content in YIPF5-knockout stem cell–derived β cells. We thank the families for participating in the study. Ellard, S. Study approval. Yoshida Y, et al. 12], PPP1R15B [ref. |, Find articles by Diagramme à utiliser pour dessiner les châtaignes (si moins de 3 épis ou autres marques) B. Formulaire de mutation. We believe this is the first report of mutations disrupting the ER-to-Golgi trafficking, resulting in diabetes. Differentiation was started 24 hours later. People who inherit two copies of C677T have a higher risk for having a child with a neural tube defect. Insulin secretion results are presented as insulin released after cell mass normalization using DNA content of the β cell percentage assessed by FACS. Expression was found in both progenitor (ventricular zone) and neuronal (intermediate zone and cortical plate) compartments. The percentage of β cells of the human islet preparations was 59% ± 5%, as determined by insulin immunofluorescence. Mutations in different functional domains of FLNC can cause various clinical phenotypes. The muscle MRI findings are similar to those observed in FLNC-myofibrillar myopathy (MIM #609524). (Ile98Ser)-β cells under basal condition (Figure 6E). PubMed | in: Nakagawa H, Hazama K, Ishida K, Komori M, Nishimura K, Matsuo S. Inhibition of PLD1 activity causes ER stress via regulation of COPII vesicle formation. 1Institute of Biomedical and Clinical Science, University of Exeter Medical School, Exeter, United Kingdom. We believe that utilizing registries and biobanks to reveal other monogenic atypical forms of diabetes is an important approach to gaining insight and suggest that an insulin sensitizer may alleviate ER stress associated with YIPF5 disruption by decreasing the demand for insulin secretion. A β cell–specific knockout of cTAGE5, a TANGO1-interacting protein, has been previously shown to impair proinsulin trafficking, induce ER stress, and cause impaired glucose tolerance in mice (49). | PubMed A missense mutation in PPP1R15B causes a syndrome including diabetes, short stature, and microcephaly. Moore F, Cunha DA, Mulder H, Eizirik DL. (F–H) Immunohistochemistry of grafts for glucagon (GCG) and insulin (INS) (F), proinsulin (PROINS) and insulin (INS) (G), and BiP and insulin (INS) (H) 3 months after implantation. Shakeri, H. ... for which only mutations in TMEM126A and ACO2 are known. Our qPCR analysis detected abundant expression in pancreatic tissue, islets, β cells, and brain (Figure 2A). After 1 week of stage 7, aggregates were exposed to 1 μM thapsigargin, 5 μg/mL tunicamycin, or 0.01 μg/mL brefeldin A in stage 7 medium. | In HeLa and CaSki cells, another human cervical cancer cell line, YIPF5 constitutively activated IRE1 and PERK signaling, with YIPF5-depleted cells showing reduced IRE1 phosphorylation, lower PERK mRNA and protein expression, and less PERK phosphorylation and downstream signaling (39). While disruption of ER-to-Golgi trafficking is known to result in at least 10 different neurological disorders (47), its involvement in the etiology of diabetes has been less clear. Human pancreata not suitable for clinical purposes were collected from nondiabetic, brain-dead organ donors after written informed consent from next of kin, and handled as described (64) with the approval of the Ethical Committee, University of Pisa. télécharger ce modèle de lettre : vous rencontrez des problèmes de santé et demandez un changement de service à votre employeur. Transient transfection was performed using 30 nM siRNA and Lipofectamine RNAiMAX lipid reagent (Invitrogen/Life Technologies) as previously described (54). télécharger ce modèle de lettre : vous rencontrez des problèmes de santé et demandez un changement de service à votre employeur. The authors revealed mutations … | JCI (C and D) Apoptosis was assessed by staining with DNA-binding dyes in vehicle- (DMSO-)treated, thapsigargin-treated, and tunicamycin-treated control and corrected (n = 10) and patient (n = 6–7) stage 7 aggregates (C) or by luminescence produced by annexin V binding in time course experiments (means ± SEM; n = 10 control and corrected lines and n = 5 patient lines) (D). | Yellow arrowheads point at insulin granules, red arrowheads at glucagon granules, and green arrowheads at ER. The percentage of cytoplasmic area stained for proinsulin per β cell of the KO was 5.5-fold higher than in their WT counterparts, while the percentage of insulin area was 70% less in the KO β cells (Figure 4C). in: | PubMed Log In. Brefeldin A treatment markedly induced YIPF5 mRNA expression in EndoC-βH1 cells and human islets (Supplemental Figure 2, A and B); a trend for YIPF5 protein induction was seen in EndoC-βH1 cells (approximately 2-fold; n = 4). YIPF5 was ubiquitously expressed, with abundant expression in pancreatic tissue, islets, β cells, and brain (Figure 2A). Family F1 was a consanguineous French family originating from Sri Lanka and of Tamil ethnicity. = Banques de données de mutations: enjeux et perspectives pour les maladies génétiques orphelines. in: The next day, embryoid bodies were resuspended in DMEM/F-12 medium (Gibco) containing Glutamax (Gibco), 10% KSR (Life Technologies), 1% NEAA (Thermo Fisher Scientific), 0.1 mM β-mercaptoethanol (Gibco), and 1% penicillin/streptomycin. Log In. Dickinson ME, et al. PubMed The milder phenotype observed in patient-iPSC- and ESCp. PubMed (A) Representative immunostaining of dispersed stage 7 aggregates stained for insulin (INS, green) and glucagon (GCG, red). | The expression in the choroid plexus is in line with potential control of brain morphogenesis and size, as this structure was recently found to secrete important morphogens and growth factors during embryonic brain development (42, 43). YIPF5 is expressed during human brain development, in adult brain and pancreatic islets. fermé pour cause de maladie Übersetzung, Franzosisch - Englisch Wörterbuch, Siehe auch , biespiele, konjugation Études sur la bière, ses maladies, causes qui les provoquent, procédé pour la rendre inaltérable; avec une théorie nouvelle de la fermentation; by Pasteur, Louis, 1822-1895 Genetic analysis. Medium was refreshed with E6 medium (Gibco), and cells were transferred to Matrigel-coated plates (Corning BV, Life Sciences). YIPF5 deficiency sensitizes human β cells to ER stress–induced apoptosis. The differentiation was continued in microwells. | Montaser, H. PubMed Google Scholar The missense mutation identified in patient I, p.(Ala181Val), was predicted to be tolerated by 2 of 4 in silico tools used, suggesting the possibility that this variant has a less severe effect on protein function. Aggregates were incubated sequentially in 3.3 mM glucose, 20 mM glucose, and 3.3 mM plus 30 mM KCl for periods of 30 minutes. Data points represent independent experiments. Samples were sequenced on an Illumina HiSeq 2500 with a mean read depth of 38.3 for patient I and 33.6 for patient II. HV and EJ performed the electron microscopic analyses. For all 6 patients, the birth weight was low (median standard deviation score –1.85 (–1.99 to –1.72)), consistent with reduced insulin secretion in utero. | Transplantations were performed on 3- to 9-month-old mice as described previously (60). In time course experiments, data are shown as mean ± SEM. At present, nearly 20% of neonatal diabetes cases have unknown causes. Human islets from nondiabetic organ donors (n = 4, 2 female and 2 male donors; age 62 ± 9 years; BMI 27 ± 3 kg/m2; cause of death: 3 cerebral hemorrhage, 1 cardiovascular disease) were isolated by collagenase digestion and density gradient purification, and cultured as previously described (53). Flanagan SE, et al. Taniguchi M, et al. One of the pathways known to be crucial for the function of both β and brain cells is the endoplasmic reticulum (ER) stress response. modèle gratuit de lettre pour demander à changer de poste ou de service en ni pour une cause prévue par le code du travail (comme pour une grossesse, in: (H) Percentage of INS+ cells at week 2 of stage 7 (n = 3–4). Similar to previously identified mutations in genes encoding proteins involved in SMC contraction, PRKG1 c.530G>A leads to PKG-1α alteration p.Arg177Gln, which increases PKG-1α activity and causes decreased phosphorylation of the RLC and thus should decrease vascular SMCs contraction. Kano F, et al. | | Error bars represent SD from the mean. Mouse blood samples were collected monthly from the saphenous vein using heparinized capillary tubes. JCI To try to account for this possibility in our functional experiments, we used a model of complete loss of function (complete KO in hESCs), as well as models of incomplete loss of function [50%–75% knockdown in human EndoC-βH1 cells, iPSCs derived from 2 of the cases with the p.(Ile98Ser) mutation, and hESCs harboring the same homozygous p.(Ile98Ser) mutation]. Saarimäki-Vire, J. Støy J, et al. Furthermore, impaired ER-to-Golgi trafficking may play a role in β cell dysfunction and death caused by environmental insults in type 2 diabetes, as exposure of β cells to the saturated free fatty acid palmitate reduces protein trafficking, thereby contributing to ER stress (19). Sa première expérience en recherche fondamentale sur la maladie Alzheimer lors de son internat, le neurologue David Wallon ne l'oubliera jamais ! | PubMed The vehicle DMSO was added to the control condition in all experiments. ATH, BH, MNO, EU, RY, TG, MY, KP, and BA analyzed the clinical data. Aggregates were exposed to 1.6 mM glucose, 16.7 mM glucose, or 16.7 mM glucose plus 10 μM forskolin for 1 hour. Insulin content and secretion. PBMC reprogramming into iPSCs and iPSC quality control. Google Scholar, Find articles by PubMed Fantuzzi, F. Sawatani, T. In keeping with the results in YIPF5-depleted EndoC-βH1 cells, the proapoptotic BCL-2 family members DP5 and PUMA were induced in ER-stressed YIPF5-mutant cells (Figure 6E). The YIPF5 patients’ iPSCs differentiated into stage 7 islet cells with somewhat fewer insulin-positive cells and slightly more glucagon-expressing cells compared with healthy control or corrected iPSCs (Figure 6, A and B). All 4 affected individuals also had insulin-dependent diabetes, highlighting the importance of the mechanisms regulating cargo exit from the ER for β cell function (48). Google Scholar cause … All compounds were from Sigma-Aldrich. De Franco, E. (A) YIPF5 mRNA expression was measured by qPCR in human tissues (n = 2–3), EndoC-βH1 cells (n = 15), and human islets (n = 4) and normalized to the geometric mean of the reference genes ACTB, GAPDH, and OAZ1. 17Université de Paris, Faculté de Médecine Paris–Diderot, U958, Paris, France. There was no significant family history of diabetes for any of the patients. | Neonatal hypoglycemia, early-onset diabetes and hypopituitarism due to the mutation in EIF2S3 gene causing MEHMO syndrome. The RNP components (Alt-R A.s. Cpf1 Ultra and crRNA) were purchased from IDT and prepared based on the manufacturer’s protocol. PUMA expression was also induced by YIPF5 silencing (Supplemental Figure 2G), while BIM expression was not altered. P2008/313). Cells were seeded at a density of 1.8 million cells per 3.5 cm well on Matrigel-coated plates with E8 medium containing 10 μM ROCK inhibitor. in: The H1 wild-type (WT), KO, and YIPF5Ile98Ser cells differentiated normally until the pancreatic endocrine stage. PubMed | Individual symbols represent independent experiments, and box plots show the median by a horizontal line, 25th and 75th percentiles at the bottom and top of the boxes, and minimum and maximum values by whiskers. JCI iPSCs from patients IIIa and IIIb differentiated into β cells are sensitive to ER stress–induced apoptosis. A dynamic unfolded protein response contributes to the control of cortical neurogenesis.
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