Gene Expression Research Articles

AcsS inhibits the hemolytic activity and thermostable direct hemolysin (TDH) gene expression in Vibrio parahaemolyticus

Vibrio parahaemolyticus produces a key virulent factor known as thermostable direct hemolysin (TDH). TDH exhibits diverse biological activities, including hemolytic activity. The β-type hemolysis observed on Wagatsuma agar due to TDH is recognized as the Kanagawa phenomenon (KP). The tdh2 gene is primarily responsible for TDH production and the associated KP. AcsS was originally identified as an activator of swimming and swarming motility in V. parahaemolyticus. However, the extent of its potential to regulate other cellular pathways remains unclear. In this study, we explored the regulatory effects of AcsS on the hemolytic activity and tdh2 expression in V. parahaemolyticus. The data showed that V. parahaemolyticus hemolytic activity and tdh2 transcription were under the negative control of AcsS. Additionally, in vitro binding assays revealed that His-AcsS could not bind to the regulatory DNA region of tdh2. However, overexpression of AcsS in an Escherichia coli strain suppressed the expression of tdh2. Collectively, these results suggested that AcsS suppresses the hemolytic activity of V. parahaemolyticus through the downregulation of tdh2 transcription. The data enhanced our understanding of the regulatory networks governing tdh2 expression and the roles of AcsS in this bacterium.

Synergistic Effects of Dietary Tryptophan and Dip Vaccination in the Immune Response of European Seabass Juveniles

Vaccination is an effective, cost-efficient method to preventing disease outbreaks. However, vaccine procedures can induce adverse reactions due to stress, increasing plasma cortisol in the short term. In this context, tryptophan may prove to be fundamental as it has been demonstrated to have various desirable neuroendocrine attributes in different fish species. Therefore, this study aimed to evaluate both short-term (3 days) and long-term (21 days) effects of dietary tryptophan supplementation on European seabass juveniles’ (26.23 ± 7.22 g) response to vaccination and disease resistance to Tenacibaculum maritimum. The short-term tryptophan-fed fish exhibited increased hepatic superoxide dismutase and plasma cortisol levels, along with the downregulation of immune-related genes. Despite these changes, disease resistance was unaffected. When fish were later dip vaccinated, tryptophan prevented the stress-induced plasma cortisol increase and upregulated the gene expression of igm, suggesting tryptophan’s role in enhancing vaccination efficiency by counteracting stress-associated effects. In the long term, the lowest supplementation dose counteracted vaccine-mediated reduced gene expression, and fish fed this diet showed a more modest molecular response. Overall, the findings suggest a complex interplay between tryptophan supplementation, immune responses, and vaccine efficiency in fish. Further research is necessary to clarify how tryptophan could consistently improve vaccine efficiency in aquaculture.

Abstract A055: Tumor muscle invasion promotes tumor heterogeneity and normal muscle reprogramming

Abstract Aggressive human prostate tumors traverse a contractile smooth muscle pseudo-capsule, in a process termed extracapsular extension (ECE), defining the pT3 pathologic stage associating with increased biochemical recurrence, bone metastases, and cancer-specific mortality. While T3 lesions can be detected by non-invasive mpMRI imaging, how the tumor invades into and through the contractile muscle is unknown. Using a mouse xenograft model system of ECE, we investigated the transcriptomic consequences of human tumor invasion into and through the contractile smooth muscle of the mouse diaphragm. Both human and mouse gene transcripts were documented and analyzed. Tumor cells were intraperitoneally injected into male NSG mice and 6 weeks later, three tumor compartments (pre-invasive, muscle-invasive, and super-invasive cells that had reached the superior diaphragm surface) were analyzed for differential bulk human and mouse gene expression. Whole genome transcriptomic sequencing and GO pathway analysis revealed that approximately 414 human genes were differentially expressed between the non- invasive, muscle-resident, and super-invasive tumor populations and at least 5 enriched pathways were involved. The unique expression gene patterns of muscle-resident tumor cells were reversible when compared to the pre-invasive and super-invasive expression patterns. Significant increases in g-H2AX and nuclear deformation were observed in the muscle-resident tumor clusters as compared to the non-invasive tumor mass. No differences in tumor cell proliferation, as detected by Ki67 staining, were found between the tumor clusters in the three compartments. Immunohistochemistry staining for a damage response cytokeratin (KRT6A) and integrin (CD49f) revealed heterogeneity within the muscle resident tumors as compared to the non- invasive cells. Taken together, these data suggest a hostile contractile muscle environment elicits specific responses by the invading tumor. Single cell analysis within each of the tumor compartments is currently underway to define the spatial heterogeneity of gene expression in invasive tumor cell clusters within the contractile muscle. A dynamic reciprocity of the tumor/muscle microenvironment is suggested by the analysis of the bulk transcriptomic sequencing demonstrating a significant increase in mouse muscle bio-synthetic gene transcription as a consequence of the human tumor penetrating the tissue. Taken together, these data indicate that human tumors, during the act of traversing the contractile muscle layer, respond to this unique microenvironment by transiently altering transcription, while sustaining nuclear damage which results in the reprogramming of the muscle. This new information suggests that novel tumor or muscle biomarkers might indicate early muscle invasion events and assist new high-resolution image analysis for precision diagnostic and/or therapeutic decisions. (Supported in part by P30 CA23074; F30 CA143924, UACC Team Science Award). Citation Format: Kendra D Marr, Beatrice S Knudsen, Rafael Sainz, Kelvin W Pond, Noel E Warfel, Belinda E Sun, Anne E Cress. Tumor muscle invasion promotes tumor heterogeneity and normal muscle reprogramming [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A055.

Robust reference gene selection in Norway spruce: essential for real-time quantitative PCR across different tissue, stress and developmental conditions

Accurate gene expression analysis in Norway spruce (Picea abies) under diverse stress conditions requires the identification of stable reference genes for normalization. Notably, the literature lacks reports on suitable reference genes in Norway spruce. Here, we aimed to address this gap by identifying suitable reference genes for quantitative real-time PCR in Norway spruce across various stress conditions (drought, heat, pathogen infection) in seedlings, tissues (needle, phloem, root), and developmental stages (seedlings, mature trees). We evaluated the stability of 15 candidate reference genes and assessed their expression stability using five statistical algorithms (ΔCt, geNorm, NormFinder, BestKeeper, and RefFinder). Our results highlight ubiquitin-protein ligase (SP1), conserved oligomeric Golgi complex (COG7), and tubby-like F-box protein (TULP6) as the most stable reference genes, while succinate dehydrogenase (SDH5) and heat shock protein 90 (HSP90) were the least stable under various experimental conditions. COG7 and TULP6 are novel candidate reference genes reported for the first time. The expression stability of the identified reference genes was further validated using dehydrin-like protein 5 (PaDhn5) under drought conditions in Norway spruce. Pairwise variation analysis suggests that two reference genes were sufficient to normalize gene expression across all sample sets. This study provides a comprehensive analysis of reference gene stability under different experimental conditions and a catalog of genes for each condition, facilitating future functional genomic research in Norway spruce and related conifers.

Inactivation of Pseudovirus Expressing the D614G Spike Protein Mutation using Nitric Oxide‐Plasma Activated Water

AbstractVariants of concern (VOCs) of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS‐CoV‐2) exhibit high infectivity due to mutations, particularly in the spike protein, that facilitate enhanced binding of virus to human angiotensin‐converting enzyme 2 (hACE2). The D614G mutation, situated in S1‐domain, promotes the open conformation of spike protein, augmenting its interaction with hACE2. Activated water neutralizes pathogens by damaging biological molecules; however, its effect on mutated SARS‐CoV‐2 or VOCs requires further exploration. Here, the efficacy of nitric oxide (NOx)‐plasma activated water (PAW) in inhibiting infections by SARS‐CoV‐2 pseudovirus expressing D614G‐mutated spike protein is investigated, which serves as a model for mutated SARS‐CoV‐2. Results demonstrated high prevalence of D614G mutation in SARS‐CoV‐2 and its VOCs. NOx‐PAW is non‐toxic to cells at high concentration, inhibiting infection by 71%. Moreover, NOx‐PAW induced structural changes in S1‐domain of spike protein, reducing its binding affinity and lowering clathrin‐mediated endocytosis‐related gene expression. Additionally, in silico analysis revealed NOx species in NOx‐PAW played key role in impairing S1‐domain function of the mutated SARS‐CoV‐2 pseudovirus by interacting directly with it. Collectively, these findings reveal the potent inactivation ability of PAW against mutated SARS‐CoV‐2 and suggest its potential application in combating emerging variants of SARS‐CoV‐2 and other viral threats.

Revealing the potential therapeutic mechanism of Lonicerae Japonicae Flos in Alzheimer’s disease: a computational biology approach

BackgroundAlzheimer’s disease (AD) is a degenerative brain disease without a cure. Lonicerae Japonicae Flos (LJF), a traditional Chinese herbal medicine, possesses a neuroprotective effect, but its mechanisms for AD are not well understood. This study aimed to investigate potential targets and constituents of LJF against AD.MethodsNetwork pharmacology and bioinformatics analyses were performed to screen potential compounds and targets. Gene Expression Omnibus (GEO) datasets related to AD patients were used to screen core targets of differential expression. Gene expression profiling interactive analysis (GEPIA) was used to validate the correlation between core target genes and major causative genes of AD. The receiver operating characteristic (ROC) analysis was used to evaluate the predictive efficacy of core targets based on GEO datasets. Molecular docking and dynamics simulation were conducted to analyze the binding affinities of effective compounds with core targets.ResultsNetwork pharmacology analysis showed that 112 intersection targets were identified. Bioinformatics analysis displayed that 32 putative core targets were identified from 112 intersection targets. Only eight core targets were differentially expressed based on GEO datasets. Finally, six core targets of MAPK8, CTNNB1, NFKB1, EGFR, BCL2, and NFE2L2 were related to AD progression and had good predictive ability based on correlation and ROC analyses. Molecular docking and dynamics simulation analyses elucidated that the component of lignan interacted with EGFR, the component of β-carotene interacted with CTNNB1 and BCL2, the component of β-sitosterol interacted with BCL2, the component of hederagenin interacted with NFKB1, the component of berberine interacted with EGFR and BCL2, and the component of baicalein interacted with NFKB1, EGFR and BCL2.ConclusionThrough a comprehensive analysis, this study revealed that six core targets (MAPK8, CTNNB1, NFKB1, EGFR, BCL2, and NFE2L2) and six practical components (lignan, β-carotene, β-sitosterol, hederagenin, berberine, and baicalein) were involved in the mechanism of action of LJF against AD. Our work demonstrated that LJF effectively treats AD through its multi-component and multi-target properties. The findings of this study will establish a theoretical basis for the expanded application of LJF in AD treatment and, hopefully, can guide more advanced experimental research in the future.

Transcriptome Analysis Reveals the Crucial Role of Phenylalanine Ammonia-Lyase in Low Temperature Response in Ammopiptanthus mongolicus

Background: Ammopiptanthus mongolicus is a rare temperate evergreen shrub with high tolerance to low temperature, and understanding the related gene expression regulatory network can help advance research on the mechanisms of plant tolerance to abiotic stress. Methods: Here, time-course transcriptome analysis was applied to investigate the gene expression network in A. mongolicus under low temperature stress. Results: A total of 12,606 differentially expressed genes (DEGs) were identified at four time-points during low temperature stress treatment, and multiple pathways, such as plant hormones, secondary metabolism, and cell membranes, were significantly enriched in the DEGs. Trend analysis found that the expression level of genes in cluster 19 continued to upregulate under low temperatures, and the genes in cluster 19 were significantly enriched in plant hormone signaling and secondary metabolic pathways. Based on the transcriptome data, the expression profiles of the genes in abscisic acid, salicylic acid, and flavonoid metabolic pathways were analyzed. It was found that biosynthesis of abscisic acid and flavonoids may play crucial roles in the response to low temperature stress. Furthermore, members of the phenylalanine ammonia-lyase (PAL) family in A. mongolicus were systematically identified and their structures and evolution were characterized. Analysis of cis-acting elements showed that the PAL genes in A. mongolicus were closely related to abiotic stress response. Expression pattern analysis showed that PAL genes responded to various environmental stresses, such as low temperature, supporting their involvement in the low temperature response in A. mongolicus. Conclusions: Our study provides important data for understanding the mechanisms of tolerance to low temperatures in A. mongolicus.

Plasticity and environment-specific relationships between gene expression and fitness in Saccharomyces cerevisiae

Plasticity and environment-specific relationships between gene expression and fitness in Saccharomyces cerevisiae

P53 and the E3 Ubiquitin Ligase MDM2 in Glaucomatous Lamina Cribrosa Cells

Lamina cribrosa (LC) cells play an integral role in extracellular matrix remodeling and fibrosis in human glaucoma. LC cells bear similarities to myofibroblasts that adopt an apoptotic-resistant, proliferative phenotype, a process linked to dysregulation of tumor suppressor-gene p53 pathways, including ubiquitin-proteasomal degradation via murine-double-minute-2 (MDM2). Here, we investigate p53 and MDM2 in glaucomatous LC cells. Primary human LC cells were isolated from glaucomatous donor eyes (GLC) and age-matched normal controls (NLC) (n = 3 donors/group). LC cells were cultured under standard conditions ± 48-h treatment with p53-MDM2-interaction inhibitor RG-7112. Markers of p53-MDM2, fibrosis, and apoptosis were analyzed by real-time polymerase chain reaction (qRT-PCR), western blotting, and immunofluorescence. Cellular proliferation and viability were assessed using colorimetric methyl-thiazolyl-tetrazolium salt assays (MTS/MTT). In GLC versus NLC cells, protein expression of p53 was significantly decreased (p < 0.05), MDM2 was significantly increased, and immunofluorescence showed reduced p53 and increased MDM2 expression in GLC nuclei. RG-7112 treatment significantly increased p53 and significantly decreased MDM2 gene and protein expression. GLC cells had significantly increased protein expression of αSMA, significantly decreased caspase-3 protein expression, and significantly increased proliferation after 96 h. RG-7112 treatment significantly decreased COL1A1 and αSMA, significantly increased BAX and caspase-3 gene expression, and significantly decreased proliferation in GLC cells. MTT-assay showed equivocal cellular viability in NLC/GLC cells with/without RG-7112 treatment. Our data suggests that proliferation and the ubiquitin-proteasomal pathway are dysregulated in GLC cells, with MDM2-led p53 protein degradation negatively impacting its protective role.

Arabidopsis GLYI4 Reveals Intriguing Insights into the JA Signaling Pathway and Plant Defense

Plant hormones play a central role in various physiological functions and mediate defense responses against (a)biotic stresses. Jasmonic acid (JA) has emerged as one of the key phytohormones involved in the response to necrotrophic pathogens. Under stressful conditions, plants can also produce small molecules, such as methylglyoxal (MG), a cytotoxic aldehyde. The enzymes glyoxalase I (GLYI) and glyoxalase II primarily detoxify MG. In Arabidopsis thaliana, GLYI4 has been recently characterized as having a crucial role in MG detoxification and emerging involvement in the JA pathway. Here, we investigated the impact of a GLYI4 loss-of-function on the Arabidopsis JA pathway and how MG affects it. The results showed that the glyI4 mutant plant had stunted growth, a smaller rosette diameter, reduced leaf size, and an altered pigment concentration. A gene expression analysis of the JA marker genes showed significant changes in the JA biosynthetic and signaling pathway genes in the glyI4 mutant. Disease resistance bioassays against the necrotroph Botrytis cinerea revealed altered patterns in the glyI4 mutant, likely due to increased oxidative stress. The MG effect has a further negative impact on plant performance. Collectively, these results contribute to clarifying the intricate interconnections between the GLYI4, MG, and JA pathways, opening up new avenues for further explorations of the intricate molecular mechanisms controlling plant stress responses.

Expression of distal limb patterning genes in Hypsibius exemplaris indicate regionalization and suggest distal identity of tardigrade legs

Expression of distal limb patterning genes in Hypsibius exemplaris indicate regionalization and suggest distal identity of tardigrade legs

MID1-COMPLEMENTING ACTIVITY regulates cell proliferation and development via Ca2+ signaling in Marchantia polymorpha

Abstract MID1-COMPLEMENTING ACTIVITY (MCA) is a land plant-specific, plasma membrane protein and Ca2+ signaling component that responds to exogenous mechanical stimuli, such as touch, gravity, and hypotonic-osmotic stress, in various plant species. MCA is essential for cell proliferation and differentiation during growth and development in rice (Oryza sativa) and maize (Zea mays). However, the mechanism by which MCA mediates cell proliferation and differentiation via Ca2+ signaling remains unknown. Here, we address this question using the liverwort Marchantia polymorpha. We show that the M. polymorpha MCA ortholog, MpMCA, is highly expressed in actively dividing regions, such as apical notches in the thalli and developing gametangiophores, and that MpMCA is a plasma membrane protein. In vivo Ca2+ imaging using a Ca2+ sensor (yellow cameleon) revealed that MpMCA is required for maintaining proper [Ca2+]cyt levels in the apical notch region, egg cells, and antheridium cells. Mpmca mutant plants showed severe cell proliferation and differentiation defects in the thalli, gametangiophores, and gametangia, resulting in abnormal development and unsuccessful fertilization. Furthermore, expression of the Arabidopsis MCA1 gene complemented most of the defects in the growth and development of the Mpmca mutant plants. Our findings indicate that MpMCA is an evolutionarily conserved Ca2+-signaling component that regulates cell proliferation and development across the life cycle of land plants.

Ectopic expression of a truncated NLR gene from wild Arachis enhances resistance to Fusarium oxysporum

Fusarium oxysporum causes devastating vascular wilt diseases in numerous crop species, resulting in substantial yield losses. The Arabidopsis thaliana-F. oxysporum f.sp. conglutinans (FOC) model system enables the identification of meaningful genotype–phenotype correlations and was applied in this study to evaluate the effects of overexpressing an NLR gene (AsTIR19) from Arachis stenosperma against pathogen infection. AsTIR19 overexpression (OE) lines exhibited enhanced resistance to FOC without any discernible phenotype penalties. To elucidate the underlying resistance mechanisms mediated by AsTIR19 overexpression, we conducted whole transcriptome sequencing of an AsTIR19-OE line and non-transgenic wild-type (WT) plants inoculated and non-inoculated with FOC using Illumina HiSeq4000. Comparative analysis revealed 778 differentially expressed genes (DEGs) attributed to transgene overexpression, while fungal inoculation induced 434 DEGs in the OE line, with many falling into defense-related Gene Ontology (GO) categories. GO and KEGG enrichment analysis showed that DEGs were enriched in the phenylpropanoid and flavonoid pathways in the OE plants. This comprehensive transcriptomic analysis underscores how AsTIR19 overexpression reprograms transcriptional networks, modulating the expression of stress-responsive genes across diverse metabolic pathways. These findings provide valuable insights into the molecular mechanisms underlying the role of this NLR gene under stress conditions, highlighting its potential to enhance resistance to Fusarium oxysporum.

Stem cell models of TAFAZZIN deficiency reveal novel tissue-specific pathologies in Barth syndrome

Abstract Barth syndrome (BTHS) is a rare mitochondrial disease caused by pathogenic variants in the gene TAFAZZIN, which leads to abnormal cardiolipin (CL) metabolism on the inner mitochondrial membrane. Although TAFAZZIN is ubiquitously expressed, BTHS involves a complex combination of tissue specific phenotypes including cardiomyopathy, neutropenia, skeletal myopathy, and growth delays, with a relatively minimal neurological burden. To understand both the developmental and functional effects of TAZ-deficiency in different tissues, we generated isogenic TAZ knockout (TAZ-KO) and WT cardiomyocytes (CMs) and neural progenitor cells (NPCs) from CRISPR-edited induced pluripotent stem cells (iPSCs). In TAZ-KO CMs we discovered evidence of dysregulated mitophagy including dysmorphic mitochondria and mitochondrial cristae, differential expression of key autophagy-associated genes, and an inability of TAZ-deficient CMs to properly initiate stress-induced mitophagy. In TAZ-deficient NPCs we identified novel phenotypes including a reduction in CIV abundance and CIV activity in the CIII2&CIV2 intermediate complex. Interestingly, while CL acyl chain manipulation was unable to alter mitophagy defects in TAZ-KO CMs, we found that linoleic acid or oleic acid supplementation was able to partially restore CIV abundance in TAZ-deficient NPCs. Taken together, our results have implications for understanding the tissue-specific pathology of BTHS and potential for tissue-specific therapeutic targeting. Moreover, our results highlight an emerging role for mitophagy in the cardiac pathophysiology of BTHS and reveal a potential neuron-specific bioenergetic phenotype.

Expression of the polyphenol oxidase gene MdPPO7 is modulated by MdWRKY3 to regulate browning in sliced apple fruit

Abstract Browning is a pervasive problem in horticultural products, substantially diminishing the appearance, flavor and nutritional value of fruit, including important fruits like apple (Malus × domestica Borkh.). In this study, we compared the physiological characteristics of the browning-resistant line ‘Rb-18’ with the susceptible variety ‘Fuji’ and found that the polyphenol oxidase (PPO) enzyme activity and phenolic content of Rb-18 were significantly lower than those in Fuji. In addition, the PPO enzyme in Fuji showed a stronger affinity for its substrate, catechol, compared to Rb-18. Through transcriptome and RT-qPCR analyses, MdPPO7 expression was identified as contributing to flesh browning after cutting. Subsequent fruit injection and stable genetic transformation of the MdPPO7 gene into apple fruit and calli determined that syringic acid, procyanidin, phloridzin, chlorogenic acid, gallic acid, catechin, and caffeic act as its catalytic substrates in the process involved in browning. Furthermore, luciferase reporter, yeast one-hybrid, β-glucuronidase reporter assays and chip-qPCR analysis demonstrated that a WRKY transcription factor(MdWRKY3) binds to the promoter region of polyphenol oxidase gene (MdPPO7) and positively regulates its expression to promote apple flesh browning. This study provides insights into the molecular regulatory mechanisms of fruit browning in fresh-cut apples and provides a theoretical basis for the generation of high-quality apple germplasm resources.

Multi-trait association analysis reveals shared genetic loci between Alzheimer’s disease and cardiovascular traits

AbstractSeveral cardiovascular traits and diseases co-occur with Alzheimer’s disease. We mapped their shared genetic architecture using multi-trait genome-wide association studies. Subsequent fine-mapping and colocalisation highlighted 16 genetic loci associated with both Alzheimer’s and cardiovascular diseases. We prioritised rs11786896, which colocalised with Alzheimer’s disease, atrial fibrillation and expression of PLEC in the heart left ventricle, and rs7529220, which colocalised with Alzheimer’s disease, atrial fibrillation and expression of C1Q family genes. Single-cell RNA-sequencing data, co-expression network and protein-protein interaction analyses provided evidence for different mechanisms of PLEC, which is upregulated in left ventricular endothelium and cardiomyocytes with heart failure and in brain astrocytes with Alzheimer’s disease. Similar common mechanisms are implicated for C1Q in heart macrophages with heart failure and in brain microglia with Alzheimer’s disease. These findings highlight inflammatory and pleomorphic risk determinants for the co-occurrence of Alzheimer’s and cardiovascular diseases and suggest PLEC, C1Q and their interacting proteins as potential therapeutic targets.

The role of miR-152 in urological tumors: potential biomarkers and therapeutic targets

Urological malignant tumors pose a significant threat to human health, with a high incidence rate each year. Prostate cancer, bladder cancer, and renal cell carcinoma are among the most prevalent and extensively researched urological malignancies. Despite advancements in research, the prognosis for these tumors remains unfavorable due to late detection, postoperative recurrence, and treatment resistance. A thorough investigation into their pathogenesis is crucial for early diagnosis and treatment. Recent studies have highlighted the close association between microRNAs (miRNAs) and cancer progression. miRNAs are small non-coding RNAs composed of 19-23 nucleotides that regulate gene expression by binding to the 3’ untranslated region (3’UTR) of target mRNAs, impacting key cellular processes such as proliferation, differentiation, apoptosis, and migration. Dysregulation of miRNAs can disrupt the expression of oncogenes and tumor suppressor genes, contributing to cancer development. Among the various miRNAs studied, miR-152 has garnered attention for its role in urological malignancies. Several studies have indicated that dysregulation of miR-152 expression is significant in these cancers, warranting a comprehensive review of the evidence. This review focuses on the expression and function of miR-152 in prostate cancer, bladder cancer, and renal cell carcinoma, elucidating its mechanisms in cancer progression and exploring its potential as a therapeutic target and biomarker in urological malignancies.

A single upstream mutation of whiB7 underlies amikacin and clarithromycin resistance in Mycobacterium abscessus

Abstract Aims We aimed to investigate the molecular mechanisms underlying the survival of Mycobacterium abscessus when faced with antibiotic combination therapy. By conducting evolution experiments and whole genome sequencing (WGS), we sought to identify genetic variants associated with stress response mechanisms, with a particular focus on drug survival and resistance. Methods and results We conducted evolution experiments on M. abscessus, exposing the bacteria to a combination therapy of amikacin and rifabutin. Genetic mutations associated with increased antibiotic survival and altered susceptibility were subsequently identified by WGS. We focused on mutations that contribute to stress response mechanisms and tolerance. Of particular interest was a novel frameshift mutation in MAB_3509c, a gene of unknown function within the upstream open reading frame of whiB7. A MAB_3509c knockout mutant was constructed, and expression of downstream drug resistance genes was assessed by RT-qPCR. Mutation of MAB_3509c results in increased RNA levels of whiB7 and downstream stress response genes such as eis2, which is responsible for aminoglycoside resistance. Conclusion Our findings demonstrate the importance of whiB7 in the adaptive stress response in M. abscessus. Moreover, our results highlight the complexity of M. abscessus adapting to drug stress and underscore the need for further research.

Single-cell transcriptomics unveils skin cell specific antifungal immune responses and IL-1Ra- IL-1R immune evasion strategies of emerging fungal pathogen Candida auris

Candida auris is an emerging multidrug-resistant fungal pathogen that preferentially colonizes and persists in skin tissue, yet the host immune factors that regulate the skin colonization of C. auris in vivo are unknown. In this study, we employed unbiased single-cell transcriptomics of murine skin infected with C. auris to understand the cell type-specific immune response to C. auris. C. auris skin infection results in the accumulation of immune cells such as neutrophils, inflammatory monocytes, macrophages, dendritic cells, T cells, and NK cells at the site of infection. We identified fibroblasts as a major non-immune cell accumulated in the C. auris infected skin tissue. The comprehensive single-cell profiling revealed the transcriptomic signatures in cytokines, chemokines, host receptors (TLRs, C-type lectin receptors, NOD receptors), antimicrobial peptides, and immune signaling pathways in individual immune and non-immune cells during C. auris skin infection. Our analysis revealed that C. auris infection upregulates the expression of the IL-1RN gene (encoding IL-1R antagonist protein) in different cell types. We found IL-1Ra produced by macrophages during C. auris skin infection decreases the killing activity of neutrophils. Furthermore, C. auris uses a unique cell wall mannan outer layer to evade IL-1R-signaling mediated host defense. Collectively, our single-cell RNA seq profiling identified the transcriptomic signatures in immune and non-immune cells during C. auris skin infection. Our results demonstrate the IL-1Ra and IL-1R-mediated immune evasion mechanisms employed by C. auris to persist in the skin. These results enhance our understanding of host defense and immune evasion mechanisms during C. auris skin infection and identify potential targets for novel antifungal therapeutics.

Ferroptosis in Cancer: Epigenetic Control and Therapeutic Opportunities

Ferroptosis, an iron-dependent form of regulated cell death driven by lipid peroxidation, has emerged as a critical pathway in cancer biology. This review delves into the epigenetic mechanisms that modulate ferroptosis in cancer cells, focusing on how DNA methylation, histone modifications, and non-coding RNAs influence the expression and function of essential genes involved in this process. By unraveling the complex interplay between these epigenetic mechanisms and ferroptosis, the article sheds light on novel gene targets and functional insights that could pave the way for innovative cancer treatments to enhance therapeutic efficacy and overcome resistance in cancer therapy.