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Sphingosine-1-phosphate lyase is encoded by the Sgpl1 gene, degrades S1P, and is crucial for S1P homeostasis in animal models and humans. S1P lyase deficient patients suffer from adrenal insufficiency, severe lymphopenia, and skin disorders. In this study, we used random mutagenesis screening to identify a mouse line carrying a missense mutation of Sgpl1 (M467K). This mutation caused similar pathologies as Sgpl1 knock-out mice in multiple organs, but greatly preserved its lifespan, which M467K mutation mice look normal under SPF conditions for over 40 weeks, in contrast, the knock-out mice live no more than 6 weeks. When treated with Imiquimod, Sgpl1M467K mice experienced exacerbated skin inflammation, as revealed by aggravated acanthosis and orthokeratotic hyperkeratosis. We also demonstrated that the IL17a producing Vγ6+ cell was enriched in Sgpl1M467K skin and caused severe pathology after imiquimod treatment. Interestingly, hyperchromic plaque occurred in the mutant mice one month after Imiquimod treatment but not in the controls, which resembled the skin disorder found in Sgpl1 deficient patients. Therefore, our results demonstrate that Sgpl1M467K point mutation mice successfully modeled a human disease after being treated with Imiquimod. We also revealed a major subset of γδT cells in the skin, IL17 secreting Vγ6 T cells were augmented by Sgpl1 deficiency and led to skin pathology. Therefore, we have, for the first time, linked the IL17a and γδT cells to SPL insufficiency.
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Hiperpigmentación , Mutación Puntual , Animales , Homeostasis , Imiquimod , Ratones , Ratones NoqueadosRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Mandibuloacral dysplasia syndromes are mainly due to recessive LMNA or ZMPSTE24 mutations, with cardinal nuclear morphological abnormalities and dysfunction. We report five homozygous null mutations in MTX2, encoding Metaxin-2 (MTX2), an outer mitochondrial membrane protein, in patients presenting with a severe laminopathy-like mandibuloacral dysplasia characterized by growth retardation, bone resorption, arterial calcification, renal glomerulosclerosis and severe hypertension. Loss of MTX2 in patients' primary fibroblasts leads to loss of Metaxin-1 (MTX1) and mitochondrial dysfunction, including network fragmentation and oxidative phosphorylation impairment. Furthermore, patients' fibroblasts are resistant to induced apoptosis, leading to increased cell senescence and mitophagy and reduced proliferation. Interestingly, secondary nuclear morphological defects are observed in both MTX2-mutant fibroblasts and mtx-2-depleted C. elegans. We thus report the identification of a severe premature aging syndrome revealing an unsuspected link between mitochondrial composition and function and nuclear morphology, establishing a pathophysiological link with premature aging laminopathies and likely explaining common clinical features.
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Acroosteólisis/metabolismo , Predisposición Genética a la Enfermedad/genética , Lipodistrofia/metabolismo , Mandíbula/anomalías , Proteínas de la Membrana/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Acroosteólisis/diagnóstico por imagen , Acroosteólisis/genética , Acroosteólisis/patología , Envejecimiento Prematuro/genética , Envejecimiento Prematuro/metabolismo , Animales , Apoptosis , Caenorhabditis elegans , Proliferación Celular , Niño , Regulación hacia Abajo , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Regulación de la Expresión Génica , Genotipo , Homocigoto , Humanos , Lipodistrofia/diagnóstico por imagen , Lipodistrofia/genética , Lipodistrofia/patología , Masculino , Mandíbula/diagnóstico por imagen , Proteínas de la Membrana/genética , Metaloendopeptidasas , Proteínas de Transporte de Membrana Mitocondrial/genética , Proteínas Mitocondriales/genética , Mutación , Fenotipo , Piel , Secuenciación Completa del GenomaRESUMEN
Recent reports highlight regulatory functions of long noncoding RNAs (lncRNAs) in neurodegeneration and aging, but biomedical implications remain limited. Here, we report an rRNA-depletion-based long RNA-Sequencing Resource of 65 substantia nigra, amygdala, and medial temporal gyrus samples from Parkinson's disease (PD) and matched control brains. Using a lncRNA-focused analysis approach to identify functionally important transcripts, we discovered and prioritized many lncRNAs dysregulated in PD. Those included pronounced elevation of the P53-induced noncoding transcript LINC-PINT in the substantia nigra of PD patients, as well as in additional models of oxidative stress and PD. Intriguingly, we found that LINC-PINT is a primarily neuronal transcript which showed conspicuous increases in maturing primary culture neurons. LINC-PINT also accumulated in several brain regions of Alzheimer's and Huntington's disease patients and decreased with healthy brain aging, suggesting a general role in aging and neurodegeneration for this lncRNA. RNAi-mediated depletion of LINC-PINT exacerbated the death of cultured N2A and SH-SY5Y cells exposed to oxidative stress, highlighting a previously undiscovered neuroprotective role for this tumor-inducible lncRNA in the brains of patients with neurodegenerative disorders.
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Neuroprotección/genética , Enfermedad de Parkinson/metabolismo , ARN Largo no Codificante/metabolismo , Sustancia Negra/metabolismo , Anciano , Anciano de 80 o más Años , Animales , Muerte Celular/efectos de los fármacos , Muerte Celular/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Estudios de Cohortes , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Ratones , Persona de Mediana Edad , Neuroblastoma/patología , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Peróxidos/farmacología , Interferencia de ARN , ARN Largo no Codificante/genética , RNA-SeqRESUMEN
When an animal is infected, its innate immune response needs to be tightly regulated across tissues and coordinated with other aspects of organismal physiology. Previous studies with Caenorhabditis elegans have demonstrated that insulin-like peptide genes are differentially expressed in response to different pathogens. They represent prime candidates for conveying signals between tissues upon infection. Here, we focused on one such gene, ins-11 and its potential role in mediating cross-tissue regulation of innate immune genes. While diverse bacterial intestinal infections can trigger the up-regulation of ins-11 in the intestine, we show that epidermal infection with the fungus Drechmeria coniospora triggers an upregulation of ins-11 in the epidermis. Using the Shigella virulence factor OpsF, a MAP kinase inhibitor, we found that in both cases, ins-11 expression is controlled cell autonomously by p38 MAPK, but via distinct transcription factors, STA-2/STAT in the epidermis and HLH-30/TFEB in the intestine. We established that ins-11, and the insulin signaling pathway more generally, are not involved in the regulation of antimicrobial peptide gene expression in the epidermis. The up-regulation of ins-11 in the epidermis does, however, affect intestinal gene expression in a complex manner, and has a deleterious effect on longevity. These results support a model in which insulin signaling, via ins-11, contributes to the coordination of the organismal response to infection, influencing the allocation of resources in an infected animal.
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Proteínas de Caenorhabditis elegans/biosíntesis , Caenorhabditis elegans/microbiología , Regulación de la Expresión Génica , Hypocreales/crecimiento & desarrollo , Hormonas Peptídicas/biosíntesis , Animales , Proteínas Bacterianas/metabolismo , Epidermis/microbiología , Intestinos/microbiología , Factores de Transcripción/metabolismo , Factores de Virulencia/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Toxins are believed to play a crucial role in Burkholderia pseudomallei pathogenicity, however to date, only a few have been identified. The discovery of additional toxic molecules is limited by the lack of a sensitive indicator of B. pseudomallei toxicity. Previously, from a whole genome transcriptome analysis of B. pseudomallei-infected Caenorhabditis elegans, we noted significant overexpression of a number of worm genes encoding detoxification enzymes, indicating the host's attempt to clear bacterial toxic molecules. One of these genes, ugt-29, a family member of UDP-glucuronosyltransferases, was the most robustly induced phase II detoxification gene. In this study, we show that strong induction of ugt-29 is restricted to infections by the most virulent species among the pathogens tested. We also noted that ugt-29 is activated upon disruption of host protein synthesis. Hence, we propose that UGT-29 could be a promising biosensor to detect B. pseudomallei toxins that compromise host protein synthesis. The identification of bactobolin, a polyketide-peptide hybrid molecule, as a toxic molecule of B. pseudomallei further verifies the utilization of this surveillance system to search for bacterial toxins. Hence, a ugt-29 based reporter should be useful in screening for other molecules that inhibit host protein synthesis.
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Toxinas Bacterianas/toxicidad , Técnicas Biosensibles , Burkholderia pseudomallei/patogenicidad , Caenorhabditis elegans/metabolismo , Biosíntesis de Proteínas/efectos de los fármacos , Inhibidores de la Síntesis de la Proteína/toxicidad , AnimalesRESUMEN
Burkholderia pseudomallei is a Gram-negative soil bacterium that infects both humans and animals. Although cell culture studies have revealed significant insights into factors contributing to virulence and host defense, the interactions between this pathogen and its intact host remain to be elucidated. To gain insights into the host defense responses to B. pseudomallei infection within an intact host, we analyzed the genome-wide transcriptome of infected Caenorhabditis elegans and identified â¼6% of the nematode genes that were significantly altered over a 12-h course of infection. An unexpected feature of the transcriptional response to B. pseudomallei was a progressive increase in the proportion of down-regulated genes, of which ELT-2 transcriptional targets were significantly enriched. ELT-2 is an intestinal GATA transcription factor with a conserved role in immune responses. We demonstrate that B. pseudomallei down-regulation of ELT-2 targets is associated with degradation of ELT-2 protein by the host ubiquitin-proteasome system. Degradation of ELT-2 requires the B. pseudomallei type III secretion system. Together, our studies using an intact host provide evidence for pathogen-mediated host immune suppression through the destruction of a host transcription factor.
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Burkholderia pseudomallei/patogenicidad , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/inmunología , Caenorhabditis elegans/microbiología , Factores de Transcripción GATA/metabolismo , Animales , Animales Modificados Genéticamente , Burkholderia pseudomallei/genética , Burkholderia pseudomallei/inmunología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Regulación hacia Abajo , Factores de Transcripción GATA/genética , Interacciones Huésped-Patógeno/inmunología , Procesamiento Postranscripcional del ARN , ARN de Helminto/genética , ARN de Helminto/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Virulencia/inmunologíaRESUMEN
The nematode Caenorhabditis elegans is hypersusceptible to Burkholderia pseudomallei infection. However, the virulence mechanisms underlying rapid lethality of C. elegans upon B. pseudomallei infection remain poorly defined. To probe the host-pathogen interaction, we constructed GFP-tagged B. pseudomallei and followed bacterial accumulation within the C. elegans intestinal lumen. Contrary to slow-killing by most bacterial pathogens, B. pseudomallei caused fairly limited intestinal lumen colonization throughout the period of observation. Using grinder-defective mutant worms that allow the entry of intact bacteria also did not result in full intestinal lumen colonization. In addition, we observed a significant decline in C. elegans defecation and pharyngeal pumping rates upon B. pseudomallei infection. The decline in defecation rates ruled out the contribution of defecation to the limited B. pseudomallei colonization. We also demonstrated that the limited intestinal lumen colonization was not attributed to slowed host feeding as bacterial loads did not change significantly when feeding was stimulated by exogenous serotonin. Both these observations confirm that B. pseudomallei is a poor colonizer of the C. elegans intestine. To explore the possibility of toxin-mediated killing, we examined the transcription of the C. elegans ABC transporter gene, pgp-5, upon B. pseudomallei infection of the ppgp-5::gfp reporter strain. Expression of pgp-5 was highly induced, notably in the pharynx and intestine, compared with Escherichia coli-fed worms, suggesting that the host actively thwarted the pathogenic assaults during infection. Collectively, our findings propose that B. pseudomallei specifically and continuously secretes toxins to overcome C. elegans immune responses.
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Transportadoras de Casetes de Unión a ATP/genética , Toxinas Bacterianas/metabolismo , Infecciones por Burkholderia/inmunología , Burkholderia pseudomallei/patogenicidad , Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/inmunología , Transportadoras de Casetes de Unión a ATP/biosíntesis , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Toxinas Bacterianas/inmunología , Infecciones por Burkholderia/patología , Caenorhabditis elegans/microbiología , Proteínas de Caenorhabditis elegans/biosíntesis , Proteínas de Caenorhabditis elegans/metabolismo , Escherichia coli/patogenicidad , Proteínas Fluorescentes Verdes/genética , Interacciones Huésped-Patógeno , Intestinos/microbiología , Proteínas Protozoarias/biosíntesis , Proteínas Protozoarias/genética , Interferencia de ARN , ARN Interferente Pequeño , VirulenciaRESUMEN
BACKGROUND: Burkholderia pseudomallei is the causative agent of melioidosis, a disease of significant morbidity and mortality in both human and animals in endemic areas. Much remains to be known about the contributions of genotypic variations within the bacteria and the host, and environmental factors that lead to the manifestation of the clinical symptoms of melioidosis. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we showed that different isolates of B. pseudomallei have divergent ability to kill the soil nematode Caenorhabditis elegans. The rate of nematode killing was also dependent on growth media: B. pseudomallei grown on peptone-glucose media killed C. elegans more rapidly than bacteria grown on the nematode growth media. Filter and bacteria cell-free culture filtrate assays demonstrated that the extent of killing observed is significantly less than that observed in the direct killing assay. Additionally, we showed that B. pseudomallei does not persistently accumulate within the C. elegans gut as brief exposure to B. pseudomallei is not sufficient for C. elegans infection. CONCLUSIONS/SIGNIFICANCE: A combination of genetic and environmental factors affects virulence. In addition, we have also demonstrated that a Burkholderia-specific mechanism mediating the pathogenic effect in C. elegans requires proliferating B. pseudomallei to continuously produce toxins to mediate complete killing.
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Burkholderia pseudomallei/patogenicidad , Caenorhabditis elegans/microbiología , Interacciones Huésped-Patógeno , Animales , Burkholderia pseudomallei/aislamiento & purificación , Difusión , Susceptibilidad a Enfermedades , Ambiente , Tracto Gastrointestinal/microbiología , Tracto Gastrointestinal/patología , Humanos , Ratones , Toxinas Biológicas/metabolismoRESUMEN
Burkholderia pseudomallei is a Gram-negative saprophytic soil bacterium, which is the etiologic agent of melioidosis, a severe and fatal infectious disease occurring in human and animals. Distinct clinical and animal isolates have been shown to exhibit differences in phenotypic trait such as growth rate, colony morphology, antimicrobial resistance, and virulence. This study was carried out to gain insight into the intrinsic differences between 4 clinical and 6 animal B. pseudomallei isolates from Malaysia. The 16S rRNA-encoding genes from these 10 isolates of B. pseudomallei were sequenced to confirm the identity of these isolates along with the avirulent Burkholderia thailandensis. The nucleotide sequences indicated that the 16S rRNA-encoding genes among the 10 B. pseudomallei isolates were identical to each other. However, the nucleotide sequence differences in the 16S rRNA-encoding genes appeared to be B. pseudomallei and B. thailandensis specific. The growth rate of all B. pseudomallei isolates was determined by generating growth curves at 37 degrees C for 72 h. The isolates were found to differ in growth rates with doubling time varying from 1.5 to 2.3 h. In addition, the B. pseudomallei isolates exhibited considerable variation in colony morphology when grown on Ashdown media, brain-heart infusion agar, and Luria-Bertani agar over 9 days of observation. Antimicrobial susceptibility tests indicated that 80% of the isolates examined were Amp(R) Cb(R) Kn(R) Gm(R) Chl(S) Te(S). Virulence of the B. pseudomallei clinical and animal isolates was evaluated in B. pseudomallei-susceptible BALB/c mice. Most of the clinical isolates were highly virulent. However, virulence did not correlate with isolate origin since 2 of the animal isolates were also highly virulent.