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Nature Communications Jun 2024Neuronal differentiation requires building a complex intracellular architecture, and therefore the coordinated regulation of defined sets of genes. RNA-binding proteins...
Neuronal differentiation requires building a complex intracellular architecture, and therefore the coordinated regulation of defined sets of genes. RNA-binding proteins (RBPs) play a key role in this regulation. However, while their action on individual mRNAs has been explored in depth, the mechanisms used to coordinate gene expression programs shaping neuronal morphology are poorly understood. To address this, we studied how the paradigmatic RBP IMP1 (IGF2BP1), an essential developmental factor, selects and regulates its RNA targets during the human neuronal differentiation. We perform a combination of system-wide and molecular analyses, revealing that IMP1 developmentally transitions to and directly regulates the expression of mRNAs encoding essential regulators of the microtubule network, a key component of neuronal morphology. Furthermore, we show that m6A methylation drives the selection of specific IMP1 mRNA targets and their protein expression during the developmental transition from neural precursors to neurons, providing a molecular principle for the onset of target selectivity.
Topics: Humans; RNA-Binding Proteins; Microtubules; Neurons; Cell Differentiation; RNA, Messenger; Methylation; Neurogenesis; Adenosine; Gene Expression Regulation, Developmental
PubMed: 38844464
DOI: 10.1038/s41467-024-49139-7 -
Haematologica Jun 2024IKZF1-deletions occur in 10-15% of patients with B-cell precursor acute lymphoblastic leukemia (BCP-ALL) and predict a poor outcome. However, the impact of IKZF1-loss on...
IKZF1-deletions occur in 10-15% of patients with B-cell precursor acute lymphoblastic leukemia (BCP-ALL) and predict a poor outcome. However, the impact of IKZF1-loss on sensitivity to drugs used in contemporary treatment protocols has remained underexplored. Here we show in experimental models and in patients that loss of IKZF1 promotes resistance to AraC, a key component of both upfront and relapsed treatment protocols. We attribute this resistance, in part, to diminished import and incorporation of cytarabine (AraC) due to reduced expression of the solute carrier hENT1. Moreover, we find elevated mRNA expression of Evi1, a known driver of therapy resistance in myeloid malignancies. Finally, a kinase directed CRISPR/Cas9-screen identified that inhibition of either mediator kinases CDK8/19 or casein kinase 2 can restore response to AraC. We conclude that this high-risk patient group could benefit from alternative antimetabolites, or targeted therapies that resensitize the cells to AraC.
PubMed: 38841778
DOI: 10.3324/haematol.2023.284357 -
Frontiers in Cellular and Infection... 2024Trimethylamine-N-oxide (TMAO) is produced by hepatic flavin-containing monooxygenase 3 (FMO3) from trimethylamine (TMA). High TMAO level is a biomarker of cardiovascular...
BACKGROUND
Trimethylamine-N-oxide (TMAO) is produced by hepatic flavin-containing monooxygenase 3 (FMO3) from trimethylamine (TMA). High TMAO level is a biomarker of cardiovascular diseases and metabolic disorders, and it also affects periodontitis through interactions with the gastrointestinal microbiome. While recent findings indicate that periodontitis may alter systemic TMAO levels, the specific mechanisms linking these changes and particular oral pathogens require further clarification.
METHODS
In this study, we established a C57BL/6J male mouse model by orally administering (, ), (, ), (, ) and PBS was used as a control. We conducted LC-MS/MS analysis to quantify the concentrations of TMAO and its precursors in the plasma and cecal contents of mice. The diversity and composition of the gut microbiome were analyzed using 16S rRNA sequencing. TMAO-related lipid metabolism and enzymes in the intestines and liver were assessed by qPCR and ELISA methods. We further explored the effect of on FMO3 expression and lipid molecules in HepG2 cells by stimulating the cells with -LPS .
RESULTS
The three oral pathogenic bacteria were orally administered to the mice for 5 weeks. The group showed a marked increase in plasma TMAO, betaine, and creatinine levels, whereas no significant differences were observed in the gut TMAO level among the four groups. Further analysis showed similar diversity and composition in the gut microbiomes of both the and groups, which were different from the and control groups. The profiles of TMA-TMAO pathway-related genera and gut enzymes were not significantly different among all groups. The group showed significantly higher liver FMO3 levels and elevated lipid factors (IL-6, TG, TC, and NEFA) in contrast to the other groups. experiments confirmed that stimulation of HepG2 cells with -LPS upregulated the expression of FMO3 and increased the lipid factors TC, TG, and IL-6.
CONCLUSION
This study conclusively demonstrates that , compared to and , plays a critical role in elevating plasma TMAO levels and significantly influences the TMA-TMAO pathway, primarily by modulating the expression of hepatic FMO3 and directly impacting hepatic lipid metabolism.
Topics: Animals; Male; Methylamines; Humans; Gastrointestinal Microbiome; Mice; Mice, Inbred C57BL; Oxygenases; Porphyromonas gingivalis; Fusobacterium nucleatum; Metabolic Networks and Pathways; Hep G2 Cells; Lipid Metabolism; Disease Models, Animal; Periodontitis; Liver; RNA, Ribosomal, 16S; Tandem Mass Spectrometry; Mouth
PubMed: 38836053
DOI: 10.3389/fcimb.2024.1413787 -
PLoS Genetics Jun 2024Splicing is an important step of gene expression regulation in eukaryotes, as there are many mRNA precursors that can be alternatively spliced in different tissues, at...
Splicing is an important step of gene expression regulation in eukaryotes, as there are many mRNA precursors that can be alternatively spliced in different tissues, at different cell cycle phases or under different external stimuli. We have developed several integrated fluorescence-based in vivo splicing reporter constructs that allow the quantification of fission yeast splicing in vivo on intact cells, and we have compared their splicing efficiency in a wild type strain and in a prp2-1 (U2AF65) genetic background, showing a clear dependency between Prp2 and a consensus signal at 5' splicing site (5'SS). To isolate novel genes involved in regulated splicing, we have crossed the reporter showing more intron retention with the Schizosaccharomyces pombe knock out collection. Among the candidate genes involved in the regulation of splicing, we have detected strong splicing defects in two of the mutants -Δcwf12, a member of the NineTeen Complex (NTC) and Δsaf5, a methylosome subunit that acts together with the survival motor neuron (SMN) complex in small nuclear ribonucleoproteins (snRNP) biogenesis. We have identified that strains with mutations in cwf12 have inefficient splicing, mainly when the 5'SS differs from the consensus. However, although Δsaf5 cells also have some dependency on 5'SS sequence, we noticed that when one intron of a given pre-mRNA was affected, the rest of the introns of the same pre-mRNA had high probabilities of being also affected. This observation points Saf5 as a link between transcription rate and splicing.
Topics: Schizosaccharomyces; Schizosaccharomyces pombe Proteins; RNA Splicing; Transcription, Genetic; Gene Expression Regulation, Fungal; Introns; Mutation; Alternative Splicing; Ribonucleoproteins, Small Nuclear; RNA Precursors; RNA Splice Sites; Splicing Factor U2AF
PubMed: 38833506
DOI: 10.1371/journal.pgen.1011316 -
Nature Communications Jun 2024The human mitochondrial genome is transcribed into two RNAs, containing mRNAs, rRNAs and tRNAs, all dedicated to produce essential proteins of the respiratory chain. The...
The human mitochondrial genome is transcribed into two RNAs, containing mRNAs, rRNAs and tRNAs, all dedicated to produce essential proteins of the respiratory chain. The precise excision of tRNAs by the mitochondrial endoribonucleases (mt-RNase), P and Z, releases all RNA species from the two RNA transcripts. The tRNAs then undergo 3'-CCA addition. In metazoan mitochondria, RNase P is a multi-enzyme assembly that comprises the endoribonuclease PRORP and a tRNA methyltransferase subcomplex. The requirement for this tRNA methyltransferase subcomplex for mt-RNase P cleavage activity, as well as the mechanisms of pre-tRNA 3'-cleavage and 3'-CCA addition, are still poorly understood. Here, we report cryo-EM structures that visualise four steps of mitochondrial tRNA maturation: 5' and 3' tRNA-end processing, methylation and 3'-CCA addition, and explain the defined sequential order of the tRNA processing steps. The methyltransferase subcomplex recognises the pre-tRNA in a distinct mode that can support tRNA-end processing and 3'-CCA addition, likely resulting from an evolutionary adaptation of mitochondrial tRNA maturation complexes to the structurally-fragile mitochondrial tRNAs. This subcomplex can also ensure a tRNA-folding quality-control checkpoint before the sequential docking of the maturation enzymes. Altogether, our study provides detailed molecular insight into RNA-transcript processing and tRNA maturation in human mitochondria.
Topics: Humans; RNA, Transfer; Mitochondria; Ribonuclease P; tRNA Methyltransferases; RNA Processing, Post-Transcriptional; Cryoelectron Microscopy; RNA, Mitochondrial; Methylation; Nucleic Acid Conformation; Models, Molecular; RNA Precursors
PubMed: 38824131
DOI: 10.1038/s41467-024-49132-0 -
Experimental Gerontology Aug 2024Chronic stress (CS) is critically involved in the Alzheimer's disease (AD) pathogenesis resulting in cognitive disturbance. Also, amyloid precursor protein (APP) related...
Quercetin ameliorates cognitive deficit, expression of amyloid precursor gene, and pro-inflammatory cytokines in an experimental models of Alzheimer's disease in Wistar rats.
Chronic stress (CS) is critically involved in the Alzheimer's disease (AD) pathogenesis resulting in cognitive disturbance. Also, amyloid precursor protein (APP) related gens, pro-inflammatory cytokines, and stress increases AD-related pathogenesis through increasing APP, all are important players in the development of AD. Herein, we explore the possible neuroprotective and anti-amnestic effect of quercetin (QUER) on cognitive deficits induced by scopolamine (SCOP) in stressed rats. Stress induction was performed by exposed of rats to 2-h chronic restraint stress for 10 days. Then rats were supplemented with QUER (25 mg/kg/day oral gavage, for 1 month). Ratswere submitted to intraperitoneal (i.p.) injection of SCOP (1 mg/kg) during the final 9 days of QUER supplementation to induce dementia like condition. Following the interventions, behavioral tests [elevated plus maze (EPM) and novel object recognition memory (NORM)] was examined to analysis the cognitive functions. Meanwhile, prefrontal cortex (PFC) and hippocampus of brain were used for gene expression and biochemical studies. Also, the plasma corticosterone (CORT) level was measured. We established that administration of QUER ameliorated the SCOP-related memory impairment. Also, QUER decreased stress related anxiety like behaviors in the EPM. QUER also altered the interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) levels in both PFC and hippocampus of SCOP treated rats in stress and non-stress conditions. We found that QUER increased APP and amyloid precursor-like protein 2 (APLP2) mRNA expression in both non-stress and stressed rats. Also, our findings imply that QUER suppress the effect of SCOP on cognitive functions. Moreover, decreased APP mRNA expression in the hippocampus were observed following pretreatment of rats with QUER in both stress and non-stress groups. Given that decreased amyloid beta (Aβ) expression in the hippocampus of stressed rats, it can be proposed that elevations in APP mRNA expression by QUER activates non-amyloidogenic pathways leading to reduction in Aβ levels. However, our findings indicate that QUER can be a therapeutic candidate, which exerts an anti-amnesic property against SCOP-induced memory decline. On the other hand, prior QUER administration in stress condition could be a promising approach against AD prevention.
Topics: Animals; Quercetin; Alzheimer Disease; Rats, Wistar; Male; Disease Models, Animal; Rats; Cytokines; Hippocampus; Amyloid beta-Protein Precursor; Stress, Psychological; Cognitive Dysfunction; Scopolamine; Neuroprotective Agents; Corticosterone; Prefrontal Cortex; Cognition
PubMed: 38821324
DOI: 10.1016/j.exger.2024.112466 -
Nature Immunology Jun 2024A key barrier to the development of vaccines that induce broadly neutralizing antibodies (bnAbs) against human immunodeficiency virus (HIV) and other viruses of high...
A key barrier to the development of vaccines that induce broadly neutralizing antibodies (bnAbs) against human immunodeficiency virus (HIV) and other viruses of high antigenic diversity is the design of priming immunogens that induce rare bnAb-precursor B cells. The high neutralization breadth of the HIV bnAb 10E8 makes elicitation of 10E8-class bnAbs desirable; however, the recessed epitope within gp41 makes envelope trimers poor priming immunogens and requires that 10E8-class bnAbs possess a long heavy chain complementarity determining region 3 (HCDR3) with a specific binding motif. We developed germline-targeting epitope scaffolds with affinity for 10E8-class precursors and engineered nanoparticles for multivalent display. Scaffolds exhibited epitope structural mimicry and bound bnAb-precursor human naive B cells in ex vivo screens, protein nanoparticles induced bnAb-precursor responses in stringent mouse models and rhesus macaques, and mRNA-encoded nanoparticles triggered similar responses in mice. Thus, germline-targeting epitope scaffold nanoparticles can elicit rare bnAb-precursor B cells with predefined binding specificities and HCDR3 features.
Topics: Animals; Humans; HIV Envelope Protein gp41; HIV Antibodies; Mice; AIDS Vaccines; Macaca mulatta; Antibodies, Neutralizing; HIV-1; HIV Infections; Vaccination; Broadly Neutralizing Antibodies; B-Lymphocytes; Nanoparticles; Female; Complementarity Determining Regions; Epitopes
PubMed: 38816615
DOI: 10.1038/s41590-024-01833-w -
Nature Communications May 2024The 5'-end capping of nascent pre-mRNA represents the initial step in RNA processing, with evidence demonstrating that guanosine addition and 2'-O-ribose methylation...
The 5'-end capping of nascent pre-mRNA represents the initial step in RNA processing, with evidence demonstrating that guanosine addition and 2'-O-ribose methylation occur in tandem with early steps of transcription by RNA polymerase II, especially at the pausing stage. Here, we determine the cryo-EM structures of the paused elongation complex in complex with RNGTT, as well as the paused elongation complex in complex with RNGTT and CMTR1. Our findings show the simultaneous presence of RNGTT and the NELF complex bound to RNA polymerase II. The NELF complex exhibits two conformations, one of which shows a notable rearrangement of NELF-A/D compared to that of the paused elongation complex. Moreover, CMTR1 aligns adjacent to RNGTT on the RNA polymerase II stalk. Our structures indicate that RNGTT and CMTR1 directly bind the paused elongation complex, illuminating the mechanism by which 5'-end capping of pre-mRNA during transcriptional pausing.
Topics: Cryoelectron Microscopy; Transcription, Genetic; RNA Polymerase II; RNA Caps; RNA Precursors; Humans; Protein Binding; Models, Molecular; RNA, Messenger; Saccharomyces cerevisiae Proteins
PubMed: 38816438
DOI: 10.1038/s41467-024-48963-1 -
Non-coding RNA May 2024Small RNAS (sRNAs) participate in regulatory RNA interference (RNAi) mechanisms in a wide range of eukaryotic organisms, including fungi. The fungus , a model for the...
Small RNAS (sRNAs) participate in regulatory RNA interference (RNAi) mechanisms in a wide range of eukaryotic organisms, including fungi. The fungus , a model for the study of secondary metabolism, contains a complete set of genes for RNAi pathways. We have analyzed by high-throughput sequencing the content of sRNAs in total RNA samples of grown in synthetic medium in the dark or after 1 h of illumination, using libraries below 150 nt, covering sRNAs and their precursors. For comparison, a parallel analysis with was carried out. The sRNA reads showed a higher proportion of 5' uracil in the RNA samples of the expected sizes in both species, indicating the occurrence of genuine sRNAs, and putative miRNA-like sRNAs (milRNAS) were identified with prediction software. carries at least one transcriptionally expressed Ty1/copia-like retrotransposable element, in which sRNAs were found in both sense and antisense DNA strands, while in skippy-like elements also show sRNA formation. The finding of sRNA in these mobile elements indicates an active sRNA-based RNAi pathway. Targeted deletion of , the only Dicer gene with significant expression under the conditions tested, did not produce appreciable phenotypic or transcriptomic alterations.
PubMed: 38804363
DOI: 10.3390/ncrna10030031 -
Communications Biology May 2024Nesfatin-1 (NESF-1) has been shown to modulate lipid metabolism. We have identified a nesfatin-1-like-peptide (NLP) processed from a related precursor nucleobindin 1...
Nesfatin-1 (NESF-1) has been shown to modulate lipid metabolism. We have identified a nesfatin-1-like-peptide (NLP) processed from a related precursor nucleobindin 1 (NUCB1). Here we determined if NLP, like NESF-1, regulates lipid accumulation in vitro, and tested if the disruption of nucb1 gene affects hepatic lipid metabolism genes in mice. Hepatocytes (HepG2/C3A cells) express NLP and NESF-1 and both peptides significantly reduced lipogenic enzyme mRNAs and enhanced beta-oxidation enzyme mRNAs. Lipid contents in oleic acid induced HepG2/C3A cells were attenuated by NESF-1 and NLP. The inhibitory effect on cellular lipid content was blocked by compound C, an inhibitor of AMPK. The disruption of nucb1 gene affected lipid metabolism-related enzyme mRNAs, endogenous nucb2 mRNA and AMPK phosphorylation. The lipid-lowering effects identified here highlights the potential of nucleobindins and peptides processed from them to address lipid disorders, and its possible benefits in metabolic disease management.
Topics: Nucleobindins; Animals; Humans; Lipid Metabolism; Hepatocytes; Nerve Tissue Proteins; Mice; Calcium-Binding Proteins; DNA-Binding Proteins; Hep G2 Cells; Male; Mice, Inbred C57BL
PubMed: 38802487
DOI: 10.1038/s42003-024-06314-2