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BioRxiv : the Preprint Server For... Jun 2024Vault RNAs (vRNAs) are evolutionarily conserved small non-coding RNAs transcribed by RNA polymerase lll. Initially described as components of the vault particle, they...
Vault RNAs (vRNAs) are evolutionarily conserved small non-coding RNAs transcribed by RNA polymerase lll. Initially described as components of the vault particle, they have since also been described as noncanonical miRNA precursors and as riboregulators of autophagy. As central molecules in these processes, vRNAs have been attributed numerous biological roles including regulation of cell proliferation and survival, response to viral infections, drug resistance, and animal development. Yet, their impact to mammalian physiology remains largely unexplored. To study vault RNAs , we generated a mouse line with a conditional loss of function allele. Because is the sole murine vRNA, this allele enables the characterization of the physiological requirements of this conserved class of small regulatory RNAs in mammals. Using this strain, we show that mice constitutively null for are viable and histologically normal but have a slight reduction in platelet counts pointing to a potential role for vRNAs in hematopoiesis. This work paves the way for further characterizations of this abundant but mysterious RNA molecule. Specifically, it enables the study of the biological consequences of constitutive or lineage-specific deletion and of the physiological requirements for an intact during normal hematopoiesis or in response to cellular stresses such as oncogene expression, viral infection, or drug treatment.
PubMed: 38895289
DOI: 10.1101/2024.06.01.596958 -
International Journal of Molecular... May 2024MicroRNAs (miRNAs) regulate approximately one-third of all human genes. The dysregulation of miRNAs has been implicated in the development of numerous human diseases,...
MicroRNAs (miRNAs) regulate approximately one-third of all human genes. The dysregulation of miRNAs has been implicated in the development of numerous human diseases, including cancers. In our investigation focusing on altering specific miRNA expression in human pancreatic cancer cells, we encountered an interesting finding. While two expression vector designs effectively enhanced miR-708 levels, they were unable to elevate mature forms of miR-29b, -1290, -2467, and -6831 in pancreatic cancer cell lines. This finding was also observed in a panel of other non-pancreatic cancer cell lines, suggesting that miRNA processing efficiency was cell line specific. Using a step-by-step approach in each step of miRNA processing, we ruled out alternative strand selection by the RISC complex and transcriptional interference at the primary miRNA (pri-miRNA) level. DROSHA processing and pri-miRNA export from the nucleus also appeared to be occurring normally. We observed precursor (pre-miRNA) accumulation only in cell lines where mature miRNA expression was not achieved, suggesting that the block was occurring at the pre-miRNA stage. To further confirm this, synthetic pre-miRNA mimics that bypass DICER processing were processed into mature miRNAs in all cases. This study has demonstrated the distinct behaviours of different miRNAs with the same vector in the same cell line, the same miRNA between the two vector designs, and with the same miRNA across different cell lines. We identified a stable vector pre-miRNA processing block. Our findings on the structural and sequence differences between successful and non-successful vector designs could help to inform future chimeric miRNA design strategies and act as a guide to other researchers on the intricate processing dynamics that can impact vector efficiency. Our research confirms the potential of miRNA mimics to surmount some of these complexities.
Topics: MicroRNAs; Humans; Pancreatic Neoplasms; RNA Processing, Post-Transcriptional; Cell Line, Tumor; Ribonuclease III; Gene Expression Regulation, Neoplastic; Transfection; RNA Precursors; Animals
PubMed: 38891854
DOI: 10.3390/ijms25115666 -
CNS Neuroscience & Therapeutics Jun 2024Islet cell autoantigen 1 (ICA1) is involved in autoimmune diseases and may affect synaptic plasticity as a neurotransmitter. Databases related to Alzheimer's disease...
AIMS
Islet cell autoantigen 1 (ICA1) is involved in autoimmune diseases and may affect synaptic plasticity as a neurotransmitter. Databases related to Alzheimer's disease (AD) have shown decreased ICA1 expression in patients with AD. However, the role of ICA1 in AD remains unclear. Here, we report that ICA1 expression is decreased in the brains of patients with AD and an AD mouse model.
RESULTS
The ICA1 increased the expression of amyloid precursor protein (APP), disintegrin and metalloprotease 10 (ADAM10), and disintegrin and metalloprotease 17 (ADAM17), but did not affect protein half-life or mRNA levels. Transcriptome sequencing analysis showed that ICA1 regulates the G protein-coupled receptor signaling pathway. The overexpression of ICA1 increased PKCα protein levels and phosphorylation.
CONCLUSION
Our results demonstrated that ICA1 shifts APP processing to non-amyloid pathways by regulating the PICK1-PKCα signaling pathway. Thus, this study suggests that ICA1 is a novel target for the treatment of AD.
Topics: Amyloid beta-Protein Precursor; Animals; Protein Kinase C-alpha; Signal Transduction; Humans; Alzheimer Disease; Mice; Carrier Proteins; Nuclear Proteins; Male; Mice, Transgenic; Female; Mice, Inbred C57BL; Amyloid Precursor Protein Secretases; Brain; Cell Cycle Proteins
PubMed: 38884369
DOI: 10.1111/cns.14754 -
BMC Cancer Jun 2024Checkpoint inhibitors (CPIs) are widely used in cancer treatment, with transformative impacts on survival. They nonetheless carry a significant risk of toxicity in the... (Observational Study)
Observational Study
BACKGROUND
Checkpoint inhibitors (CPIs) are widely used in cancer treatment, with transformative impacts on survival. They nonetheless carry a significant risk of toxicity in the form of immune-related adverse events (IrAEs), which may be sustained and life-altering. IrAEs may require high-dose and/or prolonged steroid use and represent a significant healthcare burden. They mimic immune-mediated inflammatory diseases (IMIDs) but understanding of their pathogenesis is limited. The MEDALLION project aims to determine targetable mechanisms of immune dysregulation in IrAE development, employing an immune monitoring approach to determine changes in circulating and tissue resident cells of CPI recipients who do/do not develop them and assessing the contribution of the microbiome in parallel.
METHODS
MEDALLION is a non-randomised longitudinal cohort study aiming to recruit 66 cancer patient recipients of anti-PD1/PD-L1, anti-CTLA-4 or combination therapy. Eligible participants include those with malignant melanoma in the adjuvant or metastatic setting, mesothelioma and non-small cell lung carcinoma (NSCLC) treated in the metastatic setting. Comprehensive clinical evaluation is carried out alongside blood, skin swab and stool sampling at the time of CPI initiation (baseline) and during subsequent routine hospital visits on 6 occasions over a 10-month follow-up period. It is conservatively anticipated that one third of enrolled patients will experience a "significant IrAE" (SirAE), defined according to pre-determined criteria specific to the affected tissue/organ system. Those developing such toxicity may optionally undergo a biopsy of affected tissue where appropriate, otherwise being managed according to standard of care. Peripheral blood mononuclear cells will be analysed using multi-parameter flow cytometry to investigate immune subsets, their activation status and cytokine profiles. Stool samples and skin swabs will undergo DNA extraction for 16 S ribosomal RNA (rRNA) sequencing and internal transcribed spacer (ITS) gene sequencing to determine bacterial and fungal microbiome diversity, respectively, including species associated with toxicity. Stored tissue biopsies will be available for in situ and single-cell transcriptomic evaluation. Analysis will focus on the identification of biological predictors and precursors of SirAEs.
DISCUSSION
The pathogenesis of IrAEs will be assessed through the MEDALLION cohort, with the potential to develop tools for their prediction and/or strategies for targeted prevention or treatment.
TRIAL REGISTRATION
The study was registered on 18/09/2023 in the ISRCTN registry (43,419,676).
Topics: Humans; Immune Checkpoint Inhibitors; Neoplasms; Longitudinal Studies; Immunotherapy; Cohort Studies; Monitoring, Immunologic; Melanoma
PubMed: 38877461
DOI: 10.1186/s12885-024-12468-3 -
Genome Biology and Evolution Jun 2024In flowering plants, euchromatic transposons are transcriptionally silenced by RNA-directed DNA Methylation, a small RNA-guided de novo methylation pathway. RNA-directed...
In flowering plants, euchromatic transposons are transcriptionally silenced by RNA-directed DNA Methylation, a small RNA-guided de novo methylation pathway. RNA-directed DNA Methylation requires the activity of the RNA Polymerases IV and V, which produce small RNA precursors and noncoding targets of small RNAs, respectively. These polymerases are distinguished from Polymerase II by multiple plant-specific paralogous subunits. Most RNA-directed DNA Methylation components are present in all land plants, and some have been found in the charophytic green algae, a paraphyletic group that is sister to land plants. However, the evolutionary origin of key RNA-directed DNA Methylation components, including the two largest subunits of Polymerase IV and Polymerase V, remains unclear. Here, we show that multiple lineages of charophytic green algae encode a single-copy precursor of the largest subunits of Polymerase IV and Polymerase V, resolving the two presumed duplications in this gene family. We further demonstrate the presence of a Polymerase V-like C-terminal domain, suggesting that the earliest form of RNA-directed DNA Methylation utilized a single Polymerase V-like polymerase. Finally, we reveal that charophytic green algae encode a single CLSY/DRD1-type chromatin remodeling protein, further supporting the presence of a single specialized polymerase in charophytic green algae.
Topics: DNA-Directed RNA Polymerases; Evolution, Molecular; DNA Methylation; Phylogeny; Charophyceae; Plant Proteins; Chlorophyta; Protein Subunits
PubMed: 38874416
DOI: 10.1093/gbe/evae119 -
Mathematical Biosciences and... Mar 2024B-cell acute lymphoblastic leukemia (B-ALL) is a malignant blood disorder, particularly detrimental to children and adolescents, with recurrent or unresponsive cases...
B-cell acute lymphoblastic leukemia (B-ALL) is a malignant blood disorder, particularly detrimental to children and adolescents, with recurrent or unresponsive cases contributing significantly to cancer-associated fatalities. IKBKE, associated with innate immunity, tumor promotion, and drug resistance, remains poorly understood in the context of B-ALL. Thus, this research aimed to explore the impact of the IKBKE inhibitor MCCK1 on B-ALL cells. The study encompassed diverse experiments, including clinical samples, in vitro and in vivo investigations. Quantitative real-time fluorescence PCR and protein blotting revealed heightened IKBKE mRNA and protein expression in B-ALL patients. Subsequent in vitro experiments with B-ALL cell lines demonstrated that MCCK1 treatment resulted in reduced cell viability and survival rates, with flow cytometry indicating cell cycle arrest. In vivo experiments using B-ALL mouse tumor models substantiated MCCK1's efficacy in impeding tumor proliferation. These findings collectively suggest that IKBKE, found to be elevated in B-ALL patients, may serve as a promising drug target, with MCCK1 demonstrating potential for inducing apoptosis in B-ALL cells both in vitro and in vivo.
Topics: Animals; Humans; Mice; I-kappa B Kinase; Cell Line, Tumor; Apoptosis; Cell Proliferation; Female; Cell Survival; Male; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma; Child; Adolescent; Xenograft Model Antitumor Assays; Protein Kinase Inhibitors
PubMed: 38872531
DOI: 10.3934/mbe.2024228 -
JCI Insight Jun 2024Progressive pulmonary fibrosis (PPF), defined as the worsening of various interstitial lung diseases (ILDs), currently lacks useful biomarkers. To identify novel...
Progressive pulmonary fibrosis (PPF), defined as the worsening of various interstitial lung diseases (ILDs), currently lacks useful biomarkers. To identify novel biomarkers for early detection of patients at risk of PPF, we performed a proteomic analysis of serum extracellular vesicles (EVs). Notably, the identified candidate biomarkers were enriched for lung-derived proteins participating in fibrosis-related pathways. Among them, pulmonary surfactant-associated protein B (SFTPB) in serum EVs could predict ILD progression better than the known biomarkers, serum KL-6 and SP-D, and it was identified as an independent prognostic factor from ILD-gender-age-physiology index. Subsequently, the utility of SFTPB for predicting ILD progression was evaluated further in 2 cohorts using serum EVs and serum, respectively, suggesting that SFTPB in serum EVs but not in serum was helpful. Among SFTPB forms, pro-SFTPB levels were increased in both serum EVs and lungs of patients with PPF compared with those of the control. Consistently, in a mouse model, the levels of pro-SFTPB, primarily originating from alveolar epithelial type 2 cells, were increased similarly in serum EVs and lungs, reflecting pro-fibrotic changes in the lungs, as supported by single-cell RNA sequencing. SFTPB, especially its pro-form, in serum EVs could serve as a biomarker for predicting ILD progression.
Topics: Extracellular Vesicles; Humans; Animals; Biomarkers; Mice; Male; Female; Pulmonary Fibrosis; Disease Progression; Pulmonary Surfactant-Associated Protein B; Middle Aged; Aged; Lung Diseases, Interstitial; Lung; Proteomics; Disease Models, Animal; Prognosis; Protein Precursors; Pulmonary Surfactant-Associated Proteins
PubMed: 38855869
DOI: 10.1172/jci.insight.177937 -
Journal of Autoimmunity Jun 2024In inflammatory diseases such as rheumatoid arthritis (RA), steroid metabolism is a central component mediating the actions of immuno-modulatory glucocorticoids and sex...
RATIONALE
In inflammatory diseases such as rheumatoid arthritis (RA), steroid metabolism is a central component mediating the actions of immuno-modulatory glucocorticoids and sex steroids. However, the regulation and function of cellular steroid metabolism within key leukocyte populations such as macrophages remain poorly defined. In this study, the inflammatory regulation of global steroid metabolism was assessed in RA macrophages.
METHODS
Bulk RNA-seq data from RA synovial macrophages was used to assess transcripts encoding key enzymes in steroid metabolism and signalling. Changes in metabolism were assessed in synovial fluids, correlated to measures of disease activity and functionally validated in primary macrophage cultures.
RESULTS
RNA-seq revealed a unique pattern of differentially expressed genes, including changes in genes encoding the enzymes 11β-HSD1, SRD5A1, AKR1C2 and AKR1C3. These correlated with disease activity, favouring increased glucocorticoid and androgen levels. Synovial fluid 11β-HSD1 activity correlated with local inflammatory mediators (TNFα, IL-6, IL-17), whilst 11β-HSD1, SRD5A1 and AKR1C3 activity correlated with systemic measures of disease and patient pain (ESR, DAS28 ESR, global disease activity). Changes in enzyme activity were evident in inflammatory activated macrophages in vitro and revealed a novel androgen activating role for 11β-HSD1. Together, increased glucocorticoids and androgens were able to suppress inflammation in macrophages and fibroblast-like-synoviocytes.
CONCLUSIONS
This study underscores the significant increase in androgen and glucocorticoid activation within inflammatory polarized macrophages of the synovium, contributing to local suppression of inflammation. The diminished profile of inactive steroid precursors in postmenopausal women may contribute to disturbances in this process, leading to increased disease incidence and severity.
PubMed: 38851089
DOI: 10.1016/j.jaut.2024.103263 -
Veterinary Journal (London, England :... Jun 2024Irisin is a 112-amino acid peptide hormone that is cleaved from fibronectin type III domain-containing protein 5 (FNDC5), a type I transmembrane protein abundantly found...
Irisin is a 112-amino acid peptide hormone that is cleaved from fibronectin type III domain-containing protein 5 (FNDC5), a type I transmembrane protein abundantly found in muscle tissue. Irisin is a putative mediator of the benefits of exercise, neuroprotection, bone growth, and cardiac health. However, few studies have focused on irisin in domestic animals. Further, whether processed irisin is detectable in domestic animal tissues remains uncertain. To address this, we determined FNDC5 mRNA and protein concentration in anatine (duck) and porcine (pig) skeletal muscle, and in equine (horse), swine, and anatine serum samples. RT-PCR analysis identified FNDC5 mRNA in all pig and duck skeletal muscle samples. An approximately 25 kDa band representing FNDC5 was detected in both pig and duck skeletal muscle. Fluorescence immunohistochemistry using a rabbit monoclonal FNDC5/irisin primary antibody and a goat polyclonal anti-rabbit secondary antibody localized FNDC5/irisin-like immunoreactivity in both the glandular and muscular regions of pig stomach. FNDC5/irisin-like immunoreactivity was also identified in horse, pig, and duck serum using a multispecies irisin ELISA. The average values of irisin-like immunoreactivity were 13.7 (duck), 15.4 (horse), and 7.0 (pig) ng/mL in samples tested. Our results support the presence of irisin precursor in several domestic animals. Processed irisin, however, was not detectable. Further studies are required to validate reliable tools to detect and quantify processed irisin in domestic animals.
PubMed: 38849027
DOI: 10.1016/j.tvjl.2024.106161 -
Bioactive Materials Sep 2024Osteoporosis is majorly caused by an imbalance between osteoclastic and osteogenic niches. Despite the development of nationally recognized first-line anti-osteoporosis...
Osteoporosis is majorly caused by an imbalance between osteoclastic and osteogenic niches. Despite the development of nationally recognized first-line anti-osteoporosis drugs, including alendronate (AL), their low bioavailability, poor uptake rate, and dose-related side effects present significant challenges in treatment. This calls for an urgent need for more effective bone-affinity drug delivery systems. In this study, we produced hybrid structures with bioactive components and stable fluffy topological morphology by cross-linking calcium and phosphorus precursors based on mesoporous silica to fabricate nanoadjuvants for AL delivery. The subsequent grafting of -PEG-DAsp ensured superior biocompatibility and bone targeting capacity. RNA sequencing revealed that these fluffy nanoadjuvants effectively activated adhesion pathways through CARD11 and CD34 molecular mechanisms, hence promoting cellular uptake and intracellular delivery of AL. Experiments showed that small-dose AL nanoadjuvants effectively suppress osteoclast formation and potentially promote osteogenesis. results restored the balance between osteogenic and osteoclastic niches against osteoporosis as well as the consequent significant recovery of bone mass. Therefore, this study constructed a drug nanoadjuvant with peculiar topological structures and high bone targeting capacities, efficient intracellular drug delivery as well as bone bioactivity. This provides a novel perspective on drug delivery for osteoporosis and treatment strategies for other bone diseases.
PubMed: 38846529
DOI: 10.1016/j.bioactmat.2024.05.037