RESUMO
miRNA biogenesis process is an intricate and complex event consisting of many proteins working in a highly coordinated fashion. Most of these proteins have been studied in Arabidopsis; however, their orthologs and functions have not been explored in other plant species. In the present study, we have manually curated all the experimentally verified information present in the literature regarding these proteins and found a total of 98 genes involved in miRNA biogenesis in Arabidopsis. The conservation pattern of these proteins was identified in other plant species ranging from dicots to lower organisms, and we found that a major proportion of proteins involved in the pri-miRNA processing are conserved. However, nearly 20% of the genes, mostly involved in either transcription or functioning of the miRNAs, were absent in the lower organisms. Further, we manually curated a regulatory network of the core components of the biogenesis process and found that nearly half (46%) of the proteins interact with them, indicating that the processing step is perhaps the most under surveillance/regulation. We have subsequently attempted to characterize the orthologs identified in Oryza sativa, on the basis of transcriptome and epigenetic modifications under field drought conditions in order to assess the impact of drought on the process. We found several participating genes to be differentially expressed and/or epigenetically methylated under drought, although the core components like DCL1, SE, and HYL1 remain unaffected by the stress itself. The study enhances our present understanding of the biogenesis process and its regulation.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , MicroRNAs , Oryza , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Oryza/genética , Oryza/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Processamento Pós-Transcricional do RNA , Regulação da Expressão Gênica de Plantas , Proteínas de Ligação a RNA/metabolismoRESUMO
Engineering drought tolerance in rice needs to focus on regulators that enhance tolerance while boosting plant growth and vigor. The present study delineated the concealed function and tissue-mediated interplay of the miR408/target module in imparting drought stress tolerance in rice. The plant miR408 family comprises three dominant mature forms (21 nt), including a distinct monocot variant (F-7 with 5' C) and is divided into six groups. miR408 majorly cleaves genes belonging to the blue copper protein in addition to several other species-specific targets in plants. Comparative sequence analysis in 4726 rice accessions identified 22 sequence variants (SNP and InDELs) in its promoter (15) and pre-miR408 region. Haplotype analysis of the sequence variants indicated eight haplotypes (three: Japonica-specific and five: Indica-specific) of the miR408 promoter. In drought-tolerant Nagina 22, miR408 follows flag leaf preferential expression. Under drought conditions, its levels are upregulated in flag leaf and roots which seems to be regulated by a differential fraction of methylated cytosines (mCs) in the precursor region. The active pool of miR408 regulated targets under control and drought conditions is impacted by the tissue type. Comparative expression analysis of the miR408/target module under different sets of conditions features 83 targets exhibiting antagonistic expression in rice, out of which 12 genes, including four PLANTACYANINS (OsUCL6, 7, 9 and 30), PIRIN, OsLPR1, OsCHUP1, OsDOF12, OsBGLU1, glycine-rich cell wall gene, OsDUT, and OsERF7, are among the high confidence targets. Further, overexpression of MIR408 in drought-sensitive rice cultivar (PB1) leads to the massive enhancement of vegetative growth in rice with improved ETR and Y(II) and enhanced dehydration stress tolerance. The above results suggest that miR408 is likely to act as a positive regulator of growth and vigor, as well as dehydration stress, making it a potential candidate for engineering drought tolerance in rice.
Assuntos
Oryza , Oryza/metabolismo , Secas , Desidratação/genética , Regiões Promotoras Genéticas , Folhas de Planta/metabolismo , Regulação da Expressão Gênica de Plantas , Estresse Fisiológico/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
The divergence of natural stress tolerance mechanisms between species is an intriguing phenomenon. To study it in rice, a comparative transcriptome analysis was carried out in 'heading' stage tissue (flag leaf, panicles and roots) of Nagina 22 (N22; drought-tolerant) and IR64 (drought-sensitive) plants subjected to field drought. Interestingly, N22 showed almost double the number of differentially expressed genes (DEGs) than IR64. Many DEGs colocalized within drought-related QTLs responsible for grain yield and drought tolerance and also associated with drought tolerance and critical drought-related plant traits such as leaf rolling, trehalose content, sucrose and cellulose content. Besides, co-expression analysis of the DEGs revealed several 'hub' genes known to actively regulate drought stress response. Strikingly, 1366 DEGs, including 21 'hub' genes, showed a distinct opposite regulation in the two rice varieties under similar drought conditions. Annotation of these variety-specific DEGs (VS-DEGs) revealed that they are distributed in various biological pathways. Furthermore, 103 VS-DEGs were found to physically interact with over 1300 genes, including 32 that physically interact with other VS-DEGs as well. The promoter region of these genes has sequence variations among the two rice varieties, which might be in part responsible for their unique expression pattern.
Assuntos
Secas , Oryza , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Oryza/metabolismo , Estresse Fisiológico/genética , TranscriptomaRESUMO
OBJECTIVES: Crohn disease (CD) is a chronic relapsing condition possibly caused by a dysbiotic microbiome. Approximately 30% to 60% of patients with CD have anti-Saccharomyces cerevisiae antibody (ASCA), but any association with gut microbiota is unexplored. We hypothesized that ASCA positivity would predict a signature microbial status and clinical phenotype. METHODS: Ileocolonic mucosal biopsies were obtained from children with CD (nâ=â135), and controls without inflammatory bowel disease (nâ=â45). Comparison was made between ASCA status, microbial diversity, and clinical characteristics. RESULTS: ASCA was highly specific but poorly sensitive for the diagnosis of CD. In patients with CD, ASCA positivity was associated with older age (≥10 years), ileocolonic disease, and long-term risk of surgery. Microbial alpha and beta diversity were similar in patients with CD with or without ASCA, but significantly less when compared to noninflammatory bowel disease controls. Microbial richness was similar across all 3 groups. Fourteen bacterial species were associated with ASCA-positive patients with CD and 14 species with ASCA-negative patients (Pâ<â0.05). After using a false discovery rate correction Ruminococcus torques and bacterium Yersinia enterocolitica 61 remained significantly associated with CD ASCA positivity (Pâ=â0.0178), whereas Enterobacter cloacae and Faecalibacterium prausnitzii were significantly associated with CD ASCA negativity (Pâ=â0.0178 and 0.0342). CONCLUSION: ASCA-positive and ASCA-negative patients with CD have significant differences in gut microbiome composition, which could possibly be influencing the phenotype of the disease.
Assuntos
Anticorpos Antifúngicos/imunologia , Doença de Crohn/microbiologia , Microbioma Gastrointestinal/imunologia , Proteínas de Saccharomyces cerevisiae/imunologia , Adolescente , Estudos de Casos e Controles , Criança , Pré-Escolar , Feminino , Humanos , Lactente , MasculinoRESUMO
MAIN CONCLUSION: Drought-tolerant rice variety, Nagina 22 (N22), has a unique spikelet miRNome during anthesis stage drought as well as transition from heading to anthesis. Molecular characterization of genetic diversity of rice is essential to understand the evolution and molecular basis of various agronomically important traits such as drought tolerance. miRNAs play an important role in regulating plant development as well as stress response such as drought. In this study, we characterized the yet unexplored dynamics of the spikelet miRNA population during developmental transition from 'heading' to 'anthesis' as well as anthesis stage drought stress in a drought-tolerant indica rice variety, N22. A significant proportion of miRNA population (~20 %) in N22 spikelets is modulated during transition from heading to anthesis indicating a unique miRNome at anthesis, a developmental stage highly sensitive to stress (drought/heat). Based on the analysis of degradome data, majority of differentially regulated miRNAs appear to regulate transcription factors, some of which are implicated in regulation of development and fertilization. Similarly, drought during anthesis leads to a global change in miRNA expression pattern including those which regulate ROS homeostasis. It was possible to identify several miRNAs that were not reported to be drought responsive in earlier studies. Interestingly, a significant proportion of the drought-regulated miRNAs co-localize within QTLs related to drought tolerance and associated traits. Comparison of the expression profiles between N22 and Pusa Basmati 1 (drought sensitive) identified miRNAs with variety-specific expression patterns during phase transition (miR164, miR396, miR812, and miR1881) as well as drought stress (miR1881) indicating an evolution of a distinct and variety-specific regulatory mechanism. The promoters of these miRNAs contain LREs (light-responsive elements) and are induced by dark treatment. It was also possible to identify 4 novel miRNAs including an intronic miRNA that was conserved in both rice varieties.
Assuntos
Adaptação Fisiológica/genética , Secas , Flores/fisiologia , Regulação da Expressão Gênica de Plantas , MicroRNAs/genética , Oryza/genética , Oryza/fisiologia , Adaptação Fisiológica/efeitos da radiação , Sequência de Bases , Flores/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/efeitos da radiação , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Loci Gênicos , Genoma de Planta , Luz , Dados de Sequência Molecular , Oryza/efeitos da radiação , Estresse Fisiológico/genética , Estresse Fisiológico/efeitos da radiaçãoRESUMO
MicroRNAs (miRNAs) are critical components of the multidimensional regulatory networks in eukaryotic systems. Given their diverse spectrum of function, it is apparent that the transcription, processing, and activity of the miRNAs themselves, is very dynamically regulated. One of the most important and universally implicated signaling molecules is [Ca2+]cyt. It is known to regulate a plethora of developmental and metabolic processes in both plants and animals; however, its impact on the regulation of miRNA expression is relatively less explored. The current study employed a combination of internal and external calcium channel inhibitors to establishing that [Ca2+]cyt signatures actively regulate miRNA expression in rice. Involvement of [Ca2+]cyt in the regulation of miRNA expression was further confirmed by treatment with calcimycin, the calcium ionophore. Modulation of the cytosolic calcium levels was also found to regulate the drought-responsive expression as well as ABA-mediated response of miRNA genes in rice seedlings. The study further establishes the role of calmodulins and Calmodulin-binding Transcription Activators (CAMTAs) as important components of the signal transduction schema that regulates miRNA expression. Yeast one-hybrid assay established that OsCAMTA4 & 6 are involved in the transcriptional regulation of miR156a and miR167h. Thus, the study was able to establish that [Ca2+]cyt is actively involved in regulating the expression of miRNA genes both under control and stress conditions.
RESUMO
BACKGROUND AND AIMS: The gut mucosa is the principal site where Crohn's disease [CD] inflammation occurs. Limited information is available about the gut mucosal microbiome during CD relapse and remission. The aim of our study was to characterize specific changes in the gut microbiome during relapse and remission in a large single-centre paediatric CD cohort. METHODS: We analysed the microbiome of 345 biopsies from 204 patients, including 88 CD first diagnosis [CDFD] patients, 38 relapse [CDRL] patients, 12 remission [CDRM] patients, and 66 controls. Species identification was conducted using oligotyping in combination with ARB/SILVA taxonomic annotation. RESULTS: We observed 45 bacteria to differ between CDFD samples and controls with statistical significance, with Fusobacterium being the most implicated species in CDFD patients. We also identified gender-specific differences in CD. Five species showed a strong association with CDRL patients and 10 species with CDRM patients. Three taxa showed a positive co-occurrence across the two groups. Hespellia porcina [closest taxonomic neighbour to Clostridium oroticum] was the most strongly associated with CDRL samples. Interestingly, Fusobacterium was not part of the CDRL-associated taxa group. Faecalibacterium prausnitzii was equally present in CDFD and control samples. CONCLUSION: This is the first study that has investigated the gut mucosal microbiome in a paediatric CD cohort with longitudinal sampling. Importantly, the microbiome of patients in CDRM did not return to a healthy control state. Neither did the microbiome of patients with CDRL return to the profile seen at CDFD.
Assuntos
Clostridiales/isolamento & purificação , Doença de Crohn , Fusobacterium/isolamento & purificação , Microbioma Gastrointestinal , Mucosa Intestinal , Adolescente , Austrália , Biópsia/métodos , Biópsia/estatística & dados numéricos , Criança , Estudos de Coortes , Doença de Crohn/diagnóstico , Doença de Crohn/microbiologia , Doença de Crohn/patologia , Feminino , Humanos , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Estudos Longitudinais , Masculino , Gravidade do Paciente , Estudos ProspectivosAssuntos
Autoanticorpos/efeitos dos fármacos , Hepacivirus/metabolismo , Esteroides/uso terapêutico , Linfócitos T/metabolismo , Alanina Transaminase/sangue , Alanina Transaminase/metabolismo , Feminino , Hepacivirus/efeitos dos fármacos , Hepatite C/virologia , Humanos , Lactente , Esteroides/farmacologia , Linfócitos T/efeitos dos fármacosRESUMO
Human milk samples were collected from 86 mothers induced in the study on days 3, 7, 14 and 28 of lactation. The mothers were divided into three groups on the basis of gestation viz. group 1: 37-41 weeks (n=41), group II: 33-36 (n=23) and group III: <33 weeks gestation (n=22). All the samples were analysed for the estimation of fat, lactose and protein. The results obtained from the investigations were statistically analysed. The analysis of the results revealed a lower amount of fat and lactose in preterm milk as compared to term milk (p<0.01). These were found to increase in amount with increasing postnatal age (p<0.05). The protein levels were observed to be significantly higher in preterm than term milk (p<0.01). These were observed to decrease significantly with increase in postnatal age (p<0.01).
RESUMO
MicroRNAs regulate a spectrum of developmental and biochemical processes in plants and animals. Thus, knowledge of the entire miRNome is essential to understand the complete regulatory schema of any organism. The current study attempts to unravel yet undiscovered miRNA genes in rice. Analysis of small RNA libraries from various tissues of drought-tolerant 'aus' rice variety Nagina 22 (N22) identified 71 novel miRNAs. These were validated based on precursor hairpin structure, small RNA mapping pattern, 'star' sequence, conservation and identification of targets based on degradome data. While some novel miRNAs were conserved in other monocots and dicots, most appear to be lineage-specific. They were segregated into two different classes based on the closeness to the classical miRNA definition. Interestingly, evidence of a miRNA-like cleavage was found even for miRNAs that lie beyond the classical definition. Several novel miRNAs displayed tissue-enriched and/or drought responsive expression. Generation and analysis of the degradome data from N22 along with publicly available degradome identified several high confidence targets implicated in regulation of fundamental processes such as flowering and stress response. Thus, discovery of these novel miRNAs considerably expands the dimension of the miRNA-mediated regulation in rice.
Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , MicroRNAs/genética , Oryza/fisiologia , Estresse Fisiológico , Sequência de Bases , Sequência Conservada , Secas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Biblioteca Gênica , MicroRNAs/química , Oryza/genética , RNA de Plantas/química , RNA de Plantas/genéticaRESUMO
Evolution of differential regulatory mechanisms can lead to quite distinct physiological attributes. In the present study, we have identified one such regulatory schema that regulates osa-miR408 and responds differentially in drought-sensitive and -tolerant indica rice varieties. A comparison of the drought stress response in drought-sensitive (Pusa Basmati 1 and IR64) and drought-tolerant (Nagina 22 and Vandana) indica rice varieties revealed that, during drought stress, levels of miR408 transcript decrease significantly in sensitive cultivars, whereas they remain elevated in the tolerant cultivars. The trend is reflected in young seedlings, as well as in flag leaf and spikelets of adult plants (heading stage). Members of the plastocyanin-like protein family targeted by miR408 also show the inverse expression profile and thus accumulate at a lower level in tolerant cultivars during drought. Interestingly, some members of this family are implicated in maintaining the cellular redox state and spikelet fertility in Arabidopsis. An investigation of miR408 loci (including promoter) in all four cultivars did not reveal any significant sequence variation indicating an involvement of the upstream regulatory schema. Indeed, a similar variety-specific stress response was found in the Oryza sativa squamosa promoter-binding-like 9 transcription factor that regulates miR408 expression. We further demonstrate that drought-mediated induction of miR408 in Nagina 22 is regulated by [Ca(2+)]cyt levels. However, [Ca(2+)]cyt does not appear to regulate miR408 levels in Pusa Basmati 1, suggesting a variety-specific evolution of regulatory schema in rice.
Assuntos
Secas , Regulação da Expressão Gênica de Plantas , MicroRNAs , Oryza/fisiologia , Ácido Abscísico/metabolismo , Sequência de Bases , Cálcio/metabolismo , Sequência Conservada , Citosol/metabolismo , Evolução Molecular , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plastocianina/metabolismo , Regiões Promotoras Genéticas , Especificidade da Espécie , Estresse Fisiológico/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Pruritus with abnormal liver function tests is the classical presentation of intrahepatic cholestasis of pregnancy (ICP), a condition associated with significant fetal complications. Although the etiology of ICP is unclear in many cases, certain features of the clinical presentation should alert the practitioner to the possibility of an underlying metabolic defect, which may not only affect subsequent pregnancies, but may be an indicator of more serious subsequent liver disease. We report a kindred of Anglo-Celtic descent, among whom many members present with ICP, gallstones or cholestasis related to use of oral contraception. Genetic studies revealed a novel mutation in the ABCB4 gene, which codes for a phospholipid transport protein. The clinical significance of this mutation and the importance of identifying such patients are discussed.