Post-transcriptional Regulatory Mechanisms Research Articles

Genome-wide transcriptomics and microRNAomics analyses uncover multi-faceted mechanisms to cope with copper stress in ancient macrobenthos amphioxus

The mechanisms underlying the toxicity of environmental stress are unclear for marine macrobenthos. Copper/Cu has posed the most serious threats to amphioxus, an ancient and model benthic cephalochordate. Herein, a dynamic change in the physiological parameters (GR, SOD, ATP, and MDA) was detected with ROS accumulation in Branchiostoma belcheri exposed to 0.3 mg·L−1 Cu. Transcriptomes and microRNAomes of B. belcheri were generated to investigate the molecular mechanisms by which this amphioxus copes with Cu exposure. Time-specific genes identified at different time points after exposure were involved in the stimulus and immune response, detoxification and ionic homeostasis, aging and the nervous system, sequentially, with prolongation of exposure time, forming a dynamic process of molecular response to Cu stress. In total, 57 differentially expressed miRNAs were identified under Cu stress. Transcriptomics–miRNAomics analyses indicate that these miRNAs targeted genes associated with many key biological processes such as xenobiotics degradation, oxidative stress, and energy metabolism. The constructed miRNA–mRNA-pathway network uncovered a broad post-transcriptional regulatory mechanism in B. belcheri to cope with Cu stress. Overall, this integrated analyses show that enhanced defense response, accelerated ROS elimination, and repressed ATP production constitute a comprehensive strategy to cope with Cu toxicity in the ancient macrobenthos.

Stem Cell-Associated Signatures Help to Predict Diagnosis and Prognosis in Ovarian Serous Cystadenocarcinoma.

Ovarian serous cystadenocarcinoma (OV) is a fatal gynecologic cancer with a five-year survival rate of only 46%. Resistance to platinum-based chemotherapy is a prevalent factor in OV patients, leading to increased mortality. The platinum resistance in OV is driven by transcriptome heterogeneity and tumor heterogeneity. Studies have indicated that ovarian cancer stem cells (OCSCs), which are chemoresistant and help in disease recurrence, are enriched by platinum-based chemotherapy. Stem cells have a significant influence on the OV progression and prognosis of OV patients and are key pathology mediators of OV. However, the molecular mechanisms and targets of OV have not yet been fully understood. In this study, systematic research based on the TCGA-OV dataset was conducted for the identification and construction of key stem cell-related diagnostic and prognostic models for the development of multigene markers of OV. A six-gene diagnostic and prognostic model (C19orf33, CBX2, CSMD1, INSRR, PRLR, and SLC38A4) was developed based on the differentially expressed stem cell-related gene model, which can act as a potent diagnostic biomarker and can characterize the clinicopathological properties of OV. The key genes related to stem cells were identified by screening the genes differentially expressed in OV and control samples. The mRNA-miRNA-TF molecular network for the six-gene model was constructed, and the potential biological significance of this molecular model and its impact on the infiltration of immune cells in the OV tumor microenvironment were elucidated. The differences in immune infiltration and stem cell-related biological processes were determined using gene set variation analysis (GSVA) and single-sample gene set enrichment analysis (ssGSEA) for the selection of molecular treatment options and providing a reference for elucidating the posttranscriptional regulatory mechanisms in OV.

Features of CFTR mRNA and implications for therapeutics development.

Cystic fibrosis (CF) is an autosomal recessive disease impacting ∼100,000 people worldwide. This lethal disorder is caused by mutation of the CF transmembrane conductance regulator (CFTR) gene, which encodes an ATP-binding cassette-class C protein. More than 2,100 variants have been identified throughout the length of CFTR. These defects confer differing levels of severity in mRNA and/or protein synthesis, folding, gating, and turnover. Drug discovery efforts have resulted in recent development of modulator therapies that improve clinical outcomes for people living with CF. However, a significant portion of the CF population has demonstrated either no response and/or adverse reactions to small molecules. Additional therapeutic options are needed to restore underlying genetic defects for all patients, particularly individuals carrying rare or refractory CFTR variants. Concerted focus has been placed on rescuing variants that encode truncated CFTR protein, which also harbor abnormalities in mRNA synthesis and stability. The current mini-review provides an overview of CFTR mRNA features known to elicit functional consequences on final protein conformation and function, including considerations for RNA-directed therapies under investigation. Alternative exon usage in the 5'-untranslated region, polypyrimidine tracts, and other sequence elements that influence splicing are discussed. Additionally, we describe mechanisms of CFTR mRNA decay and post-transcriptional regulation mediated through interactions with the 3'-untranslated region (e.g. poly-uracil sequences, microRNAs). Contributions of synonymous single nucleotide polymorphisms to CFTR transcript utilization are also examined. Comprehensive understanding of CFTR RNA biology will be imperative for optimizing future therapeutic endeavors intended to address presently untreatable forms of CF.

Analysis of alternative splicing in chicken macrophages infected with avian pathogenic E. coli (APEC)

Colibacillosis is a complex disease that caused by avian pathogenic Escherichia coli (APEC), resulting in huge economic loss to the global poultry industry and threatening to human health. Alternative splicing (AS) is a universal post-transcriptional regulatory mechanism, which can simultaneously produce many proteins from a single gene to involve in various diseases and individual development. Herein, we characterized genome-wide AS events in wild type macrophages (WT) and APEC infected macrophages (APEC) by high-throughput RNA sequencing technology. A total of 751 differentially expressed (DE) AS genes were identified in the comparison of APEC vs. WT, including 587 of SE, 114 of MXE, 25 of RI, 17 of A3 and 8 of A5 event. Functional analysis showed that these identified DE AS genes were involved in ‘Endocytosis’, ‘p53 signaling pathway’, ‘MAPK signaling pathway’, ‘NOD-like receptor signaling pathway’, ‘Ubiquitin mediated proteolysis’ and ‘Focal adhesion’ immune related pathways. In summary, we comprehensively investigate AS events during APEC infection. This study has expanded our understanding of the process of APEC infection and provided new insights for further treatment options for APEC infection.

Cassava MeRS40 is required for the regulation of plant salt tolerance

Soil salinity affects the expression of serine/arginine-rich (SR) genes and isoforms by alternative splicing, which in turn regulates the adaptation of plants to stress. We previously identified the cassava spliceosomal component 35 like (SCL) and SR subfamilies, belonging to the SR protein family, which are extensively involved in responses to abiotic stresses. However, the post-transcriptional regulatory mechanism of cassava arginine/serine-rich (RS) subfamily in response to salt stress remains to be explored. In the current study, we identified 37 genes of the RS subfamily from 11 plant species and systematically investigated the transcript levels of the RS40 and RS31 genes under diverse abiotic stress conditions. Subsequently, an analysis of the conserved protein domains revealed that plant RS subfamily genes were likely to preserve their conserved molecular functions and played critical functional roles in responses to abiotic stresses. Importantly, we found that overexpression of MeRS40 in Arabidopsis enhanced salt tolerance by maintaining reactive oxygen species homeostasis and up-regulating the salt-responsive genes. However, overexpression of MeRS40 gene in cassava reduced salt tolerance due to the depression of its endogenous gene expression by negative autoregulation of its own pre-mRNA. Moreover, the MeRS40 protein interacted with MeU1-70Ks (MeU1-70Ka and MeU1-70Kb) in vivo and in vitro, respectively. Therefore, our ?ndings highlight the critical role of cassava SR proteins in responses to salt stress in plants.

Abstract SY04-01: Understanding and targeting RNA splicing in breast cancer

Abstract Upregulation of SR proteins, a family of 14 essential splicing factors (SFs), often occurs in solid tumors, including in breast, and promotes mammary cell transformation and metastasis in breast cancer models. SR protein alterations often occur without genomic copy number changes, suggesting misregulation at the transcriptional or post-transcriptional levels; however the underlying molecular mechanisms are incompletely understood. Here we investigate the transcriptional and post-transcriptional regulatory mechanisms that control SF-levels in normal mammary cells and their misregulation in cancer cells. We demonstrate the MYC oncogene directly regulates a network of splicing factors that are co-expressed in tumors and act together to regulate their downstream targets and promote cell invasion. In parallel, our work reveals the role of non-coding ‘poison exon’ in regulating SR proteins levels during cell differentiation and in tumors. We uncover an extensive cross-regulatory network used by the SR protein family to control their expression. Finally, we develop splice-switching antisense oligonucleotides to reverse the increased skipping of TRA2β poison exon detected in tumors, leading to selective anti-proliferative effects across a variety of cancer models. In summary, our work provides new insights into the regulatory mechanisms of SR proteins and identifies novel oncogenic spliced isoforms that represent potential biomarkers and targets for therapeutics development. Citation Format: Olga A. Anczukow. Understanding and targeting RNA splicing in breast cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr SY04-01.

Iron restriction increases the expression of a cytotoxic cysteine proteinase TvCP2 by a novel mechanism of tvcp2 mRNA alternative polyadenylation in Trichomonas vaginalis

Trichomonas vaginalis TvCP2 (TVAG_057000) is a cytotoxic cysteine proteinase (CP) expressed under iron-limited conditions. This work aimed to identify one of the mechanisms of tvcp2 gene expression regulation by iron at the posttranscriptional level. We checked tvcp2 mRNA stability under both iron-restricted (IR) and high iron (HI) conditions in the presence of actinomycin D. Greater stability of the tvcp2 mRNA under the IR than in HI conditions was observed, as expected. In silico analysis of the 3′ regulatory region showed the presence of two putative polyadenylation signals in the tvcp2 transcript. By 3′-RACE assays, we demonstrated the existence of two isoforms of the tvcp2 mRNA with different 3′-UTR that resulted in more TvCP2 protein under IR than in HI conditions detected by WB assays. Additionally, we searched for homologs of the trichomonad polyadenylation machinery by an in silico analysis in the genome database, TrichDB. 16 genes that encode proteins that could be part of the trichomonad polyadenylation machinery were found. qRT-PCR assays showed that most of these genes were positively regulated by iron. Thus, our results show the presence of alternative polyadenylation as a novel iron posttranscriptional regulatory mechanism in T. vaginalis for the tvcp2 gene expression.

Meet the First Authors.

Meet the First Authors.

The RNA-binding protein hnRNP F is required for the germinal center B cell response

The T cell-dependent (TD) antibody response involves the generation of high affinity, immunoglobulin heavy chain class-switched antibodies that are generated through germinal center (GC) response. This process is controlled by coordinated transcriptional and post-transcriptional gene regulatory mechanisms. RNA-binding proteins (RBPs) have emerged as critical players in post-transcriptional gene regulation. Here we demonstrate that B cell-specific deletion of RBP hnRNP F leads to diminished production of class-switched antibodies with high affinities in response to a TD antigen challenge. B cells deficient in hnRNP F are characterized by defective proliferation and c-Myc upregulation upon antigenic stimulation. Mechanistically, hnRNP F directly binds to the G-tracts of Cd40 pre-mRNA to promote the inclusion of Cd40 exon 6 that encodes its transmembrane domain, thus enabling appropriate CD40 cell surface expression. Furthermore, we find that hnRNP A1 and A2B1 can bind to the same region of Cd40 pre-mRNA but suppress exon 6 inclusion, suggesting that these hnRNPs and hnRNP F might antagonize each-other’s effects on Cd40 splicing. In summary, our study uncovers an important posttranscriptional mechanism regulating the GC response.

Integrated transcriptome and proteome analysis reveals posttranscriptional regulation of ribosomal genes in human brain organoids.

During development of the human cerebral cortex, multipotent neural progenitors generate excitatory neurons and glial cells. Investigations of the transcriptome and epigenome have revealed important gene regulatory networks underlying this crucial developmental event. However, the posttranscriptional control of gene expression and protein abundance during human corticogenesis remains poorly understood. We addressed this issue by using human telencephalic brain organoids grown using a dual reporter cell line to isolate neural progenitors and neurons and performed cell class and developmental stage-specific transcriptome and proteome analysis. Integrating the two datasets revealed modules of gene expression during human corticogenesis. Investigation of one such module uncovered mTOR-mediated regulation of translation of the 5'TOP element-enriched translation machinery in early progenitor cells. We show that in early progenitors partial inhibition of the translation of ribosomal genes prevents precocious translation of differentiation markers. Overall, our multiomics approach proposes novel posttranscriptional regulatory mechanisms crucial for the fidelity of cortical development.

A Putative TRAPα Protein of Microsporidia Nosema bombycis Exhibits Non-Canonical Alternative Polyadenylation in Transcripts.

Microsporidia are obligate intracellular eukaryotic parasites that have significantly reduced genomes and that have lost most of their introns. In the current study, we characterized a gene in microsporidia Nosema bombycis, annotated as TRAPα (HNbTRAPα). The homologous of TRAPα are a functional component of ER translocon and facilitates the initiation of protein translocation in a substrate-specific manner, which is conserved in animals but absent from most fungi. The coding sequence of HNbTRAPα consists of 2226 nucleotides, longer than the majority of homologs in microsporidia. A 3' RACE analysis indicated that there were two mRNA isoforms resulting from non-canonical alternative polyadenylation (APA), and the polyadenylate tail was synthesized after the C951 or C1167 nucleotide, respectively. Indirect immunofluorescence analysis showed two different localization characteristics of HNbTRAPα, which are mainly located around the nuclear throughout the proliferation stage and co-localized with the nuclear in mature spores. This study demonstrated that the post-transcriptional regulation mechanism exists in Microsporidia and expands the mRNA isoform repertoire.

HnRNPA2B1 Aggravates Inflammation by Promoting M1 Macrophage Polarization.

Macrophages have critical contributions to both acute and chronic inflammatory diseases, for example, bowel disease and obesity, respectively. However, little is known about the post-transcriptional regulatory mechanisms in macrophage-mediated inflammatory diseases. hnRNPA2B1 (A2B1) is an RNA binding protein for mRNA fate determination. We showed that hnRNPA2B1 mRNA levels were increased in colon in dextran sodium sulfate (DSS)-induced colitis mice and in epididymal white adipose tissue (eWAT) and spleen of high-fat-diet (HFD)-induced obese mice. Consistently, mice with haploinsufficiency of A2B1 (A2B1 HET) are protected against DSS-induced acute colitis and HFD-induced obesity, with decreased M1 macrophages polarization in colon, eWAT and spleen. Mechanistically, A2B1 mRNA and protein levels were increased in LPS-stimulated RAW 264.7 macrophages, and A2B1 enhanced RNA stability of pro-inflammatory genes Tnfα, Il-6 and Il-1β for the regulation of macrophages polarization. Interestingly, A2B1 HET mice exhibited reduced white fat expansion, which was influenced by macrophages, since conditioned medium from macrophages with A2B1 manipulation significantly changed preadipocyte proliferation. Our data demonstrate that A2B1 plays a vital role in macrophage-mediated inflammation via regulating mRNA stability, suggesting that A2B1 may be served as a promising target for the intervention of acute and chronic inflammatory diseases.

Disentangling the Potential Functions of miRNAs in the Synthesis of Terpenoids during the Development of Cinnamomum burmannii Leaves

The essential oil of Cinnamomum burmannii (Nees and T. Nees) Blume is rich in monoterpenes and sesquiterpenes. The post-transcriptional regulatory mechanisms controlling the expression of terpenoid-related genes have not yet been clarified in C. burmannii. Here, we conducted a metabolomic analysis of the leaves of C. burmannii across four developmental stages using gas chromatography–mass spectrometry. We also identified miRNAs and their target genes involved in terpenoid biosynthesis using small RNA sequencing. A total of 135 differentially expressed metabolites were detected, including 65 terpenoids, 15 aldehydes, and 13 benzenes. A total of 876 miRNAs from 148 families were detected, among which 434 miRNAs were differentially expressed, including three known miRNAs and 431 novel miRNAs. Four miRNAs (gma-miR5368, novel_miR_377, novel_miR_111, and novel_miR_251) were predicted to regulate the expression of four differential expressed genes involved in the monoterpenoid and sesquiterpenoid synthesis. miRNAs families miR396, miR5185, and miR9408 were predicted to play diverse regulatory roles in monoterpenoid and sesquiterpenoid synthesis during the leaf development of C. burmannii. The results of our study shed new light on the roles of regulatory genes in terpenoid biosynthesis. Our findings also have implications for the further promotion of essential oil production using the leaves of C. burmannii.

Global Analysis of Dark- and Heat-Regulated Alternative Splicing in Arabidopsis

Alternative splicing (AS) is one of the major post-transcriptional regulation mechanisms that contributes to plant responses to various environmental perturbations. Darkness and heat are two common abiotic factors affecting plant growth, yet the involvement and regulation of AS in the plant responses to these signals remain insufficiently examined. In this study, we subjected Arabidopsis seedlings to 6 h of darkness or heat stress and analyzed their transcriptome through short-read RNA sequencing. We revealed that both treatments altered the transcription and AS of a subset of genes yet with different mechanisms. Dark-regulated AS events were found enriched in photosynthesis and light signaling pathways, while heat-regulated AS events were enriched in responses to abiotic stresses but not in heat-responsive genes, which responded primarily through transcriptional regulation. The AS of splicing-related genes (SRGs) was susceptible to both treatments; while dark treatment mostly regulated the AS of these genes, heat had a strong effect on both their transcription and AS. PCR analysis showed that the AS of the Serine/Arginine-rich family gene SR30 was reversely regulated by dark and heat, and heat induced the upregulation of multiple minor SR30 isoforms with intron retention. Our results suggest that AS participates in plant responses to these two abiotic signals and reveal the regulation of splicing regulators during these processes.

Inhibition of Nonsense-Mediated Decay Induces Nociceptive Sensitization through Activation of the Integrated Stress Response.

RNA stability is meticulously controlled. Here, we sought to determine whether an essential post-transcriptional regulatory mechanism plays a role in pain. Nonsense-mediated decay (NMD) safeguards against translation of mRNAs that harbor premature termination codons and controls the stability of ∼10% of typical protein-coding mRNAs. It hinges on the activity of the conserved kinase SMG1. Both SMG1 and its target, UPF1, are expressed in murine DRG sensory neurons. SMG1 protein is present in both the DRG and sciatic nerve. Using high-throughput sequencing, we examined changes in mRNA abundance following inhibition of SMG1. We confirmed multiple NMD stability targets in sensory neurons, including ATF4. ATF4 is preferentially translated during the integrated stress response (ISR). This led us to ask whether suspension of NMD induces the ISR. Inhibition of NMD increased eIF2-α phosphorylation and reduced the abundance of the eIF2-α phosphatase constitutive repressor of eIF2-α phosphorylation. Finally, we examined the effects of SMG1 inhibition on pain-associated behaviors. Peripheral inhibition of SMG1 results in mechanical hypersensitivity in males and females that persists for several days and priming to a subthreshold dose of PGE2. Priming was fully rescued by a small-molecule inhibitor of the ISR. Collectively, our results indicate that suspension of NMD promotes pain through stimulation of the ISR.SIGNIFICANCE STATEMENT Nociceptors undergo long-lived changes in their plasticity which may contribute to chronic pain. Translational regulation has emerged as a dominant mechanism in pain. Here, we investigate the role of a major pathway of RNA surveillance called nonsense-mediated decay (NMD). Modulation of NMD is potentially beneficial for a broad array of diseases caused by frameshift or nonsense mutations. Our results suggest that inhibition of the rate-limiting step of NMD drives behaviors associated with pain through activation of the ISR. This work reveals complex interconnectivity between RNA stability and translational regulation and suggests an important consideration in harnessing the salubrious benefits of NMD disruption.

SRSF1 regulates primordial follicle formation and number determination during meiotic prophase I

BackgroundOvarian folliculogenesis is a tightly regulated process leading to the formation of functional oocytes and involving successive quality control mechanisms that monitor chromosomal DNA integrity and meiotic recombination. A number of factors and mechanisms have been suggested to be involved in folliculogenesis and associated with premature ovarian insufficiency, including abnormal alternative splicing (AS) of pre-mRNAs. Serine/arginine-rich splicing factor 1 (SRSF1; previously SF2/ASF) is a pivotal posttranscriptional regulator of gene expression in various biological processes. However, the physiological roles and mechanism of SRSF1 action in mouse early-stage oocytes remain elusive. Here, we show that SRSF1 is essential for primordial follicle formation and number determination during meiotic prophase I.ResultsThe conditional knockout (cKO) of Srsf1 in mouse oocytes impairs primordial follicle formation and leads to primary ovarian insufficiency (POI). Oocyte-specific genes that regulate primordial follicle formation (e.g., Lhx8, Nobox, Sohlh1, Sohlh2, Figla, Kit, Jag1, and Rac1) are suppressed in newborn Stra8-GFPCre Srsf1Fl/Fl mouse ovaries. However, meiotic defects are the leading cause of abnormal primordial follicle formation. Immunofluorescence analyses suggest that failed synapsis and an inability to undergo recombination result in fewer homologous DNA crossovers (COs) in the Srsf1 cKO mouse ovaries. Moreover, SRSF1 directly binds and regulates the expression of the POI-related genes Six6os1 and Msh5 via AS to implement the meiotic prophase I program.ConclusionsAltogether, our data reveal the critical role of an SRSF1-mediated posttranscriptional regulatory mechanism in the mouse oocyte meiotic prophase I program, providing a framework to elucidate the molecular mechanisms of the posttranscriptional network underlying primordial follicle formation.

Lengthening of 3′ Untranslated Regions of mRNAs by Alternative Polyadenylation Is Associated With Tumor Progression and Poor Prognosis of Clear Cell Renal Cell Carcinoma

Alternative polyadenylation (APA) is emerging as a major posttranscriptional mechanism for gene regulation in cancer. A prevailing hypothesis is that shortening of the 3′ untranslated region (3′UTR) increases oncoprotein expression because of the loss of miRNA-binding sites (MBSs). We showed that the longer 3′UTR is associated with a more advanced tumor stage in patients with clear cell renal cell carcinoma (ccRCC). More surprisingly, 3′UTR shortening is correlated with better overall survival in patients with ccRCC. Furthermore, we identified a mechanism by which longer transcripts lead to increased oncogenic protein and decreased tumor-suppressive protein expression compared to the shorter transcripts. In our model, shortening of 3′UTRs by APA may increase the mRNA stability of the majority of the potential tumor-suppressor genes due to the loss of MBSs and AU-rich elements (AREs). Unlike potential tumor-suppressor genes, the potential oncogenes display much lower MBS and ARE density and globally much higher m6A density in distal 3′UTRs. As a result, 3′UTRs shortening decreases the mRNA stability of potential oncogenes and enhances the mRNA stability of potential tumor-suppressor genes. Our findings highlight the cancer-specific pattern of APA regulation and extend our understanding of the mechanism of APA-mediated 3′UTR length changes in cancer biology.

Alternative splicing in shaping the molecular landscape of the cochlea.

The cochlea is a complex organ comprising diverse cell types with highly specialized morphology and function. Until now, the molecular underpinnings of its specializations have mostly been studied from a transcriptional perspective, but accumulating evidence points to post-transcriptional regulation as a major source of molecular diversity. Alternative splicing is one of the most prevalent and well-characterized post-transcriptional regulatory mechanisms. Many molecules important for hearing, such as cadherin 23 or harmonin, undergo alternative splicing to produce functionally distinct isoforms. Some isoforms are expressed specifically in the cochlea, while some show differential expression across the various cochlear cell types and anatomical regions. Clinical phenotypes that arise from mutations affecting specific splice variants testify to the functional relevance of these isoforms. All these clues point to an essential role for alternative splicing in shaping the unique molecular landscape of the cochlea. Although the regulatory mechanisms controlling alternative splicing in the cochlea are poorly characterized, there are animal models with defective splicing regulators that demonstrate the importance of RNA-binding proteins in maintaining cochlear function and cell survival. Recent technological breakthroughs offer exciting prospects for overcoming some of the long-standing hurdles that have complicated the analysis of alternative splicing in the cochlea. Efforts toward this end will help clarify how the remarkable diversity of the cochlear transcriptome is both established and maintained.

Alternative Splicing in the Regulatory Circuit of Plant Temperature Response.

As sessile organisms, plants have evolved complex mechanisms to rapidly respond to ever-changing ambient temperatures. Temperature response in plants is modulated by a multilayer regulatory network, including transcriptional and post-transcriptional regulations. Alternative splicing (AS) is an essential post-transcriptional regulatory mechanism. Extensive studies have confirmed its key role in plant temperature response, from adjustment to diurnal and seasonal temperature changes to response to extreme temperatures, which has been well documented by previous reviews. As a key node in the temperature response regulatory network, AS can be modulated by various upstream regulations, such as chromatin modification, transcription rate, RNA binding proteins, RNA structure and RNA modifications. Meanwhile, a number of downstream mechanisms are affected by AS, such as nonsense-mediated mRNA decay (NMD) pathway, translation efficiency and production of different protein variants. In this review, we focus on the links between splicing regulation and other mechanisms in plant temperature response. Recent advances regarding how AS is regulated and the following consequences in gene functional modulation in plant temperature response will be discussed. Substantial evidence suggests that a multilayer regulatory network integrating AS in plant temperature response has been unveiled.

Transcriptomics-proteomics Integration reveals alternative polyadenylation driving inflammation-related protein translation in patients with diabetic nephropathy

BackgroundDiabetic nephropathy (DN) is a complex disease involving the upregulation of many inflammation-related proteins. Alternative polyadenylation (APA), a crucial post-transcriptional regulatory mechanism, has been proven to play vital roles in many inflammatory diseases. However, it is largely unknown whether and how APA exerts function in DN.MethodsWe performed transcriptomics and proteomics analysis of glomeruli samples isolated from 50 biopsy-proven DN patients and 25 control subjects. DaPars and QAPA algorithms were adopted to identify APA events from RNA-seq data. The qRT-PCR analysis was conducted to verify 3′UTR length alteration. Short and long 3ʹUTRs isoforms were also overexpressed in podocytes under hyperglycemia condition for examining protein expression.ResultsWe detected transcriptome-wide 3′UTR APA events in DN, and found that APA-mediated 3ʹUTR lengthening of genes (APA genes) increased their expression at protein but not mRNA level. Increased protein level of 3′UTR lengthening gene was validated in podocytes under hyperglycemia condition. Pathway enrichment analysis showed that APA genes were enriched in inflammation-related biological processes including endoplasmic reticulum stress pathways, NF-κB signaling and autophagy. Further bioinformatics analysis demonstrated that 3′UTR APA of genes probably altered the binding sites for RNA-binding proteins, thus enhancing protein translation.ConclusionThis study revealed for the first time that 3′UTR lengthening of APA genes contributed to the progression of DN by elevating the translation of corresponding proteins, providing new insight and a rich resource for investigating DN mechanisms.