RN7SK small nuclear RNA controls bidirectional transcription of highly expressed gene pairs in skin

Pausing of RNA polymerase II (Pol II) close to promoters is a common regulatory step in RNA synthesis, and is coordinated by a ribonucleoprotein complex scaffolded by the noncoding RNA RN7SK. The function of RN7SK-regulated gene transcription in adult tissue homoeostasis is currently unknown. Here, we deplete RN7SK during mouse and human epidermal stem cell differentiation. Unexpectedly, loss of this small nuclear RNA specifically reduces transcription of numerous cell cycle regulators leading to cell cycle exit and differentiation. Mechanistically, we show that RN7SK is required for efficient transcription of highly expressed gene pairs with bidirectional promoters, which in the epidermis co-regulated cell cycle and chromosome organization. The reduction in transcription involves impaired splicing and RNA decay, but occurs in the absence of chromatin remodelling at promoters and putative enhancers. Thus, RN7SK is directly required for efficient Pol II transcription of highly transcribed bidirectional gene pairs, and thereby exerts tissue-specific functions, such as maintaining a cycling cell population in the epidermis.

Long Non-Coding RNA SNHG12 Contributes to Cisplatin Resistance by Mediating WEE1 via miR-503-5p in Cervical Cancer

Background: Emerging evidences have indicated that the aberrant expression of long noncoding RNAs (lncRNAs) was responsible for drug resistance, which represents a major obstacle for chemotherapy failure. Our previous study has showed that small nuclear RNA host gene 12 (SNHG12) was increased and contributed to cell growth and invasion in cervical cancer. In the present study, we aimed to investigate the role of the lncRNA SNHG12 in cisplatin (DDP) resistance and elucidate its underlying mechanisms in cervical cancer.Methods: The expression and prognosis of SNHG12 in cervical cancer tissues were evaluated based on bioinformatics. MTT, colony formation assay and flow cytometer were performed to detect cell viability. Further, Molecular relationships among CTD-3252C9.4, IRF1 and IFI6 were investigated via luciferase reporter assay, western blot, and qRT-PCR. Finally, subcutaneous xenograft model was established to verify our findings.Results: In the present study, we evaluated the cell apoptosis and half maximal inhibitory concentration (IC50) of cervical cancer upon DDP treatment. Mechanically, we found that SNHG12 upregulated WEE1 expression to regulate cell and DDP resistance via sponging miR-503-5p. Moreover, SNHG12 silencing inhibited the growth of DDP-resistant cervical cancer tumors in vivo. Conclusions: Taken together, our findings suggested that a SNHG12/miR-503-5p/ WEE1 axis which modulated the chemoresistance of cervical cancer cell to DDP, and provided promising targets for dealing with the chemoresistance of cervical cancer.

Differential expression of small nuclear RNA activating complex polypeptide 2 in human epithelial ovarian cancer.

Epithelial ovarian cancer (EOC) is the most lethal gynecologic cancer (1). We performed discovery of genes associated with epithelial ovarian cancer and of the high-grade serous ovarian cancer (HGSC) subtype, using published and public microarray data (2, 3) to compare global gene expression profiles of normal ovary or fallopian tube with that of primary tumors from women diagnosed with epithelial ovarian cancer or HGSC. We identified the gene encoding small nuclear RNA activating complex polypeptide 2, SNAPC2, as among the genes whose expression was most different in epithelial ovarian cancer as compared to the normal fallopian tube. SNAPC2 expression was significantly lower in high-grade serous ovarian tumors relative to normal fallopian tube. SNAPC2 expression correlated with overall survival in patients with ovarian cancer. These data indicate that expression of SNAPC2 is perturbed in epithelial ovarian cancers broadly and in ovarian cancers of the HGSC subtype. SNAPC2 may be relevant to pathways underlying ovarian cancer initiation (transformation) or progression.

The snoGloBe interaction predictor enables a broader study of box C/D snoRNA functions and mechanisms

Box C/D small nucleolar RNAs (snoRNAs) are a conserved class of noncoding RNA known to serve as guides for the site-specific 2'-O-ribose methylation of ribosomal RNAs and the U6 small nuclear RNA, through direct base pairing with the target. In recent years however, several examples of box C/D snoRNAs regulating different levels of gene expression including transcript stability and splicing have been reported. These regulatory interactions typically require direct binding of the target but do not always involve the guide region. Supporting these new box C/D snoRNA functions, high-throughput RNA-RNA interaction datasets detect many interactions between box C/D snoRNAs and messenger RNAs. To facilitate the study of box C/D snoRNA functionality, we created snoGloBe, a box C/D snoRNA machine learning target predictor based on a gradient boosting classifier and considering snoRNA and target sequence and position as well as target type. SnoGloBe convincingly outperforms general RNA duplex predictors and PLEXY, the only box C/D snoRNA-specific target predictor available. The study of snoGloBe human transcriptome-wide predictions identifies enrichment in snoRNA interactions in exons and on exon-intron junctions. Some specific snoRNAs are predicted to target groups of functionally-related transcripts on common regulatory elements and the exact position of the predicted targets strongly overlaps binding sites of RNA-binding proteins involved in relevant molecular functions. SnoGloBe was also applied to predicting interactions between human box C/D snoRNAs and the SARS-CoV-2 transcriptome, identifying known and novel interactions. Overall, snoGloBe is a timely new tool that will accelerate our understanding of C/D snoRNA targets and function.

An alternative D. melanogaster 7SK snRNP

Background The 7SK small nuclear RNA (snRNA) found in most metazoans is a key regulator of P-TEFb which in turn regulates RNA polymerase II elongation. Although its primary sequence varies in protostomes, its secondary structure and function are conserved across evolutionary distant taxa. Results Here, we describe a novel ncRNA sharing many features characteristic of 7SK RNAs, in D. melanogaster. We examined the structure of the corresponding gene and determined the expression profiles of the encoded RNA, called snRNA:7SK:94F, during development. It is probably produced from the transcription of a lncRNA which is processed into a mature snRNA. We also addressed its biological function and we show that, like dm7SK, this alternative 7SK interacts in vivo with the different partners of the P-TEFb complex, i.e. HEXIM, LARP7 and Cyclin T. This novel RNA is widely expressed across tissues. Conclusion We propose that two distinct 7SK genes might contribute to the formation of the 7SK snRNP complex in D. melanogaster.

Mutational Landscape and Interaction of SARS-CoV-2 with Host Cellular Components

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its rapid evolution has led to a global health crisis. Increasing mutations across the SARS-CoV-2 genome have severely impacted the development of effective therapeutics and vaccines to combat the virus. However, the new SARS-CoV-2 variants and their evolutionary characteristics are not fully understood. Host cellular components such as the ACE2 receptor, RNA-binding proteins (RBPs), microRNAs, small nuclear RNA (snRNA), 18s rRNA, and the 7SL RNA component of the signal recognition particle (SRP) interact with various structural and non-structural proteins of the SARS-CoV-2. Several of these viral proteins are currently being examined for designing antiviral therapeutics. In this review, we discuss current advances in our understanding of various host cellular components targeted by the virus during SARS-CoV-2 infection. We also summarize the mutations across the SARS-CoV-2 genome that directs the evolution of new viral strains. Considering coronaviruses are rapidly evolving in humans, this enables them to escape therapeutic therapies and vaccine-induced immunity. In order to understand the virus’s evolution, it is essential to study its mutational patterns and their impact on host cellular machinery. Finally, we present a comprehensive survey of currently available databases and tools to study viral–host interactions that stand as crucial resources for developing novel therapeutic strategies for combating SARS-CoV-2 infection.

Structural insights into how Prp5 proofreads the pre-mRNA branch site

During the splicing of introns from precursor messenger RNAs (pre-mRNAs), the U2 small nuclear ribonucleoprotein (snRNP) must undergo stable integration into the spliceosomal A complex—a poorly understood, multistep process that is facilitated by the DEAD-box helicase Prp5 (refs. 1–4). During this process, the U2 small nuclear RNA (snRNA) forms an RNA duplex with the pre-mRNA branch site (the U2–BS helix), which is proofread by Prp5 at this stage through an unclear mechanism5. Here, by deleting the branch-site adenosine (BS-A) or mutating the branch-site sequence of an actin pre-mRNA, we stall the assembly of spliceosomes in extracts from the yeast Saccharomyces cerevisiae directly before the A complex is formed. We then determine the three-dimensional structure of this newly identified assembly intermediate by cryo-electron microscopy. Our structure indicates that the U2–BS helix has formed in this pre-A complex, but is not yet clamped by the HEAT domain of the Hsh155 protein (Hsh155HEAT), which exhibits an open conformation. The structure further reveals a large-scale remodelling/repositioning of the U1 and U2 snRNPs during the formation of the A complex that is required to allow subsequent binding of the U4/U6.U5 tri-snRNP, but that this repositioning is blocked in the pre-A complex by the presence of Prp5. Our data suggest that binding of Hsh155HEAT to the bulged BS-A of the U2–BS helix triggers closure of Hsh155HEAT, which in turn destabilizes Prp5 binding. Thus, Prp5 proofreads the branch site indirectly, hindering spliceosome assembly if branch-site mutations prevent the remodelling of Hsh155HEAT. Our data provide structural insights into how a spliceosomal helicase enhances the fidelity of pre-mRNA splicing.

Differential expression of LSM4 homolog, U6 small nuclear RNA and mRNA degradation associated in human epithelial ovarian cancer.

Epithelial ovarian cancer (EOC) is the most lethal gynecologic cancer (1). We performed discovery of genes associated with epithelial ovarian cancer and of the high-grade serous ovarian cancer (HGSC) subtype, using published microarray data (2, 3) to compare global gene expression profiles of normal ovary or fallopian tube with that of primary tumors from women diagnosed with epithelial ovarian cancer or HGSC. We identified the gene encoding LSM4 homolog, U6 small nuclear RNA and mRNA degradation associated, LSM4, as among the genes whose expression was most different in epithelial ovarian cancer as compared to the normal fallopian tube. LSM4 expression was significantly higher in high-grade serous ovarian tumors relative to normal fallopian tube. LSM4 expression correlated with overall survival in patients with ovarian cancer. These data indicate that expression of LSM4 is perturbed in epithelial ovarian cancers broadly and in ovarian cancers of the HGSC subtype. LSM4 may be relevant to pathways underlying ovarian cancer initiation (transformation) or progression.

TBP and SNAP50 transcription factors bind specifically to the Pr77 promoter sequence from trypanosomatid non-LTR retrotransposons

Background Trypanosomatid genomes are colonized by active and inactive mobile DNA elements, such as LINE, SINE-like, SIDER and DIRE retrotransposons. These elements all share a 77-nucleotide-long sequence at their 5′ ends, known as Pr77, which activates transcription, thereby generating abundant unspliced and translatable transcripts. However, transcription factors that mediates this process have still not been reported. Methods TATA-binding protein (TBP) and small nuclear RNA-activating protein 50 kDa (SNAP50) recombinant proteins and specific antibodies raised against them were generated. Protein capture assay, electrophoretic mobility-shift assays (EMSA) and EMSA competition assays carried out using these proteins and nuclear proteins of the parasite together to specific DNA sequences used as probes allowed detecting direct interaction of these transcription factors to Pr77 sequence. Results This study identified TBP and SNAP50 as part of the DNA-protein complex formed by the Pr77 promoter sequence and nuclear proteins of Trypanosoma cruzi. TBP establishes direct and specific contact with the Pr77 sequence, where the DPE and DPE downstream regions are docking sites with preferential binding. TBP binds cooperatively (Hill coefficient = 1.67) to Pr77 and to both strands of the Pr77 sequence, while the conformation of this highly structured sequence is not involved in TBP binding. Direct binding of SNAP50 to the Pr77 sequence is weak and may be mediated by protein–protein interactions through other trypanosomatid nuclear proteins. Conclusions Identification of the transcription factors that mediate Pr77 transcription may help to elucidate how these retrotransposons are mobilized within the trypanosomatid genomes and their roles in gene regulation processes in this human parasite. Graphic abstract

Cytogenetic markers using single-sequence probes reveal chromosomal locations of tandemly repetitive genes in scleractinian coral Acropora pruinosa

The short and similar sized chromosomes of Acropora pose a challenge for karyotyping. Conventional methods, such as staining of heterochromatic regions, provide unclear banding patterns that hamper identification of such chromosomes. In this study, we used short single-sequence probes from tandemly repetitive 5S ribosomal RNA (rRNA) and core histone coding sequences to identify specific chromosomes of Acropora pruinosa. Both the probes produced intense signals in fluorescence in situ hybridization, which distinguished chromosome pairs. The locus of the 5S rDNA probe was on chromosome 5, whereas that of core histone probe was on chromosome 8. The sequence of the 5S rDNA probe was composed largely of U1 and U2 spliceosomal small nuclear RNA (snRNA) genes and their interspacers, flanked by short sequences of the 5S rDNA. This is the first report of a tandemly repetitive linkage of snRNA and 5S rDNA sequences in Cnidaria. Based on the constructed tentative karyogram and whole genome hybridization, the longest chromosome pair (chromosome 1) was heteromorphic. The probes also hybridized effectively with chromosomes of other Acropora species and population, revealing an additional core histone gene locus. We demonstrated the applicability of short-sequence probes as chromosomal markers with potential for use across populations and species of Acropora.