Mining sponge phenomena in RNA expression data

In the last few years, the interactions among competing endogenous RNAs (ceRNAs) have been recognized as a key post-transcriptional regulatory mechanism in cell differentiation, tissue development, and disease. Notably, such sponge phenomena substracting active microRNAs from their silencing targets have been recognized as having a potential oncosuppressive, or oncogenic, role in several cancer types. Hence, the ability to predict sponges from the analysis of large expression data sets (e.g. from international cancer projects) has become an important data mining task in bioinformatics. We present a technique designed to mine sponge phenomena whose presence or absence may discriminate between healthy and unhealthy populations of samples in tumoral or normal expression data sets, thus providing lists of candidates potentially relevant in the pathology. With this aim, we search for pairs of elements acting as ceRNA for a given miRNA, namely, we aim at discovering miRNA-RNA pairs involved in phenomena which are clearly present in one population and almost absent in the other one. The results on tumoral expression data, concerning five different cancer types, confirmed the effectiveness of the approach in mining interesting knowledge. Indeed, 32 out of 33 miRNAs and 22 out of 25 protein-coding genes identified as top scoring in our analysis are corroborated by having been similarly associated with cancer processes in independent studies. In fact, the subset of miRNAs selected by the sponge analysis results in a significant enrichment of annotation for the KEGG32 pathway “microRNAs in cancer” when tested with the commonly used bioinformatic resource DAVID. Moreover, often the cancer datasets where our sponge analysis identified a miRNA as top scoring match the one reported already in the pertaining literature.

Tobramycin suppresses cystic fibrosis lung inflammation by increasing 5′ tRNA-fMet halves secreted by P. aeruginosa

Although inhaled tobramycin increases lung function in people with cystic fibrosis (pwCF), the density of P. aeruginosa in the lungs is only modestly reduced; hence, the mechanism whereby tobramycin improves lung function remains unclear. Previously, we demonstrated that P. aeruginosa secretes outer membrane vesicles (OMVs) that fuse with bronchial epithelial cells (HBECs), delivering small RNAs (sRNAs) that suppress the host immune response. Thus, we hypothesized that tobramycin modifies the sRNA content of OMVs leading to reduced inflammation and neutrophil-mediated lung damage. We found that tobramycin increased the amount of two 5′ tRNA-fMet halves in OMVs (Tobi-OMVs) and that Tobi-OMVs elicited less IL-8 secretion by CF-HBECs than control OMVs (ctrl-OMVs). A specific 5′ tRNA-fMet halves inhibitor reduced the ability of Tobi-OMVs to suppress IL-8 secretion. Tobi-OMVs were also less effective in stimulating KC secretion and neutrophil recruitment in mouse lungs compared to ctrl-OMVs. Tobramycin also reduced IL-8 and neutrophil abundance in bronchoalveolar lavage fluid obtained from pwCF. The 5′ tRNA-fMet halves reduced IL-8 secretion by an AGO2-mediated post-transcriptional regulatory mechanism. The clinical benefit of tobramycin is partly due to an increase in the secretion of 5′ tRNA-fMet halves in OMVs, leading to the attenuation of IL-8 and neutrophil-mediated CF lung damage.

N6-Methyladenosine in Cancer Immunotherapy: An Undervalued Therapeutic Target

N6-methylation of adenosine (m6A), a post-transcriptional regulatory mechanism, is the most abundant nucleotide modification in almost all types of RNAs. The biological function of m6A in regulating the expression of oncogenes or tumor suppressor genes has been widely investigated in various cancers. However, recent studies have addressed a new role of m6A modification in the anti-tumor immune response. By modulating the fate of targeted RNA, m6A affects tumor-associated immune cell activation and infiltration in the tumor microenvironment (TME). In addition, m6A-targeting is found to affect the efficacy of classical immunotherapy, which makes m6A a potential target for immunotherapy. Although m6A modification together with its regulators may play the exact opposite role in different tumor types, targeting m6A regulators has been shown to have wide implications in several cancers. In this review, we discussed the link between m6A modification and tumor with an emphasis on the importance of m6A in anti-tumor immune response and immunotherapy.

Alternative Splicing: A New Cause and Potential Therapeutic Target in Autoimmune Disease

Alternative splicing (AS) is a complex coordinated transcriptional regulatory mechanism. It affects nearly 95% of all protein-coding genes and occurs in nearly all human organs. Aberrant alternative splicing can lead to various neurological diseases and cancers and is responsible for aging, infection, inflammation, immune and metabolic disorders, and so on. Though aberrant alternative splicing events and their regulatory mechanisms are widely recognized, the association between autoimmune disease and alternative splicing has not been extensively examined. Autoimmune diseases are characterized by the loss of tolerance of the immune system towards self-antigens and organ-specific or systemic inflammation and subsequent tissue damage. In the present review, we summarized the most recent reports on splicing events that occur in the immunopathogenesis of systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) and attempted to clarify the role that splicing events play in regulating autoimmune disease progression. We also identified the changes that occur in splicing factor expression. The foregoing information might improve our understanding of autoimmune diseases and help develop new diagnostic and therapeutic tools for them.

Genome-wide comprehensive analysis of miRNAs and their target genes expressed in resistant and susceptible Capsicum annuum landrace during Phytophthora capsici infection

MiRNAs regulate plants responses to fungal infection and immunity by modulating the gene expression. Despite extensive works on miRNA’s role during plant-fungus interaction, work in Capsicum annuum-Phytophthora capsici pathosystem is limited. Therefore, in the current study, genome-wide known and novel miRNAs were identified in two contrasting chilli pepper landraces, i.e. GojamMecha_9086 (resistant) and Dabat_80045 (susceptible) during P. capsici infection. The small RNA deep sequencing resulted in 79 known miRNAs corresponding to 24 miRNAs families and 477 novel miRNAs along with 22,895 potential targets, including 30 defence-related genes against P. capsici infection. The expression analysis of ∼29 known & 157 novel miRNAs in resistant and 30 known and 176 novel miRNAs in susceptible landrace revealed differential accumulation pattern. RT-qPCR of a set of 8 defence related miRNAs representing 4 novel (Pz-novel-miR428-1, Pz-novel-miR160-1, Pz-novel-miR1028-1, Pz-novel-miR204-1) and 4 known (Pz-known-miR803-1, Pz-known-miR2059-1, Pz-known-miR2560-1, Pz-known-miR1872-1) revealed differential accumulation pattern in both resistant and susceptible landrace. Additionally, validation of 8 target genes of corresponding miRNAs using RA-PCR, which as good as 5’ RLM-RACE, revealed an inverse relation with their corresponding miRNAs suggesting their key role during disease response. This study provides comprehensive genome-wide information about the repertoire of miRNAs and their target genes expressed in resistant and susceptible chilli pepper landrace, which can serve as a valuable resource for better understanding the post-transcriptional regulatory mechanism during C. annuum - P. capsici pathosystem.

Cleavage and Polyadenylation Specific Factor 1 Promotes Tumor Progression via Alternative Polyadenylation and Splicing in Hepatocellular Carcinoma

Alternative polyadenylation (APA) is an important post-transcriptional regulatory mechanism required for cleavage and polyadenylation (CPA) of the 3′ untranslated region (3′ UTR) of mRNAs. Several aberrant APA events have been reported in hepatocellular carcinoma (HCC). However, the regulatory mechanisms underlying APA remain unclear. In this study, we found that the expression of cleavage and polyadenylation specific factor 1 (CPSF1), a major component of the CPA complex, was significantly increased in HCC tissues and correlated with unfavorable survival outcomes. Knockdown of CPSF1 inhibited HCC cell proliferation and migration, whereas overexpression of CPSF1 caused the opposite effect. Based on integrative analysis of Iso-Seq and RNA-seq data from HepG2.2.15 cells, we identified a series of transcripts with differential 3′ UTR lengths following the knockdown of CPSF1. These transcripts were related to the biological functions of gene transcription, cytoskeleton maintenance, and endomembrane system transportation. Moreover, knockdown of CPSF1 induced an increase in alternative splicing (AS) events in addition to APA. Taken together, this study provides new insights into our understanding of the post-transcriptional regulatory mechanisms in HCC and implies that CPSF1 may be a potential prognostic biomarker and therapeutic target for HCC.

Alternative splicing events implicated in carcinogenesis and prognosis of thyroid gland cancer

Alternative splicing (AS), a critical post-transcriptional regulatory mechanism, expands gene expression patterns, thereby leading to increased protein diversity. Indeed, more than 95% of human genes undergo alternative splicing events (ASEs). In this study, we drew an all-around AS profile of thyroid cancer cells based on RNA-seq data. In total, there were 45,150 AS in 10,446 thyroid cancer cell genes derived from 506 patients, suggesting that ASEs is a common process in TC. Moreover, 1819 AS signatures were found to be significantly associated with the overall survival (OS) of TC patients. Kaplan–Meier survival analyses suggested that seven types of ASEs were associated with poor prognosis of TC (P < 0.05). Among them, exon skipping (ES) was the most common, with alternate promoter (AP) and alternate terminator (AT) coming second and third, respectively. Our results indicated that acceptor sites (AA) (AUC: 0.937), alternate donor sites (AD) (AUC: 0.965), AT (AUC: 0.964), ES (AUC: 0.999), mutually exclusive exons (ME) (AUC: 0.999), and retained intron (RI) (AUC: 0.837) exhibited an AUC greater than 0.6. In addition, age and risk score (All) were risk factors for TC patients. We also evaluated whether TC-ASEs are regulated by various splicing factors (SFs). We found that the expression of 90 SFs was associated with 469 ASEs and OS of TC patients. Our findings provide an insight into the role of spliceosomes in TC, which may offer novel perspectives in tumor research.

A fruit ripening-associated transcription factor CsMADS5 positively regulates carotenoid biosynthesis in citrus

Carotenoids in citrus contribute the quality of the fruit, but the transcriptional regulatory mechanism is still limitedly known. Here, we characterized a citrus FUL-like MADS gene, CsMADS5, that was ripening-inducible and acted as a nucleus-localized trans-activator. Transient overexpression of CsMADS5 in citrus induced fruit coloration and enhanced carotenoid contents. The expression levels of carotenogenic genes including phytoene synthase (PSY), phytoene desaturase (PDS), and lycopene β-cyclase 1 (LCYb1) were significantly increased in the peel of fruits overexpressing CsMADS5. Similar results were observed from stable overexpression of CsMADS5 in tomato fruits and citrus calli, even though the effect of CsMADS5 on the carotenoid metabolism in transgenic citrus calli was limited. Further biochemical analyses demonstrated that CsMADS5 activated the transcription of PSY, PDS, and LCYb1 by directly binding to their promoters. It is concluded that CsMADS5 positively regulates carotenoid biosynthesis in fruits by directly activating the transcription of carotenogenic genes. Moreover, CsMADS5 physically interacted with a positive regulator CsMADS6, indicating that CsMADS5 may form an enhancer complex with CsMADS6 to synergistically promote carotenoid accumulation. These findings expand our understanding of the complex transcriptional regulatory hierarchy for carotenoid biosynthesis during fruit ripening.

Genome-wide analysis of DNA G-quadruplex motifs across 37 species provides insights into G4 evolution

G-quadruplex (G4) structures have been predicted in the genomes of many organisms and proven to play regulatory roles in diverse cellular activities. However, there is little information on the evolutionary history and distribution characteristics of G4s. Here, whole-genome characteristics of potential G4s were studied in 37 evolutionarily representative species. During evolution, the number, length, and density of G4s generally increased. Immunofluorescence in seven species confirmed G4s’ presence and evolutionary pattern. G4s tended to cluster in chromosomes and were enriched in genetic regions. Short-loop G4s were conserved in most species, while loop-length diversity also existed, especially in mammals. The proportion of G4-bearing genes and orthologue genes, which appeared to be increasingly enriched in transcription factors, gradually increased. The antagonistic relationship between G4s and DNA methylation sites was detected. These findings imply that organisms may have evolutionarily developed G4 into a novel reversible and elaborate transcriptional regulatory mechanism benefiting multiple physiological activities of higher organisms.