The forskolin-induced swelling assay (FIS) in patient-derived intestinal organoids (PDIOs), used to determine in vitro responsiveness to elexacaftor/tezacaftor/ivacaftor (ETI), showed variability in swelling among PDIOs obtained from people with CF (pwCF) carrying the same F508del/F508del CFTR genotype. We aimed to characterise the effect of ETI on the transcriptional activity of PDIOs-derived cells to understand the intracellular processes triggered by ETI and the differences in treatment response. Six high- and six low-responding PDIOs to ETI, derived from F508del/F508del pwCF, were incubated with or without ETI for 2 to 6 h. Gene expression was assessed using 3'-mRNA sequencing and modelled using negative binomial models. Incubation with ETI resulted in a significant upregulation of several biological processes: mostly related to chemokines and signalling, chemotaxis, and tissue development processes. No changes were observed in abundance of the CFTR transcripts or in CFTR-related gene sets and pathways. The genes and pathways associated with ETI did not overlap with those whose expression changed with time only. PDIOs with a high FIS response did not significantly differ in any interpretable gene from the FIS-low organoids. The changes in the PDIOs gene expression upon the exposure to ETI cannot explain differences in the magnitude of PDIOs FIS-measured response to ETI. In conclusion, on incubation with ETI, genes of the CFTR-related pathways do not change their transcriptional activity; instead, overexpression was observed in genes of inflammatory-like cytokine response and receptor activation pathways.
- MeSH
- aktivátory chloridových kanálů terapeutické užití farmakologie MeSH
- aminofenoly * terapeutické užití farmakologie MeSH
- benzodioxoly * terapeutické užití farmakologie MeSH
- chinolony * farmakologie terapeutické užití MeSH
- cystická fibróza * genetika farmakoterapie MeSH
- fixní kombinace léků MeSH
- indoly * farmakologie MeSH
- lidé MeSH
- organoidy * metabolismus MeSH
- protein CFTR genetika MeSH
- pyrazoly * farmakologie MeSH
- pyridiny farmakologie MeSH
- pyrrolidiny farmakologie MeSH
- pyrroly farmakologie MeSH
- stanovení celkové genové exprese metody MeSH
- střeva účinky léků MeSH
- transkriptom MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Clear-cell renal cell carcinoma (ccRCC) is a common urological malignancy with an increasing incidence. The development of molecular biomarkers that can predict the response to treatment and guide personalized therapy selection would substantially improve patient outcomes. Dysregulation of non-coding RNA (ncRNA) has been shown to have a role in the pathogenesis of ccRCC. Thus, an increasing number of studies are being carried out with a focus on the identification of ncRNA biomarkers in ccRCC tissue samples and the connection of these markers with patients' prognosis, pathological stage and grade (including metastatic potential), and therapy outcome. RNA sequencing analysis led to the identification of several ncRNA biomarkers that are dysregulated in ccRCC and might have a role in ccRCC development. These ncRNAs have the potential to be prognostic and predictive biomarkers for ccRCC, with prospective applications in personalized treatment selection. Research on ncRNA biomarkers in ccRCC is advancing, but clinical implementation remains preliminary owing to challenges in validation, standardization and reproducibility. Comprehensive studies and integration of ncRNAs into clinical trials are essential to accelerate the clinical use of these biomarkers.
- MeSH
- karcinom z renálních buněk * genetika diagnóza MeSH
- lidé MeSH
- nádorové biomarkery * genetika MeSH
- nádory ledvin * genetika diagnóza MeSH
- nekódující RNA * genetika MeSH
- prognóza MeSH
- regulace genové exprese u nádorů MeSH
- stanovení celkové genové exprese MeSH
- transkriptom * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Euglenids have long been studied due to their unique physiology and versatile metabolism, providing underpinnings for much of our understanding of photosynthesis and biochemistry, and a growing opportunity in biotechnology. Until recently there has been a lack of genetic studies due to their large and complex genomes, but recently new technologies have begun to unveil their genetic capabilities. Whilst much research has focused on the model organism Euglena gracilis, other members of the euglenids have now started to receive due attention. Currently only poor nuclear genome assemblies of E. gracilis and Rhabdomonas costata are available, but there are many more plastid genome sequences and an increasing number of transcriptomes. As more assemblies become available, there are great opportunities to understand the fundamental biology of these organisms and to exploit them for biotechnology.
Transcriptional activity and gene expression are critical for the development of mature, meiotically competent oocytes. Our study demonstrates that the absence of cyclin-dependent kinase 12 (CDK12) in oocytes leads to complete female sterility, as fully developed oocytes capable of completing meiosis I are absent from the ovaries. Mechanistically, CDK12 regulates RNA polymerase II activity in growing oocytes and ensures the maintenance of the physiological maternal transcriptome, which is essential for protein synthesis that drives further oocyte growth. Notably, CDK12-deficient growing oocytes exhibit a 71% reduction in transcriptional activity. Furthermore, impaired oocyte development disrupts folliculogenesis, leading to premature ovarian failure without terminal follicle maturation or ovulation. In conclusion, our findings identify CDK12 as a key master regulator of the oocyte transcriptional program and gene expression, indispensable for oocyte growth and female fertility. A schematic illustrating the effects of loss of CDK12 in mammalian oocytes on the regulation of transcription by polymerase II and the concomitant effects on translation. This disruption leads to an aberrant transcriptome and translatome, resulting in the absence of fully mature oocytes and ultimately female sterility.
- MeSH
- cyklin-dependentní kinasy * metabolismus genetika MeSH
- meióza genetika MeSH
- myši MeSH
- oocyty * metabolismus MeSH
- RNA-polymerasa II metabolismus MeSH
- transkriptom genetika MeSH
- ženská infertilita * genetika patologie metabolismus MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Epitranscriptomics, the study of RNA modifications such as N6-methyladenosine (m6A), provides a novel layer of gene expression regulation with implications for numerous biological processes, including cellular adaptation to hypoxia. Hypoxia-inducible factor-1 (HIF-1), a master regulator of the cellular response to low oxygen, plays a critical role in adaptive and pathological processes, including cancer, ischemic heart disease, and metabolic disorders. Recent discoveries accent the dynamic interplay between m6A modifications and HIF-1 signaling, revealing a complex bidirectional regulatory network. While the roles of other RNA modifications in HIF-1 regulation remain largely unexplored, emerging evidence suggests their potential significance. MAIN BODY: This review examines the reciprocal regulation between HIF-1 and epitranscriptomic machinery, including m6A writers, readers, and erasers. HIF-1 modulates the expression of key m6A components, while its own mRNA is regulated by m6A modifications, positioning HIF-1 as both a regulator and a target in this system. This interaction enhances our understanding of cellular hypoxic responses and opens avenues for clinical applications in treating conditions like cancer and ischemic heart disease. Promising progress has been made in developing selective inhibitors targeting the m6A-HIF-1 regulatory axis. However, challenges such as off-target effects and the complexity of RNA modification dynamics remain significant barriers to clinical translation. CONCLUSION: The intricate interplay between m6A and HIF-1 highlights the critical role of epitranscriptomics in hypoxia-driven processes. Further research into these regulatory networks could drive therapeutic innovation in cancer, ischemic heart disease, and other hypoxia-related conditions. Overcoming challenges in specificity and off-target effects will be essential for realizing the potential of these emerging therapies.
- MeSH
- adenosin analogy a deriváty metabolismus MeSH
- epigeneze genetická * MeSH
- faktor 1 indukovatelný hypoxií * metabolismus genetika MeSH
- lidé MeSH
- posttranskripční úpravy RNA MeSH
- regulace genové exprese MeSH
- signální transdukce MeSH
- transkriptom MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Východiska: Signální dráha UPR (unfolded protein response, tj. odpověď na chybně složené proteiny) pomáhá myelomovým buňkám vyrovnat se se stresovými podmínkami vzniklými v důsledku nadměrné proteosyntézy, a představuje tak pro myelomové buňky prostředek umožňující jejich přežití. Extramedulární onemocnění je agresivnější forma mnohočetného myelomu, při které myelomové buňky ztrácí svoji závislost na mikroprostředí kostní dřeně a mohou infiltrovat jiné tkáně a orgány. Patogeneze vzniku extramedulárního onemocnění není dosud zcela objasněna. Cílem této studie bylo zjistit, zda existuje rozdíl v expresi genů spjatých s UPR mezi plazmatickými buňkami kostní dřeně od pacientů s mnohočetným myelomem a extramedulárním onemocněním. Materiál a metody: Pomocí reverzní transkripce ve spojení s kvantitativní polymerázovou řetězovou reakcí byla analyzována exprese šesti genů spjatých s UPR (ERN1, DDIT3, EIF2AK3, TUSC3, XBP1, HSPA5). Použito bylo celkem 76 vzorků plazmatických buněk kostní dřeně, z toho 44 bylo od pacientů s mnohočetným myelomem a 32 od pacientů s extramedulárním onemocněním. Výsledky: Byl pozorován statisticky významný rozdíl v expresi genů HSPA5, DDIT3, EIF2AK3 a ERN1 mezi skupinou mnohočetného myelomu a extramedulárního onemocnění; exprese byla ve všech případech vyšší u vzorků od pacientů s extramedulárním onemocněním. V případě genů XBP1 a TUSC3 nebyl pozorován statisticky významný rozdíl. Prokázáno bylo také několik statisticky významných korelací mezi hladinou exprese analyzovaných genů a klinickými daty pacientů. Závěr: Výsledky poukazují na možný význam signální dráhy UPR v patogenezi extramedulárního onemocnění. UPR se jeví jako vhodný směr dalšího výzkumu.
Background: The unfolded protein response (UPR) enables myeloma cells to overcome the stress conditions arising from excessive proteosynthesis and thus provides a survival advantage for myeloma cells. Extramedullary disease is a more aggressive form of multiple myeloma in which myeloma cells lose their dependence on the bone marrow microenvironment and are able to infiltrate other tissues and organs. The pathogenesis of extramedullary disease is not fully elucidated yet. The aim of this study was to determine whether there is a difference in the expression of UPR-related genes between bone marrow plasma cells from multiple myeloma and extramedullary disease patients. Materials and methods: Gene expression of six genes involved in UPR (ERN1, DDIT3, EIF2AK3, TUSC3, XBP1, HSPA5) was analyzed by quantitative reverse transcription polymerase chain reaction. In total, 76 bone marrow plasma cell samples were used, of which 44 were from patients with multiple myeloma and 32 from patients with extramedullary disease. Results: A statistically significant difference was observed between the multiple myeloma and extramedullary disease groups regarding the expression of HSPA5, DDIT3, EIF2AK3, and ERN1 genes. However, in the case of XBP1 and TUSC3 genes, no statistically significant difference in the expression was found. Several statistically significant correlations between the expression levels of the analyzed genes and the clinical data of the patients were observed as well. Conclusion: Our results suggest the importance of UPR in the pathogenesis of extramedullary disease. UPR appears to be a promising avenue for further research.
Background and Objectives: Liver cirrhosis is a serious and chronic condition that affects the liver, causing irreversible damage and scarring. The present study is designed to find out a possible correlation of hepatitis B virus (HBV) with cirrhosis by microRNA.Methods: The fold expression of the identified microRNAs by RT qPCR was determined to estimate the concentrations of circulating microRNAs in all samples. This study's main objective was to examine microRNA21-5p expression in liver cirrhosis patients. To extract RNA from samples of whole blood from all specimens in EDTA tubes, the study entailed collecting 60 specimens from the perspective of patients and samples pooled from 60 healthy participants (control group). Data on the patient are gathered for the study.Result: Researchers compared the levels of miRNA-146a in individuals with hepatic cirrhosis and control subjects, and the findings were clear. individuals with hepatic cirrhoses and controls, correspondingly, had mean levels of miRNA-146a of 2.38 3.25 and 1.12 1.01; this alteration was statistically noteworthy (P = 0.002). Both the miRNA-146a cut-off value and the prediction of liver cirrhosis disease should be considered diagnostic or adjuvant diagnostic tests. With sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and area under the curve (AUC) of 58.3%, 53.3%, 55.6%, 56.1%, and 0.610 (0.508 -0.711), the miRNA-146a cut-off value was > 0.91-fold. The current findings suggest that miRNA-146a is a subpar diagnostic marker.Conclusion: Compared to controls, patients with cirrhosis had significantly higher levels of micro-RNA146A. When compared to those who are healthy, this finding demonstrates that micro-RNA 146A may influence the prognosis of cirrhosis.
- MeSH
- biochemická analýza krve metody MeSH
- exprese genu MeSH
- hepatitida B diagnóza genetika MeSH
- jaterní cirhóza * diagnóza genetika MeSH
- klinická studie jako téma metody MeSH
- lidé MeSH
- mikro RNA * genetika MeSH
- polymerázová řetězová reakce s reverzní transkripcí metody MeSH
- senzitivita a specificita MeSH
- Check Tag
- lidé MeSH
Dual reporters encoding two distinct proteins within the same mRNA have had a crucial role in identifying and characterizing unconventional mechanisms of eukaryotic translation. These mechanisms include initiation via internal ribosomal entry sites (IRESs), ribosomal frameshifting, stop codon readthrough and reinitiation. This design enables the expression of one reporter to be influenced by the specific mechanism under investigation, while the other reporter serves as an internal control. However, challenges arise when intervening test sequences are placed between these two reporters. Such sequences can inadvertently impact the expression or function of either reporter, independent of translation-related changes, potentially biasing the results. These effects may occur due to cryptic regulatory elements inducing or affecting transcription initiation, splicing, polyadenylation and antisense transcription as well as unpredictable effects of the translated test sequences on the stability and activity of the reporters. Unfortunately, these unintended effects may lead to misinterpretation of data and the publication of incorrect conclusions in the scientific literature. To address this issue and to assist the scientific community in accurately interpreting dual-reporter experiments, we have developed comprehensive guidelines. These guidelines cover experimental design, interpretation and the minimal requirements for reporting results. They are designed to aid researchers conducting these experiments as well as reviewers, editors and other investigators who seek to evaluate published data.
- MeSH
- Eukaryota genetika MeSH
- lidé MeSH
- messenger RNA genetika metabolismus MeSH
- proteosyntéza genetika MeSH
- reportérové geny * MeSH
- směrnice jako téma MeSH
- výzkumný projekt normy MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
The molecular basis of increased hemoglobin in Andean Aymara highlanders is unknown. We conducted an integrative analysis of whole-genome-sequencing and granulocytes transcriptomics from Aymara and Europeans in Bolivia to explore genetic basis of the Aymara high hemoglobin. Differentially expressed and spliced genes in Aymaras were associated with inflammatory and hypoxia-related pathways. We identified transcripts with 4th or 5th exon skipping of NFKB1 (AS-NFKB1), key part of NF-kB complex, and their splicing quantitative trait loci; these were increased in Aymaras. AS-NFKB1 transcripts correlated with both transcripts and protein levels of inflammatory and HIF-regulated genes, including hemoglobin. While overexpression of the AS-NFKB1 variant led to increased expression of inflammatory and HIF-targeted genes; under inflammatory stress, NF-kB protein translocation to the nucleus was attenuated, resulting in reduced expression of these genes. Our study reveals AS-NFKB1 splicing events correlating with increased hemoglobin in Aymara and their possible protective mechanisms against excessive inflammation.
- MeSH
- alternativní sestřih * genetika MeSH
- dospělí MeSH
- exony genetika MeSH
- faktor 1 indukovatelný hypoxií - podjednotka alfa genetika metabolismus MeSH
- granulocyty metabolismus MeSH
- hemoglobiny * metabolismus genetika MeSH
- lidé MeSH
- lokus kvantitativního znaku MeSH
- NF-kappa B - podjednotka p50 * metabolismus genetika MeSH
- regulace genové exprese MeSH
- transkriptom MeSH
- zánět * genetika metabolismus MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Bolívie MeSH
The RNA chaperone Hfq plays crucial roles in bacterial gene expression and is a major facilitator of small regulatory RNA (sRNA) action. The toroidal architecture of the Hfq hexamer presents three well-characterized surfaces that allow it to bind sRNAs to stabilize them and engage target transcripts. Hfq-interacting sRNAs are categorized into two classes based on the surfaces they use to bind Hfq. By characterizing a systematic alanine mutant library of Hfq to identify amino acid residues that impact survival of Escherichia coli experiencing nitrogen (N) starvation, we corroborated the important role of the three RNA-binding surfaces for Hfq function. We uncovered two, previously uncharacterized, conserved residues, V22 and G34, in the hydrophobic core of Hfq, to have a profound impact on Hfq's RNA-binding activity in vivo. Transcriptome-scale analysis revealed that V22A and G34A Hfq mutants cause widespread destabilization of both sRNA classes, to the same extent as seen in bacteria devoid of Hfq. However, the alanine substitutions at these residues resulted in only modest alteration in stability and structure of Hfq. We propose that V22 and G34 have impact on Hfq function, especially critical under cellular conditions when there is an increased demand for Hfq, such as N starvation.
- MeSH
- bakteriální RNA * metabolismus genetika chemie MeSH
- dusík metabolismus MeSH
- Escherichia coli * genetika metabolismus MeSH
- hydrofobní a hydrofilní interakce * MeSH
- konzervovaná sekvence MeSH
- malá nekódující RNA * metabolismus genetika chemie MeSH
- mutace MeSH
- protein hostitelského faktoru 1 * metabolismus genetika chemie MeSH
- proteiny z Escherichia coli * metabolismus genetika chemie MeSH
- regulace genové exprese u bakterií MeSH
- stabilita RNA * genetika MeSH
- stanovení celkové genové exprese MeSH
- transkriptom genetika MeSH
- vazba proteinů MeSH
- Publikační typ
- časopisecké články MeSH
