RESUMO
Hepatitis B virus (HBV) is an enveloped DNA human virus belonging to the Hepadnaviridae family. Perhaps its main distinguishable characteristic is the replication of its genome through a reverse transcription process. The HBV circular genome encodes only four overlapping reading frames, encoding for the main canonical proteins named core, P, surface, and X (or HBx protein). However, pre- and post-transcriptional gene regulation diversifies the full HBV proteome into diverse isoform proteins. In line with this, hepatitis B virus X protein (HBx) is a viral multifunctional and regulatory protein of 16.5 kDa, whose canonical reading frame presents two phylogenetically conserved internal in-frame translational initiation codons, and which results as well in the expression of two divergent N-terminal smaller isoforms of 8.6 and 5.8 kDa, during translation. The canonical HBx, as well as the smaller isoform proteins, displays different roles during viral replication and subcellular localizations. In this article, we reviewed the different mechanisms of pre- and post-transcriptional regulation of protein expression that take place during viral replication. We also investigated all the past and recent evidence about HBV HBx gene regulation and its divergent N-terminal isoform proteins. Evidence has been collected for over 30 years. The accumulated evidence simply strengthens the concept of a new paradigm of the canonical HBx, and its smaller divergent N-terminal isoform proteins, not only during viral replication, but also throughout cell pathogenesis.
RESUMO
The scale of post-transcriptional regulation and the implications of its interplay with other forms of regulation in environmental acclimation are underexplored for organisms of the domain Archaea. Here, we have investigated the scale of post-transcriptional regulation in the extremely halophilic archaeon Halobacterium salinarum NRC-1 by integrating the transcriptome-wide locations of transcript processing sites (TPSs) and SmAP1 binding, the genome-wide locations of antisense RNAs (asRNAs), and the consequences of RNase_2099C knockout on the differential expression of all genes. This integrated analysis has discovered that 54% of all protein-coding genes in the genome of this haloarchaeon are likely targeted by multiple mechanisms for putative post-transcriptional processing and regulation, with about 20% of genes likely being regulated by combinatorial schemes involving SmAP1, asRNAs, and RNase_2099C. Comparative analysis of mRNA levels (transcriptome sequencing [RNA-Seq]) and protein levels (sequential window acquisition of all theoretical fragment ion spectra mass spectrometry [SWATH-MS]) for 2,579 genes over four phases of batch culture growth in complex medium generated additional evidence for the conditional post-transcriptional regulation of 7% of all protein-coding genes. We demonstrate that post-transcriptional regulation may act to fine-tune specialized and rapid acclimation to stressful environments, e.g., as a switch to turn on gas vesicle biogenesis to promote vertical relocation under anoxic conditions and modulate the frequency of transposition by insertion sequence (IS) elements of the IS200/IS605, IS4, and ISH3 families. Findings from this study are provided as an atlas in a public Web resource (https://halodata.systemsbiology.net). IMPORTANCE While the transcriptional regulation landscape of archaea has been extensively investigated, we currently have limited knowledge about post-transcriptional regulation and its driving mechanisms in this domain of life. In this study, we collected and integrated omics data from multiple sources and technologies to infer post-transcriptionally regulated genes and the putative mechanisms modulating their expression at the protein level in Halobacterium salinarum NRC-1. The results suggest that post-transcriptional regulation may drive environmental acclimation by regulating hallmark biological processes. To foster discoveries by other research groups interested in the topic, we extended our integrated data to the public in the form of an interactive atlas (https://halodata.systemsbiology.net).
Assuntos
Archaea , Transcriptoma , Humanos , Archaea/genética , Transcriptoma/genética , Genoma , RNA Antissenso/genética , Ribonucleases/genéticaRESUMO
Trypanosoma cruzi has three biochemically and morphologically distinct developmental stages that are programmed to rapidly respond to environmental changes the parasite faces during its life cycle. Unlike other eukaryotes, Trypanosomatid genomes contain protein coding genes that are transcribed into polycistronic pre-mRNAs and have their expression controlled by post-transcriptional mechanisms. Transcriptome analyses comparing three stages of the T. cruzi life cycle revealed changes in gene expression that reflect the parasite adaptation to distinct environments. Several genes encoding RNA binding proteins (RBPs), known to act as key post-transcriptional regulatory factors, were also differentially expressed. We characterized one T. cruzi RBP, named TcZH3H12, which contains a zinc finger domain and is up-regulated in epimastigotes compared to trypomastigotes and amastigotes. TcZC3H12 knockout (KO) epimastigotes showed decreased growth rates and increased capacity to differentiate into metacyclic trypomastigotes. Transcriptome analyses comparing wild type and TcZC3H12 KOs revealed a TcZC3H12-dependent expression of epimastigote-specific genes such as genes encoding amino acid transporters and proteins associated with differentiation (PADs). RNA immunoprecipitation assays showed that transcripts from the PAD family interact with TcZC3H12. Taken together, these findings suggest that TcZC3H12 positively regulates the expression of genes involved in epimastigote proliferation and also acts as a negative regulator of metacyclogenesis.
Assuntos
Expressão Gênica , Proteínas de Protozoários/genética , Trypanosoma cruzi/genética , Dedos de Zinco/genética , Sequência de Aminoácidos , Filogenia , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Alinhamento de Sequência , Trypanosoma cruzi/metabolismoRESUMO
Angiogenesis is a pivotal process of homeostasis and tissue repair, but it also favours neovascularisation syndromes and cancer nutrition. The chemical mediation of angiogenesis is complex, involving a balance between serine proteases and their inhibitors. We addressed the mechanisms of action of a Kunitz serine protease inhibitor (KPI) on spontaneous angiogenesis, using Amblyomin-X, a KPI designed from the cDNA library of the Amblyomma cajennense tick. Amblyomin-X treatment (10-1000 ng/10 µL; each 48 h; 3 times) reduced the number of vessels in the subcutaneous dorsal tissue of male Swiss mice, as measured by intravital microscopy, haematoxylin-eosin staining, and PECAM-1 immunofluorescence labeling. Incubation of Amblyomin-X with t-End endothelial cells, a murine endothelial microvascular lineage, did not alter cell proliferation, cell-cycle phases, necrosis and apoptosis, and the production of nitric oxide and prostaglandin E2. Nevertheless, Amblyomin-X treatment reduced t-End migration and adhesion to Matrigel(®), and inhibited the VEGF-A secretion and VCAM-1 and ß3 integrin expressions by posttranscriptional pathways. Together, data herein outline novel posttranscriptional mechanisms of KPIs on endothelial cells during angiogenesis and point out the possible application of Amblyomin-X as a local inhibitor to undesired neovascularisation process.
Assuntos
Adesão Celular/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Regulação da Expressão Gênica , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteínas e Peptídeos Salivares/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Apoptose/efeitos dos fármacos , Proteínas de Artrópodes , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Dinoprostona/metabolismo , Células Endoteliais/metabolismo , Biblioteca Gênica , Masculino , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos , Óxido Nítrico/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas e Peptídeos Salivares/genética , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
The downregulation of PTA genes in mTECs is associated with the loss of self-tolerance, and the role of miRNAs in this process is not fully understood. Therefore, we studied the expression of mRNAs and miRNAs in mTECs from autoimmune NOD mice during the period when loss of self-tolerance occurs in parallel with non-autoimmune BALB/c mice. Although the expression of the transcriptional regulator Aire was unchanged, we observed downregulation of a set of PTA mRNAs. A set of miRNAs was also differentially expressed in these mice. The reconstruction of miRNA-mRNA interaction networks identified the controller miRNAs and predicted the PTA mRNA targets. Interestingly, the known Aire-dependent PTAs exhibited pronounced refractoriness in the networking interaction with miRNAs. This study reveals the existence of a new mechanism in mTECs, and this mechanism may have importance in the control of self-tolerance.
Assuntos
Antígenos/genética , Epistasia Genética , Células Epiteliais/metabolismo , MicroRNAs/genética , Interferência de RNA , RNA Mensageiro/genética , Timo/citologia , Fatores de Transcrição/metabolismo , Animais , Animais Recém-Nascidos , Antígenos/imunologia , Análise por Conglomerados , Células Epiteliais/imunologia , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Camundongos , Tolerância a Antígenos Próprios , Fatores de Transcrição/genética , Transcrição Gênica , Proteína AIRERESUMO
In plants, sugars such as glucose act as signalling molecules that promote changes in gene expression programmes that impact on growth and development. Recent evidence has revealed the potential importance of controlling mRNA decay in some aspects of glucose-mediated regulatory responses suggesting a role of microRNAs (miRNAs) in these responses. In order to get a better understanding of glucose-mediated development modulation involving miRNA-related regulatory pathways, early seedling development of mutants impaired in miRNA biogenesis (hyl1-2 and dcl1-11) and miRNA activity (ago1-25) was evaluated. All mutants exhibited a glucose hyposensitive phenotype from germination up to seedling establishment, indicating that miRNA regulatory pathways are involved in the glucose-mediated delay of early seedling development. The expression profile of 200 miRNA primary transcripts (pri-miRs) was evaluated by large-scale quantitative real-time PCR profiling, which revealed that 38 pri-miRs were regulated by glucose. For several of them, the corresponding mature miRNAs are known to participate directly or indirectly in plant development, and their accumulation was shown to be co-regulated with the pri-miR by glucose. Furthermore, the expression of several miRNA target genes was found to be deregulated in response to glucose in the miRNA machinery mutants ago1-25, dcl1-11, and hyl1-2. Also, in these mutants, glucose promoted misexpression of genes for the three abscisic acid signalling elements ABI3, ABI4, and ABI5. Thus, miRNA regulatory pathways play a role in the adjustments of growth and development triggered by glucose signalling.
Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Redes Reguladoras de Genes/genética , Glucose/farmacologia , MicroRNAs/metabolismo , Plântula/crescimento & desenvolvimento , Plântula/genética , Arabidopsis/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Redes Reguladoras de Genes/efeitos dos fármacos , Germinação/efeitos dos fármacos , Germinação/genética , MicroRNAs/genética , Mutação/genética , Fenótipo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Plântula/efeitos dos fármacosRESUMO
In eukaryotic cells, a group of messenger ribonucleic acids (mRNAs) encoding functionally interrelated proteins together with the trans-acting factors that coordinately modulate their expression is termed a post-transcriptional regulon, due to their partial analogy to a prokaryotic polycistron. This mRNA clustering is organized by sequence-specific RNA-binding proteins (RBPs) that bind cis-regulatory elements in the noncoding regions of genes, and mediates the synchronized control of their fate. These recognition motifs are often characterized by conserved sequences and/or RNA structures, and it is likely that various classes of cis-elements remain undiscovered. Current evidence suggests that RNA regulons govern gene expression in trypanosomes, unicellular parasites which mainly use post-transcriptional mechanisms to control protein synthesis. In this study, we used motif discovery tools to test whether groups of functionally related trypanosomatid genes contain a common cis-regulatory element. We obtained conserved structured RNA motifs statistically enriched in the noncoding region of 38 out of 53 groups of metabolically related transcripts in comparison with a random control. These motifs have a hairpin loop structure, a preferred sense orientation and are located in close proximity to the open reading frames. We found that 15 out of these 38 groups represent unique motifs in which most 3'-UTR signature elements were group-specific. Two extensively studied Trypanosoma cruzi RBPs, TcUBP1 and TcRBP3 were found associated with a few candidate RNA regulons. Interestingly, 13 motifs showed a strong correlation with clusters of developmentally co-expressed genes and six RNA elements were enriched in gene clusters affected after hyperosmotic stress. Here we report a systematic genome-wide in silico screen to search for novel RNA-binding sites in transcripts, and describe an organized network of several coordinately regulated cohorts of mRNAs in T. cruzi. Moreover, we found that structured RNA elements are also conserved in other human pathogens. These results support a model of regulation of gene expression by multiple post-transcriptional regulons in trypanosomes.
RESUMO
Trypanosoma cruzi, a protozoan parasite that causes Chagas disease, exhibits unique mechanisms for gene expression such as constitutive polycistronic transcription of protein-coding genes, RNA editing and trans-splicing. In the absence of mechanism controlling transcription initiation, organized subsets of T. cruzi genes must be post-transcriptionally co-regulated in response to extracellular signals. The mechanisms that regulate stage-specific gene expression in this parasite have become much clearer through sequencing its whole genome as well as performing various proteomic and microarray analyses, which have demonstrated that at least half of the T. cruzi genes are differentially regulated during its life cycle. In this review, we attempt to highlight the recent advances in characterising cis and trans-acting elements in the T. cruzi genome that are involved in its post-transcriptional regulatory machinery.