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How cytokine-driven changes in chromatin topology are converted into gene regulatory circuits during inflammation still remains unclear. Here, we show that interleukin (IL)-1α induces acute and widespread changes in chromatin accessibility via the TAK1 kinase and NF-κB at regions that are highly enriched for inflammatory disease-relevant SNPs. Two enhancers in the extended chemokine locus on human chromosome 4 regulate the IL-1α-inducible IL8 and CXCL1-3 genes. Both enhancers engage in dynamic spatial interactions with gene promoters in an IL-1α/TAK1-inducible manner. Microdeletions of p65-binding sites in either of the two enhancers impair NF-κB recruitment, suppress activation and biallelic transcription of the IL8/CXCL2 genes, and reshuffle higher-order chromatin interactions as judged by i4C interactome profiles. Notably, these findings support a dominant role of the IL8 "master" enhancer in the regulation of sustained IL-1α signaling, as well as for IL-8 and IL-6 secretion. CRISPR-guided transactivation of the IL8 locus or cross-TAD regulation by TNFα-responsive enhancers in a different model locus supports the existence of complex enhancer hierarchies in response to cytokine stimulation that prime and orchestrate proinflammatory chromatin responses downstream of NF-κB.
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Cromatina/metabolismo , Elementos de Facilitación Genéticos/genética , Regulación de la Expresión Génica/efectos de los fármacos , Mediadores de Inflamación/metabolismo , Interleucina-1alfa/farmacología , Quinasas Quinasa Quinasa PAM/metabolismo , FN-kappa B/metabolismo , Sitios de Unión , Células Cultivadas , Quimiocinas/metabolismo , Cromatina/química , Cromatina/genética , Células HeLa , Humanos , Quinasas Quinasa Quinasa PAM/genética , FN-kappa B/genética , Transducción de Señal , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
BACKGROUND: Cleidocranial dysplasia (CCD) is a rare skeletal dysplasia with significant clinical variability. Patients with CCD typically present with delayed closure of fontanels and cranial sutures, dental anomalies, clavicular hypoplasia or aplasia and short stature. Runt-related transcription factor 2 (RUNX2) is currently the only known disease-causing gene for CCD, but several studies have suggested locus heterogeneity. METHODS: The cohort consists of eight subjects from five unrelated families partially identified through GeneMatcher. Exome or genome sequencing was applied and in two subjects the effect of the variant was investigated at RNA level. RESULTS: In each subject a heterozygous pathogenic variant in CBFB was detected, whereas no genomic alteration involving RUNX2 was found. Three CBFB variants (one splice site alteration, one nonsense variant, one 2 bp duplication) were shown to result in a premature stop codon. A large intragenic deletion was found to delete exon 4, without affecting CBFB expression. The effect of a second splice site variant could not be determined but most likely results in a shortened or absent protein. Affected individuals showed similarities with RUNX2-related CCD, including dental and clavicular abnormalities. Normal stature and neurocognitive problems were however distinguishing features. CBFB encodes the core-binding factor ß subunit, which can interact with all RUNX proteins (RUNX1, RUNX2, RUNX3) to form heterodimeric transcription factors. This may explain the phenotypic differences between CBFB-related and RUNX2-related CCD. CONCLUSION: We confirm the previously suggested locus heterogeneity for CCD by identifying five pathogenic variants in CBFB in a cohort of eight individuals with clinical and radiographic features reminiscent of CCD.
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Displasia Cleidocraneal , Subunidad beta del Factor de Unión al Sitio Principal , Humanos , Secuencia de Bases , Displasia Cleidocraneal/genética , Displasia Cleidocraneal/patología , Codón sin Sentido , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad beta del Factor de Unión al Sitio Principal/genética , ExonesRESUMEN
Depending on its location, size, and proximity to the cardiac structures, an intrapericardial teratoma may lead to severe circulatory disturbances and even fetal demise. A 34-year-old G2P1 presented at 20w5d with a solid cystic mass in the right thorax of the fetus, originating from the right atrium or lung, with signs of non-immune fetal hydrops, soon resulting in intrauterine fetal death. Detailed post-mortem autopsy revealed signs of hydrops fetalis universalis due to a spherical tumor mass originating from the aortic root. Histologic examination of the tumor showed the characteristic morphology of a teratoma. A 1.6-Mb microdeletion at 3q29 was identified by single nucleotide polymorphism array. This is the first report presenting the diagnosis of an intrapericardial teratoma in a fetus with a microdeletion of 3q29. Intrapericardial teratoma has a poor prognosis and the fetal outcome relies on the development of hydrops. A post-mortem examination is essential in order to make a definitive diagnosis, which underlines the status of the fetal pathologist and the need for interdisciplinary cooperation.
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OBJECTIVE: We aimed to investigate how the presence of fetal anomalies and different X chromosome variants influences Cell-free DNA (cfDNA) screening results for monosomy X. METHODS: From a multicenter retrospective survey on 673 pregnancies with prenatally suspected or confirmed Turner syndrome, we analyzed the subgroup for which prenatal cfDNA screening and karyotype results were available. A cfDNA screening result was defined as true positive (TP) when confirmatory testing showed 45,X or an X-chromosome variant. RESULTS: We had cfDNA results, karyotype, and phenotype data for 55 pregnancies. cfDNA results were high risk for monosomy X in 48/55, of which 23 were TP and 25 were false positive (FP). 32/48 high-risk cfDNA cases did not show fetal anomalies. Of these, 7 were TP. All were X-chromosome variants. All 16 fetuses with high-risk cfDNA result and ultrasound anomalies were TP. Of fetuses with abnormalities, those with 45,X more often had fetal hydrops/cystic hygroma, whereas those with "variant" karyotypes had different anomalies. CONCLUSION: Both, 45,X or X-chromosome variants can be detected after a high-risk cfDNA result for monosomy X. When there are fetal anomalies, the result is more likely a TP. In the absence of fetal anomalies, it is most often an FP or X-chromosome variant.
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Ácidos Nucleicos Libres de Células , Síndrome de Down , Síndrome de Turner , Embarazo , Humanos , Femenino , Síndrome de Turner/diagnóstico , Síndrome de Turner/genética , Síndrome de Down/diagnóstico , Estudios Retrospectivos , Cromosoma X , Diagnóstico Prenatal/métodosRESUMEN
OBJECTIVE: Precursors of peptide hormones undergo posttranslational modifications within the trans-Golgi network (TGN). Dysfunction of proteins involved at different steps of this process cause several complex syndromes affecting the central nervous system (CNS). We aimed to clarify the genetic cause in a group of patients characterized by hypopituitarism in combination with brain atrophy, thin corpus callosum, severe developmental delay, visual impairment, and epilepsy. METHODS: Whole exome sequencing was performed in seven individuals of six unrelated families with these features. Postmortem histopathological and HID1 expression analysis of brain tissue and pituitary gland were conducted in one patient. Functional consequences of the homozygous HID1 variant p.R433W were investigated by Seahorse XF Assay in fibroblasts of two patients. RESULTS: Bi-allelic variants in the gene HID1 domain-containing protein 1 (HID1) were identified in all patients. Postmortem examination confirmed cerebral atrophy with enlarged lateral ventricles. Markedly reduced expression of pituitary hormones was found in pituitary gland tissue. Colocalization of HID1 protein with the TGN was not altered in fibroblasts of patients compared to controls, while the extracellular acidification rate upon stimulation with potassium chloride was significantly reduced in patient fibroblasts compared to controls. INTERPRETATION: Our findings indicate that mutations in HID1 cause an early infantile encephalopathy with hypopituitarism as the leading presentation, and expand the list of syndromic CNS diseases caused by interference of TGN function. ANN NEUROL 2021;90:149-164.
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Encefalopatías/genética , Epilepsia/genética , Hipopituitarismo/genética , Alelos , Encefalopatías/patología , Preescolar , Epilepsia/patología , Femenino , Humanos , Hipopituitarismo/patología , Lactante , Masculino , Hipófisis/patología , Secuenciación del Exoma , Adulto JovenRESUMEN
Given the intimate link between inflammation and dysregulated cell proliferation in cancer, we investigated cytokine-triggered gene expression in different cell cycle stages. Transcriptome analysis revealed that G1 release through cyclin-dependent kinase 6 (CDK6) and CDK4 primes and cooperates with the cytokine-driven gene response. CDK6 physically and functionally interacts with the NF-κB subunit p65 in the nucleus and is found at promoters of many transcriptionally active NF-κB target genes. CDK6 recruitment to distinct chromatin regions of inflammatory genes was essential for proper loading of p65 to its cognate binding sites and for the function of p65 coactivators, such as TRIP6. Furthermore, cytokine-inducible nuclear translocation and chromatin association of CDK6 depends on the kinase activity of TAK1 and p38. These results have widespread biological implications, as aberrant CDK6 expression or activation that is frequently observed in human tumors modulates NF-κB to shape the cytokine and chemokine repertoires in chronic inflammation and cancer.
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Cromatina/metabolismo , Quinasa 6 Dependiente de la Ciclina/fisiología , FN-kappa B/genética , Ciclo Celular/genética , Quinasa 4 Dependiente de la Ciclina/genética , Quinasa 4 Dependiente de la Ciclina/metabolismo , Quinasa 4 Dependiente de la Ciclina/fisiología , Quinasa 6 Dependiente de la Ciclina/análisis , Quinasa 6 Dependiente de la Ciclina/metabolismo , Regulación de la Expresión Génica , Células HeLa , Humanos , Interleucina-1/metabolismo , Interleucina-1/fisiología , Interleucina-8/genética , Interleucina-8/metabolismo , Quinasas Quinasa Quinasa PAM/genética , Quinasas Quinasa Quinasa PAM/metabolismo , Quinasas Quinasa Quinasa PAM/fisiología , Regiones Promotoras Genéticas , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismo , Factor de Transcripción ReIA/fisiología , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/fisiologíaRESUMEN
Influenza A viruses (IAVs) quickly adapt to new environments and are well known to cross species barriers. To reveal a molecular basis for these phenomena, we compared the Ser/Thr and Tyr phosphoproteomes of murine lung epithelial cells early and late after infection with mouse-adapted SC35M virus or its nonadapted SC35 counterpart. With this analysis we identified a large set of upregulated Ser/Thr phosphorylations common to both viral genotypes, while Tyr phosphorylations showed little overlap. Most of the proteins undergoing massive changes of phosphorylation in response to both viruses regulate chromatin structure, RNA metabolism, and cell adhesion, including a focal adhesion kinase (FAK)-regulated network mediating the regulation of actin dynamics. IAV also affected phosphorylation of activation loops of 37 protein kinases, including FAK and several phosphatases, many of which were not previously implicated in influenza virus infection. Inhibition of FAK proved its contribution to IAV infection. Novel phosphorylation sites were found on IAV-encoded proteins, and the functional analysis of selected phosphorylation sites showed that they either support (NA Ser178) or inhibit (PB1 Thr223) virus propagation. Together, these data allow novel insights into IAV-triggered regulatory phosphorylation circuits and signaling networks.IMPORTANCE Infection with IAVs leads to the induction of complex signaling cascades, which apparently serve two opposing functions. On the one hand, the virus highjacks cellular signaling cascades in order to support its propagation; on the other hand, the host cell triggers antiviral signaling networks. Here we focused on IAV-triggered phosphorylation events in a systematic fashion by deep sequencing of the phosphoproteomes. This study revealed a plethora of newly phosphorylated proteins. We also identified 37 protein kinases and a range of phosphatases that are activated or inactivated following IAV infection. Moreover, we identified new phosphorylation sites on IAV-encoded proteins. Some of these phosphorylations support the enzymatic function of viral components, while other phosphorylations are inhibitory, as exemplified by PB1 Thr223 modification. Our global characterization of IAV-triggered patterns of phospho-proteins provides a rich resource to further understand host responses to infection at the level of phosphorylation-dependent signaling networks.
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Antivirales/farmacología , Virus de la Influenza A/metabolismo , Infecciones por Orthomyxoviridae/metabolismo , Proteoma/análisis , Transducción de Señal/efectos de los fármacos , Animales , Línea Celular , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Genoma , Interacciones Huésped-Patógeno/fisiología , Humanos , Virus de la Influenza A/genética , Ratones , Modelos Moleculares , Fosforilación , Conformación Proteica , Proteínas Virales/química , Proteínas Virales/metabolismoRESUMEN
Homeostasis of solid tissue is characterized by a low proliferative activity of differentiated cells while special conditions like tissue damage induce regeneration and proliferation. For some cell types it has been shown that various tissue-specific functions are missing in the proliferating state, raising the possibility that their proliferation is not compatible with a fully differentiated state. While endothelial cells are important players in regenerating tissue as well as in the vascularization of tumors, the impact of proliferation on their features remains elusive. To examine cell features in dependence of proliferation, we established human endothelial cell lines in which proliferation is tightly controlled by a doxycycline-dependent, synthetic regulatory unit. We observed that uptake of macromolecules and establishment of cell-cell contacts was more pronounced in the growth-arrested state. Tube-like structures were formed in vitro in both proliferating and non-proliferating conditions. However, functional vessel formation upon transplantation into immune-compromised mice was restricted to the proliferative state. Kaposi's sarcoma-associated herpes virus (KSHV) infection resulted in reduced expression of endothelial markers. Upon transplantation of infected cells, drastic differences were observed: proliferation arrested cells acquired a high migratory activity while the proliferating counterparts established a tumor-like phenotype, similar to Kaposi Sarcoma lesions. The study gives evidence that proliferation governs endothelial functions. This suggests that several endothelial functions are differentially expressed during angiogenesis. Moreover, since proliferation defines the functional properties of cells upon infection with KSHV, this process crucially affects the fate of virus-infected cells.
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Células Endoteliales/citología , Células Endoteliales/metabolismo , Animales , Antígenos CD34/genética , Antígenos CD34/metabolismo , Antígeno CD146/genética , Antígeno CD146/metabolismo , Línea Celular , Proliferación Celular , Regulación hacia Abajo , Endoglina/genética , Endoglina/metabolismo , Células Endoteliales/trasplante , Perfilación de la Expresión Génica , Herpesvirus Humano 8/genética , Herpesvirus Humano 8/fisiología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Ratones , Ratones Noqueados , Microscopía Fluorescente , Óxido Nítrico/metabolismo , Sarcoma de Kaposi/etiología , Regulación hacia ArribaRESUMEN
De novo heterozygous mutations changing R179 to histidine, leucine, or cysteine in the ACTA2 gene are associated with Multisystemic Smooth Muscle Dysfunction Syndrome (MSMDS). Characteristic hallmarks of this condition, caused only by these specific ACTA2 mutations, are congenital mydriasis (mid-dilated, non-reactive pupils), a large persistent ductus arteriosus (PDA), aortic aneurysms evolving during childhood, and cerebrovascular anomalies. We describe two patients, a 3-day-old newborn and a 26-year-old woman, with this unique mutation in association with a huge PDA and an aorto-pulmonary window. In addition, one showed a coarctation of the aortic arch and the other a complete interruption of the aortic arch type A; thereby expanding the spectrum of cardiac congenital heart defect of this syndrome. Each patient displayed a huge PDA and an extra-cardiovascular phenotype consistent with MSMDS. These observations exemplify that a functional alpha 2 smooth muscle actin is necessary for proper cardiovascular organ development, and demonstrate that a very exceptional congenital heart defect (aortopulmonary window) can be caused by a mutation in a gene encoding a contractile protein of vascular smooth muscle cells. © 2017 Wiley Periodicals, Inc.
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Actinas/genética , Aneurisma de la Aorta/patología , Conducto Arterioso Permeable/patología , Enfermedades Hereditarias del Ojo/patología , Cardiopatías Congénitas/patología , Mutación , Midriasis/patología , Adulto , Aneurisma de la Aorta/diagnóstico por imagen , Aneurisma de la Aorta/genética , Conducto Arterioso Permeable/diagnóstico por imagen , Conducto Arterioso Permeable/genética , Enfermedades Hereditarias del Ojo/diagnóstico por imagen , Enfermedades Hereditarias del Ojo/genética , Femenino , Expresión Génica , Cardiopatías Congénitas/diagnóstico por imagen , Cardiopatías Congénitas/genética , Heterocigoto , Humanos , Recién Nacido , Imagen por Resonancia Magnética , Músculo Liso/metabolismo , Músculo Liso/patología , Midriasis/diagnóstico por imagen , Midriasis/genética , Fenotipo , SíndromeRESUMEN
BACKGROUND: Mutations in PRRT2 cause autosomal dominant paroxysmal kinesigenic dyskinesia with infantile convulsions (PKD/IC). CASE PRESENTATION: A previously not recognized intronic PRRT2 mutation (c.880-35G > A; p.S294Lfs*29) was found in an 18 month old girl with IC and in her mother with classical presentation of PKD. The mutation results in a novel splice acceptor site in intron 2 of PRRT2. Due to frameshift and a subsequent premature stop-codon the resulting transcript appears to render the PRRT2 protein non/dysfunctional and is the likely cause of disease in this family. CONCLUSION: Our findings expand the mutational spectrum of this disease.
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Discinesias/genética , Distonía/genética , Epilepsia Benigna Neonatal/genética , Mutación del Sistema de Lectura , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Sitios de Empalme de ARN , Convulsiones/genética , Secuencia de Bases , Codón sin Sentido , Exones , Femenino , Humanos , Recién Nacido , Intrones , Datos de Secuencia Molecular , Fenotipo , Alineación de Secuencia , Análisis de Secuencia de ADNRESUMEN
Chudley-McCullough syndrome (CMS) is a rare autosomal recessive disorder characterized by sensorineural deafness, agenesis of the corpus callosum, frontal polymicrogyria, interhemispheric cyst, and ventricular enlargement. CMS is caused by mutations in the GPSM2 gene, but until now no more than eight different mutations are on record. We describe two dizygotic twins with a novel homozygous loss-of-function mutation (c.1093C > T; p.Arg365*). While one child developed hydrocephalus-prompting shunt implantation immediately after birth, the other sibling did not. The combination of sensorineural hearing loss and partial agenesis of the corpus callosum is a highly recognizable clinico-radiological entity that should prompt mutational analysis of the GPSM2 gene.
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Agenesia del Cuerpo Calloso/genética , Quistes Aracnoideos/genética , Pérdida Auditiva Sensorineural/genética , Hidrocefalia/cirugía , Péptidos y Proteínas de Señalización Intracelular/genética , Gemelos Dicigóticos/genética , Agenesia del Cuerpo Calloso/complicaciones , Agenesia del Cuerpo Calloso/diagnóstico por imagen , Quistes Aracnoideos/complicaciones , Quistes Aracnoideos/diagnóstico por imagen , Progresión de la Enfermedad , Femenino , Genotipo , Pérdida Auditiva Sensorineural/complicaciones , Pérdida Auditiva Sensorineural/diagnóstico por imagen , Homocigoto , Humanos , Hidrocefalia/etiología , Lactante , Recién Nacido , Imagen por Resonancia Magnética , Masculino , Fenotipo , Derivación VentriculoperitonealRESUMEN
BACKGROUND & AIMS: Activation of the transcription factor nuclear factor-κB (NF-κB) has been associated with the development of inflammatory bowel disease (IBD). Copper metabolism MURR1 domain containing 1 (COMMD1), a regulator of various transport pathways, has been shown to limit NF-κB activation. We investigated the roles of COMMD1 in the pathogenesis of colitis in mice and IBD in human beings. METHODS: We created mice with a specific disruption of Commd1 in myeloid cells (Mye-knockout [K/O] mice); we analyzed immune cell populations and functions and expression of genes regulated by NF-κB. Sepsis was induced in Mye-K/O and wild-type mice by cecal ligation and puncture or intraperitoneal injection of lipopolysaccharide (LPS), colitis was induced by administration of dextran sodium sulfate, and colitis-associated cancer was induced by administration of dextran sodium sulfate and azoxymethane. We measured levels of COMMD1 messenger RNA in colon biopsy specimens from 29 patients with IBD and 16 patients without (controls), and validated findings in an independent cohort (17 patients with IBD and 22 controls). We searched for polymorphisms in or near COMMD1 that were associated with IBD using data from the International IBD Genetics Consortium and performed quantitative trait locus analysis. RESULTS: In comparing gene expression patterns between myeloid cells from Mye-K/O and wild-type mice, we found that COMMD1 represses expression of genes induced by LPS. Mye-K/O mice had more intense inflammatory responses to LPS and developed more severe sepsis and colitis, with greater mortality. More Mye-K/O mice with colitis developed colon dysplasia and tumors than wild-type mice. We observed a reduced expression of COMMD1 in colon biopsy specimens and circulating leukocytes from patients with IBD. We associated single-nucleotide variants near COMMD1 with reduced expression of the gene and linked them with increased risk for ulcerative colitis. CONCLUSIONS: Expression of COMMD1 by myeloid cells has anti-inflammatory effects. Reduced expression or function of COMMD1 could be involved in the pathogenesis of IBD.
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Proteínas Adaptadoras Transductoras de Señales/fisiología , Colitis/prevención & control , Colitis/fisiopatología , Neoplasias del Colon/prevención & control , Neoplasias del Colon/fisiopatología , Inflamación/genética , Inflamación/fisiopatología , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Azoximetano/efectos adversos , Biopsia , Estudios de Casos y Controles , Colitis/inducido químicamente , Colon/metabolismo , Colon/patología , Neoplasias del Colon/inducido químicamente , Sulfato de Dextran/efectos adversos , Modelos Animales de Enfermedad , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Polimorfismo de Nucleótido Simple/genética , ARN Mensajero/metabolismoRESUMEN
Histone deacetylase (HDAC) 3, as a cofactor in co-repressor complexes containing silencing mediator for retinoid or thyroid-hormone receptors (SMRT) and nuclear receptor co-repressor (N-CoR), has been shown to repress gene transcription in a variety of contexts. Here, we reveal a novel role for HDAC3 as a positive regulator of IL-1-induced gene expression. Various experimental approaches involving RNAi-mediated knockdown, conditional gene deletion or small molecule inhibitors indicate a positive role of HDAC3 for transcription of the majority of IL-1-induced human or murine genes. This effect was independent from the gene regulatory effects mediated by the broad-spectrum HDAC inhibitor trichostatin A (TSA) and thus suggests IL-1-specific functions for HDAC3. The stimulatory function of HDAC3 for inflammatory gene expression involves a mechanism that uses binding to NF-κB p65 and its deacetylation at various lysines. NF-κB p65-deficient cells stably reconstituted to express acetylation mimicking forms of p65 (p65 K/Q) had largely lost their potential to stimulate IL-1-triggered gene expression, implying that the co-activating property of HDAC3 involves the removal of inhibitory NF-κB p65 acetylations at K122, 123, 314 and 315. These data describe a novel function for HDAC3 as a co-activator in inflammatory signaling pathways and help to explain the anti-inflammatory effects frequently observed for HDAC inhibitors in (pre)clinical use.
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Histona Desacetilasas/fisiología , Interleucina-1/farmacología , Factor de Transcripción ReIA/metabolismo , Acetilación , Animales , Línea Celular , Quimiocina CXCL2/genética , Quimiocina CXCL2/metabolismo , Regulación hacia Abajo , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Ácidos Hidroxámicos/farmacología , Interleucina-8/genética , Interleucina-8/metabolismo , Ratones , FN-kappa B/metabolismo , Fosforilación , ARN Polimerasa II/metabolismo , Transducción de Señal , Transcripción Genética/efectos de los fármacosRESUMEN
The molecular mechanisms of progressive right heart failure are incompletely understood. In this study, we systematically examined transcriptomic changes occurring over months in isolated cardiomyocytes or whole heart tissues from failing right and left ventricles in rat models of pulmonary artery banding (PAB) or aortic banding (AOB). Detailed bioinformatics analyses resulted in the identification of gene signature, protein and transcription factor networks specific to ventricles and compensated or decompensated disease states. Proteomic and RNA-FISH analyses confirmed PAB-mediated regulation of key genes and revealed spatially heterogeneous mRNA expression in the heart. Intersection of rat PAB-specific gene sets with transcriptome datasets from human patients with chronic thromboembolic pulmonary hypertension (CTEPH) led to the identification of more than 50 genes whose expression levels correlated with the severity of right heart disease, including multiple matrix-regulating and secreted factors. These data define a conserved, differentially regulated genetic network associated with right heart failure in rats and humans.
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Insuficiencia Cardíaca , Ventrículos Cardíacos , Animales , Humanos , Insuficiencia Cardíaca/genética , Insuficiencia Cardíaca/metabolismo , Ventrículos Cardíacos/metabolismo , Ratas , Modelos Animales de Enfermedad , Transcriptoma , Masculino , Perfilación de la Expresión Génica , Miocitos Cardíacos/metabolismo , Redes Reguladoras de Genes , Ratas Sprague-Dawley , Hipertensión Pulmonar/genética , Proteómica , Disfunción Ventricular Derecha/genética , Disfunción Ventricular Derecha/fisiopatologíaRESUMEN
Paroxysmal kinesigenic dyskinesia with infantile convulsions (PKD/IC) is caused by mutations in the gene PRRT2 located in 16p11.2. A deletion syndrome 16p11.2 is well established and is characterized by intellectual disability, speech delay, and autism. PKD/IC, however, is extremely rare in this syndrome. We describe a case of PKD/IC and 16p11.2 deletion syndrome and discuss modifiers of PRRT2 activity to explain the rare concurrence of both syndromes.
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Trastorno Autístico/genética , Corea/genética , Trastornos de los Cromosomas/genética , Epilepsia Benigna Neonatal/genética , Discapacidad Intelectual/genética , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Trastorno Autístico/complicaciones , Niño , Corea/etiología , Deleción Cromosómica , Trastornos de los Cromosomas/complicaciones , Cromosomas Humanos Par 16/genética , Distonía , Epilepsia Benigna Neonatal/etiología , Genes Modificadores , Humanos , Discapacidad Intelectual/complicaciones , Masculino , Polimorfismo de Nucleótido SimpleRESUMEN
The influenza A virus (IAV)-encoded matrix protein 1 (M1) acts as a master regulator of virus replication and fulfills multiple structural and regulatory functions in different cell compartments. Therefore, the spatiotemporal regulation of M1 is achieved by different mechanisms, including its structural and pH-dependent flexibility, differential association with cellular factors, and posttranslational modifications. Here, we investigated the function of M1 phosphorylation at the evolutionarily conserved threonine 108 (T108) and found that its mutation to a nonphosphorylatable alanine prohibited virus replication. Absent T108, phosphorylation led to strongly increased self-association of M1 at the cell membrane and consequently prohibited its ability to enter the nucleus and to contribute to viral ribonucleoprotein nuclear export. M1 T108 phosphorylation also controls the binding affinity to the cellular STRIPAK (striatin-interacting phosphatases and kinases) complex, which contains different kinases and the phosphatase PP2A to shape phosphorylation-dependent signaling networks. IAV infection led to the redistribution of the STRIPAK scaffolding subunits STRN and STRN3 from the cell membrane to cytosolic and perinuclear clusters, where it colocalized with M1. Inactivation of the STRIPAK complex resulted in compromised M1 polymerization and IAV replication. IMPORTANCE Influenza viruses pose a major threat to human health and cause annual epidemics and occasional pandemics. Many virus-encoded proteins exert various functions in different subcellular compartments, as exemplified by the M1 protein, but the molecular mechanisms endowing the multiplicity of functions remain incompletely understood. Here, we report that phosphorylation of M1 at T108 is essential for virus replication and controls its propensity for self-association and nuclear localization. This phosphorylation also controls binding affinity of the M1 protein to the STRIPAK complex, which contributes to M1 polymerization and virus replication.
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Virus de la Influenza A , Gripe Humana , Humanos , Autoantígenos/metabolismo , Proteínas de Unión a Calmodulina/química , Proteínas de Unión a Calmodulina/genética , Proteínas de Unión a Calmodulina/metabolismo , Virus de la Influenza A/genética , Fosforilación , Fosfotransferasas/metabolismo , Transducción de Señal , Replicación ViralRESUMEN
The unbiased identification of cytokine-induced, secreted proteins from cells cultured in serum-containing medium is challenging. Here, we describe an experimental and bioinformatics workflow to label interleukin-1α-regulated proteins in living cells with the methionine analogue L-homopropargylglycine. We detail their purification and identification by means of CLICK-chemistry-based biotinylation followed by nanoHPLC-MS/MS. A side-by-side comparison of enriched proteins and their ontologies to serum-free conditions demonstrates the sensitivity and specificity of this approach to study the inducible secreted proteomes of epithelial cells.
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Infection or cell damage triggers the release of pro-inflammatory cytokines such as interleukin(IL)-1α or ß and tumor necrosis factor (TNF)α which are key mediators of the host immune response. Following their identification and the elucidation of central signaling pathways, recent results show a highly complex crosstalk between various cytokines and their signaling effectors. The molecular mechanisms controlling signaling thresholds, signal integration and the function of feed-forward and feedback loops are currently revealed by combining methods from biochemistry, genetics and in silico analysis. Increasing evidence is mounted that defects in information processing circuits or their components can be causative for chronic or overshooting inflammation. As progress in biosciences has always benefitted from the use of well-studied model systems, research on inflammatory cytokines may function as a paradigm to reveal general principles of signal integration, crosstalk mechanisms and signaling networks.
Asunto(s)
Comunicación Celular , Citocinas/metabolismo , Mediadores de Inflamación/metabolismo , Inflamación/metabolismo , Transducción de Señal , Animales , Humanos , Inflamación/inmunologíaRESUMEN
In response to stimuli that activate p53, cells can undergo either apoptosis or cell cycle arrest, depending on the precise pattern of p53 target genes that is activated. We show here that Zbtb4, a transcriptional repressor protein, associates with the Sin3/histone deacetylase co-repressor and represses expression of P21CIP1 as part of a heterodimeric complex with Miz1. In vivo, expression of ZBTB4 is downregulated in advanced stages of multiple human tumours. In cell culture, depletion of ZBTB4 promotes cell cycle arrest in response to activation of p53 and suppresses apoptosis through regulation of P21CIP1, thereby promoting long-term cell survival. Our data suggest that Zbtb4 is a critical determinant of the cellular response to p53 activation and reinforce the notion that p21Cip1 can provide an essential survival signal in cells with activated p53.