Overexpression Of Transcription Factor Research Articles

Physiological and molecular responses of 'Hamlin' sweet orange trees expressing the VvmybA1 gene under cold stress conditions.

Overexpression of VvmybA1 transcription factor in 'Hamlin' citrus enhances cold tolerance by increasing anthocyanin accumulation. This results in improved ROS scavenging, altered gene expression, and stomatal regulation, highlighting anthocyanins' essential role in citrus cold acclimation. Cold stress is a significant threat to citrus cultivation, impacting tree health and productivity. Anthocyanins are known for their role as pigments and have emerged as key mediators of plant defense mechanisms against environmental stressors. This study investigated the potential of anthocyanin overexpression regulated by grape (Vitis vinifera) VvmybA1 transcription factor to enhance cold stress tolerance in citrus trees. Transgenic 'Hamlin' citrus trees overexpressing VvmybA1 were exposed to a 30-day cold stress period at 4°C along with the control wild-type trees. Our findings reveal that anthocyanin accumulation significantly influences chlorophyll content and their fluorescence parameters, affecting leaf responses to cold stress. Additionally, we recorded enhanced ROS scavenging capacity and distinct expression patterns of key transcription factors and antioxidant-related genes inthe transgenic leaves. Furthermore, VvmybA1 overexpression affected stomatal aperture regulation by moderating ABA biosynthesis, resulting in differential responses in a stomatal opening between transgenic and wild-type trees under cold stress. Transgenic trees exhibited reduced hydrogen peroxide levels, enhanced flavonoids, radical scavenging activity, and altered phytohormonal profiles. These findings highlighted the role of VvmybA1-mediated anthocyanin accumulation in enhancing cold tolerance. The current study alsounderlines the potential of anthocyanin overexpression as a critical regulator of the cold acclimation process by scavenging ROS in plant tissues.

Novel transcriptional regulation of the GAP promoter in Pichia pastoris towards high expression of heterologous proteins

BackgroundPichia pastoris (Komagataella phaffii) is a promising production host, but the usage of methanol limits its application in the medicine and food industries.ResultsTo improve the constitutive expression of heterologous proteins in P. pastoris, four new potential transcription regulators (Loc1p, Msn2p, Gsm1p, Hot1p) of the glyceraldehyde triphosphate dehydrogenase promoter (pGAP) were revealed in this study by using cellulase E4 as reporter gene. On this basis, a series of P. pastoris strains with knockout or overexpression of transcription factors were constructed and the deletion of transcription factor binding sites on pGAP was confirmed. The results showed that Loc1p and Msn2p can inhibit the activity of pGAP, while Gsm1p and Hot1p can enhance the activity of pGAP; Loc1p, Gsm1p and Hot1p can bind directly to pGAP, while Msn2p must be treated to expose the C-terminal domain to bind to pGAP. Moreover, manipulating a single transcription factor led to a 0.96-fold to 2.43-fold increase in xylanase expression. In another model protein, aflatoxin oxidase, knocking out Loc1 based on AFO-∆Msn2 strain resulted in a 0.63-fold to 1.4-fold increase in expression. It can be demonstrated that the combined use of transcription factors can further improve the expression of exogenous proteins in P. pastoris.ConclusionThese findings will contribute to the construction of pGAP-based P. pastoris systems towards high expression of heterologous proteins, hence improving the application potential of yeast.

Efficient Dlx2-mediated astrocyte-to-neuron conversion and inhibition of neuroinflammation by NeuroD1.

In vivo astrocyte-to-neuron (AtN) conversion induced by overexpression of neural transcriptional factors has great potential for neural regeneration and repair. Here, we demonstrate that a single neural transcriptional factor, Dlx2, converts mouse striatal astrocytes into neurons in a dose-dependent manner. Lineage-tracing studies in Aldh1l1-CreERT2 mice confirm that Dlx2 can convert striatal astrocytes into DARPP32+ and Ctip2+ medium spiny neurons (MSNs). Time-course studies reveal a gradual conversion from astrocytes to neurons in 1 month, with a distinct intermediate state in between astrocytes and neurons. Interestingly, when Dlx2-infected astrocytes start to lose astrocytic markers, the other local astrocytes proliferate to maintain astrocytic levels in the converted areas. Unexpectedly, although Dlx2 efficiently reprograms astrocytes into neurons in the gray matter striatum, it also induces partial reprogramming of astrocytes in the white matter corpus callosum. Such partial reprogramming of white matter astrocytes is associated with neuroinflammation, which can be suppressed by the addition of NeuroD1. Our results highlight the importance of investigating AtN conversion in both the gray matter and white matter to thoroughly evaluate therapeutic potentials. This study also unveils the critical role of anti-inflammation by NeuroD1 during AtN conversion.

NMR Fingerprinting of Conventional and Genetically Modified Soybean Plants with AtAREB1 Transcription Factors.

Drought stress impacts soybean yields and physiological processes. However, the insertion of the activated form of the AtAREB1 gene in the soybean cultivar BR16, which is sensitive to water deficit, improved the drought response of the genetically modified plants. Thus, in this study, we used 1H NMR in solution and solid-state NMR to investigate the response of genetically modified soybean overexpressing AtAREB1 under water deficiency conditions. We achieved that drought-tolerant soybean yields high content of amino acids isoleucine, leucine, threonine, valine, proline, glutamate, aspartate, asparagine, tyrosine, and phenylalanine after 12 days of drought stress conditions, as compared to drought-sensitive soybean under the same conditions. Specific target compounds, including sugars, organic acids, and phenolic compounds, were identified as involved in controlling sensitive soybean during the vegetative stage. Solid-state NMR was used to study the impact of drought stress on starch and cellulose contents in different soybean genotypes. The findings provide insights into the metabolic adjustments of soybean overexpressing AREB transcription factors in adapting to dry climates. This study presents NMR techniques for investigating the metabolome of transgenic soybean plants in response to the water deficit. The approach allowed for the identification of physiological and morphological changes in drought-resistant and drought-tolerant soybean tissues. The findings indicate that drought stress significantly alters micro- and macromolecular metabolism in soybean plants. Differential responses were observed among roots and leaves as well as drought-tolerant and drought-sensitive cultivars, highlighting the complex interplay between overexpressed transcription factors and drought stress in soybean plants.

Distinct mechanisms of non-autonomous UPRER mediated by GABAergic, glutamatergic, and octopaminergic neurons.

The capacity to deal with stress declines during the aging process, and preservation of cellular stress responses is critical to healthy aging. The unfolded protein response of the endoplasmic reticulum (UPRER) is one such conserved mechanism, which is critical for the maintenance of several major functions of the ER during stress, including protein folding and lipid metabolism. Hyperactivation of the UPRER by overexpression of the major transcription factor, xbp-1s, solely in neurons drives lifespan extension as neurons send a neurotransmitter-based signal to other tissue to activate UPRER in a non-autonomous fashion. Previous work identified serotonergic and dopaminergic neurons in this signaling paradigm. To further expand our understanding of the neural circuitry that underlies the non-autonomous signaling of ER stress, we activated UPRER solely in glutamatergic, octopaminergic, and GABAergic neurons in C. elegans and paired whole-body transcriptomic analysis with functional assays. We found that UPRER-induced signals from glutamatergic neurons increased expression of canonical protein homeostasis pathways and octopaminergic neurons promoted pathogen response pathways, while minor, but statistically significant changes were observed in lipid metabolism-related genes with GABAergic UPRER activation. These findings provide further evidence for the distinct role neuronal subtypes play in driving the diverse response to ER stress.

Comprehensive network modeling approaches unravel dynamic enhancer-promoter interactions across neural differentiation.

Increasing evidence suggests that a substantial proportion of disease-associated mutations occur in enhancers, regions of non-coding DNA essential to gene regulation. Understanding the structures and mechanisms of regulatory programs this variation affects can shed light on the apparatuses of human diseases. We collected epigenetic and gene expression datasets from seven early time points during neural differentiation. Focusing on this model system, we constructed networks of enhancer-promoter interactions, each at an individual stage of neural induction. These networks served as the base for a rich series of analyses, through which we demonstrated their temporal dynamics and enrichment for various disease-associated variants. We applied the Girvan-Newman clustering algorithm to these networks to reveal biologically relevant substructures of regulation. Additionally, we demonstrated methods to validate predicted enhancer-promoter interactions using transcription factor overexpression and massively parallel reporter assays. Our findings suggest a generalizable framework for exploring gene regulatory programs and their dynamics across developmental processes. This includes a comprehensive approach to studying the effects of disease-associated variation on transcriptional networks. The techniques applied to our networks have been published alongside our findings as a computational tool, E-P-INAnalyzer. Our procedure can be utilized across different cellular contexts and disorders.

Transcriptomic responses to drought stress in the Patagonian southern beech Nothofagus alpina

BackgroundDeciphering the genetic architecture of drought tolerance could allow the candidate genes identification responding to water stress. In the Andean Patagonian forest, the genus Nothofagus represents an ecologically relevant species to be included in different genomic studies. These studies are scarce in South American ecosystems however represent an important source of genomic data in order to interpret future climate-change environment scenarios of these emblematic forests. Here, we achieved the assemblage of the transcriptome of N. alpina while searching for key genes of activated or suppressed metabolic pathways in response to drought stress.ResultsDe novo transcriptome assembly resulted in 104,030 transcripts. Following confirmation of drought conditions, based on reduction of leaf water potential and stomatal conductance, a differential gene expression analysis resulted in 2720 significantly expressed genes (1601 up-regulated and 1119 down-regulated). Enrichment analysis (over-representation analysis and gene set enrichment analysis) resulted in more than one hundred stress-responsive term ontologies (i.e. biological processes) and pathways. Terms such as response to abscisic acid and pathways such as plant hormone signal transduction or starch and sucrose metabolism were over-represented. Protein–protein interaction assessment resulted in networks with significantly expressed top common hub gene clusters (e.g. plant-type cell wall biogenesis among down-regulated or ABA-signalling among up-regulated). These networks evidenced important regulators at gene expression such as transcriptional factors.ConclusionsResponses of N. alpina seedlings to drought stress were evidenced by the activation of several genes linked to GO biological processes and KEGG pathways, which were mainly based on over-expression of specific protein kinases, phosphatases, synthases and transcription factors. This suggests an up-regulation of signalling pathways, triggered through plant hormones such as abscisic acid or auxin, which could counteract the osmotic stress created as a probable immediate response to drought. On the other hand, groups of carbon fixation genes related to the galactose metabolism, photosynthesis, secondary wall biogenesis, and fatty acid biosynthesis degradation were down-regulated under drought. Overall, our results provide new genomic data for understanding how non-model long-lived trees of Patagonian forests would acclimate to environmental changes.

A Case Study from the Overexpression of OsTZF5, Encoding a CCCH Tandem Zinc Finger Protein, in Rice Plants Across Nineteen Yield Trials

BackgroundDevelopment of transgenic rice overexpressing transcription factors involved in drought response has been previously reported to confer drought tolerance and therefore represents a means of crop improvement. We transformed lowland rice IR64 with OsTZF5, encoding a CCCH-tandem zinc finger protein, under the control of the rice LIP9 stress-inducible promoter and compared the drought response of transgenic lines and nulls to IR64 in successive screenhouse paddy and field trials up to the T6 generation.ResultsCompared to the well-watered conditions, the level of drought stress across experiments varied from a minimum of − 25 to − 75 kPa at a soil depth of 30 cm which reduced biomass by 30–55% and grain yield by 1–92%, presenting a range of drought severities. OsTZF5 transgenic lines showed high yield advantage under drought over IR64 in early generations, which was related to shorter time to flowering, lower shoot biomass and higher harvest index. However, the increases in values for yield and related traits in the transgenics became smaller over successive generations despite continued detection of drought-induced transgene expression as conferred by the LIP9 promoter. The decreased advantage of the transgenics over generations tended to coincide with increased levels of homozygosity. Background cleaning of the transgenic lines as well as introgression of the transgene into an IR64 line containing major-effect drought yield QTLs, which were evaluated starting at the BC3F1 and BC2F3 generation, respectively, did not result in consistently increased yield under drought as compared to the respective checks.ConclusionsAlthough we cannot conclusively explain the genetic factors behind the loss of yield advantage of the transgenics under drought across generations, our results help in distinguishing among potential drought tolerance mechanisms related to effectiveness of the transgenics, since early flowering and harvest index most closely reflected the levels of yield advantage in the transgenics across generations while reduced biomass did not.

EP300/CREBBP acetyltransferase inhibition limits steroid receptor and FOXA1 signaling in prostate cancer cells

The androgen receptor (AR) is a primary target for treating prostate cancer (PCa), forming the bedrock of its clinical management. Despite their efficacy, resistance often hampers AR-targeted therapies, necessitating new strategies against therapy-resistant PCa. These resistances involve various mechanisms, including AR splice variant overexpression and altered activities of transcription factors like the glucocorticoid receptor (GR) and FOXA1. These factors rely on common coregulators, such as EP300/CREBBP, suggesting a rationale for coregulator-targeted therapies. Our study explores EP300/CREBBP acetyltransferase inhibition’s impact on steroid receptor and FOXA1 signaling in PCa cells using genome-wide techniques. Results reveal that EP300/CREBBP inhibition significantly disrupts the AR-regulated transcriptome and receptor chromatin binding by reducing the AR-gene expression. Similarly, GR’s regulated transcriptome and receptor binding were hindered, not linked to reduced GR expression but to diminished FOXA1 chromatin binding, restricting GR signaling. Overall, our findings highlight how EP300/CREBBP inhibition distinctively curtails oncogenic transcription factors’ signaling, suggesting the potential of coregulatory-targeted therapies in PCa.

Promising approaches for simultaneous enhancement of medicinally significant benzylisoquinoline alkaloids in opium poppy.

Benzylisoquinoline alkaloids (BIAs) produced in opium poppy have been evidenced to heal patients suffering from various diseases. They, therefore, hold an integral position in the herbal drug industry. Despite the adoption of several approaches for the large-scale production of BIAs, opium poppy remains the only platform in this purpose. The only disadvantage associated with producing BIAs in the plant is their small quantity. Thus, recruiting strategies that boost their levels is deemed necessary. All the methods which have been employed so far are just able to enhance a maximum of two BIAs. Thus, if these methods are utilized, a sizable amount of time and budget must be spent on the synthesis of all BIAs. Hence, the exploitation of strategies which increase the content of all BIAs at the same time is more commercially effective and time-saving, avoiding the laborious step of resolving the biosynthetic pathway of each compound. Exposure to biotic and abiotic elicitors, development of a synthetic auto-tetraploid, overexpression of a WRKY transcription factor, formation of an artificial metabolon, and suppression of a gene in the shikimate pathway and miRNA are strategies that turn opium poppy into a versatile bioreactor for the concurrent and massive production of BIAs. The last three strategies have never been applied for BIA biosynthetic pathways.

Overexpression of a grapevine VqWRKY2 transcription factor in Arabidopsis thaliana increases resistance to powdery mildew

Overexpression of a grapevine VqWRKY2 transcription factor in Arabidopsis thaliana increases resistance to powdery mildew

Restored glyoxylate metabolism after AGXT gene correction and direct reprogramming of primary hyperoxaluria type 1 fibroblasts

Primary hyperoxaluria type 1 (PH1) is a rare inherited metabolic disorder characterized by oxalate overproduction in the liver, resulting in renal damage. It is caused by mutations in the AGXT gene. Combined liver and kidney transplantation is currently the only permanent curative treatment. We combined locus-specific gene correction and hepatic direct cell reprogramming to generate autologous healthy induced hepatocytes (iHeps) from PH1 patient-derived fibroblasts. First, site-specific AGXT corrected cells were obtained by homology directed repair (HDR) assisted by CRISPR-Cas9, following two different strategies: accurate point mutation (c.731T>C) correction or knockin of an enhanced version of AGXT cDNA. Then, iHeps were generated, by overexpression of hepatic transcription factors. Generated AGXT-corrected iHeps showed hepatic gene expression profile and exhibited invitro reversion of oxalate accumulation compared to non-edited PH1-derived iHeps. This strategy set up a potential alternative cellular source for liver cell replacement therapy and a personalized PH1 invitro disease model.

The use of a SOX10 reporter toward ameliorating oligodendrocyte lineage differentiation from human induced pluripotent stem cells.

Oligodendrocytes (OLs) are key players in the central nervous system, critical for the formation and maintenance of the myelin sheaths insulating axons, ensuring efficient neuronal communication. In the last decade, the use of human induced pluripotent stem cells (iPSCs) has become essential for recapitulating and understanding the differentiation and role of OLs in vitro. Current methods include overexpression of transcription factors for rapid OL generation, neglecting the complexity of OL lineage development. Alternatively, growth factor-based protocols offer physiological relevance but struggle with efficiency and cell heterogeneity. To address these issues, we created a novel SOX10-P2A-mOrange iPSC reporter line to track and purify oligodendrocyte precursor cells. Using this reporter cell line, we analyzed an existing differentiation protocol and shed light on the origin of glial cell heterogeneity. Additionally, we have modified the differentiation protocol, toward enhancing reproducibility, efficiency, and terminal maturity. Our approach not only advances OL biology but also holds promise to accelerate research and translational work with iPSC-derived OLs.

Impaired innate and adaptive immune responses to BNT162b2 SARS-CoV-2 vaccination in systemic lupus erythematosus.

Understanding the immune responses to SARS-CoV-2 vaccination is critical to optimizing vaccination strategies for individuals with autoimmune diseases, such as systemic lupus erythematosus (SLE). Here, we comprehensively analyzed innate and adaptive immune responses in 19 patients with SLE receiving a complete 2-dose Pfizer-BioNTech mRNA vaccine (BNT162b2) regimen compared with a control cohort of 56 healthy control (HC) volunteers. Patients with SLE exhibited impaired neutralizing antibody production and antigen-specific CD4+ and CD8+ T cell responses relative to HC. Interestingly, antibody responses were only altered in patients with SLE treated with immunosuppressive therapies, whereas impairment of antigen-specific CD4+ and CD8+ T cell numbers was independent of medication. Patients with SLE also displayed reduced levels of circulating CXC motif chemokine ligands, CXCL9, CXCL10, CXCL11, and IFN-γ after secondary vaccination as well as downregulation of gene expression pathways indicative of compromised innate immune responses. Single-cell RNA-Seq analysis reveals that patients with SLE showed reduced levels of a vaccine-inducible monocyte population characterized by overexpression of IFN-response transcription factors. Thus, although 2 doses of BNT162b2 induced relatively robust immune responses in patients with SLE, our data demonstrate impairment of both innate and adaptive immune responses relative to HC, highlighting a need for population-specific vaccination studies.

Melatonin supplementation enhances browning suppression and improves transformation efficiency and regeneration of transgenic rough lemon plants (Citrus × jambhiri).

Enzymatic browning poses a significant challenge that limits in vitro propagation and genetic transformation of plant tissues. This research focuses on investigating how adding antioxidant substances can suppress browning, leading to improved efficiency in transforming plant tissues using Agrobacterium and subsequent plant regeneration from rough lemon (Citrus × jambhiri). When epicotyl segments of rough lemon were exposed to Agrobacterium, they displayed excessive browning and tissue decay. This was notably different from the 'Hamlin' explants, which did not exhibit the same issue. The regeneration process failed completely in rough lemon explants, and they accumulated high levels of total phenolic compounds (TPC) and polyphenol oxidase (PPO), which contribute to browning. To overcome these challenges, several antioxidant and osmoprotectant compounds, including lipoic acid, melatonin, glycine betaine, and proline were added to the tissue culture medium to reduce the oxidation of phenolic compounds and mitigate browning. Treating epicotyl segments with 100 or 200 μM melatonin led to a significant reduction in browning and phenolic compound accumulation. This resulted in enhanced shoot regeneration, increased transformation efficiency, and reduced tissue decay. Importantly, melatonin supplementation effectively lowered the levels of TPC and PPO in the cultured explants. Molecular and physiological analyses also confirmed the successful overexpression of the CcNHX1 transcription factor, which plays a key role in imparting tolerance to salinity stress. This study emphasizes the noteworthy impact of supplementing antioxidants in achieving successful genetic transformation and plant regeneration in rough lemon. These findings provide valuable insights for developing strategies to address enzymatic browning and enhance the effectiveness of plant tissue culture and genetic engineering methods with potential applications across diverse plant species.

IGF-1-mediated FOXC1 overexpression induces stem-like properties through upregulating CBX7 and IGF-1R in esophageal squamous cell carcinoma.

Substantial evidence attests to the pivotal role of cancer stem cells (CSC) in both tumorigenesis and drug resistance. A member of the forkhead box (FOX) family, FOXC1, assumes significance in embryonic development and organogenesis. Furthermore, FOXC1 functions as an overexpressed transcription factor in various tumors, fostering proliferation, enhancing migratory capabilities, and promoting drug resistance, while maintaining stem-cell-like properties. Despite these implications, scant attention has been devoted to its role in esophageal squamous cell carcinoma. Our investigation revealed a pronounced upregulation of FOXC1 expression in ESCC, correlating with a poor prognosis. The downregulation of FOXC1 demonstrated inhibitory effects on ESCC tumorigenesis, proliferation, and tolerance to chemotherapeutic agents, concurrently reducing the levels of stemness-related markers CD133 and CD44. Further studies validated that FOXC1 induces ESCC stemness by transactivating CBX7 and IGF-1R. Additionally, IGF-1 activated the PI3K/AKT/NF-κB and MEK/ERK/NF-κB pathways through its binding to IGF-1R, thereby augmenting FOXC1 expression. Conversely, suppressing FOXC1 impeded ESCC stemness induced by IGF-1. The presence of a positive feedback loop, denoted by IGF-1-FOXC1-IGF-1R, suggests the potential of FOXC1 as a prognostic biomarker for ESCC. Taken together, targeting the IGF-1-FOXC1-IGF-1R axis emerges as a promising approach for anti-CSC therapy in ESCC.

Exploring the Interplay of RUNX2 and CXCR4 in Melanoma Progression.

Overexpression of the Runt-related transcription factor 2 (RUNX2) has been reported in several cancer types, and the C-X-C motif chemokine receptor 4 (CXCR4) has an important role in tumour progression. However, the interplay between CXCR4 and RUNX2 in melanoma cells remains poorly understood. In the present study, we used melanoma cells and a RUNX2 knockout (RUNX2-KO) in vitro model to assess the influence of RUNX2 on CXCR4 protein levels along with its effects on markers associated with cell invasion and autophagy. Osteotropism was assessed using a 3D microfluidic model. Moreover, we assessed the impact of CXCR4 on the cellular levels of key cellular signalling proteins involved in autophagy. We observed that melanoma cells express both RUNX2 and CXCR4. Restored RUNX2 expression in RUNX2 KO cells increased the expression levels of CXCR4 and proteins associated with the metastatic process. The protein markers of autophagy LC3 and beclin were upregulated in response to increased CXCR4 levels. The CXCR4 inhibitor WZ811 reduced osteotropism and activated the mTOR and p70-S6 cell signalling proteins. Our data indicate that the RUNX2 transcription factor promotes the expression of the CXCR4 chemokine receptor on melanoma cells, which in turn promotes autophagy, cell invasiveness, and osteotropism, through the inhibition of the mTOR signalling pathway. Our data suggest that RUNX2 promotes melanoma progression by upregulating CXCR4, and we identify the latter as a key player in melanoma-related osteotropism.

Transcription factors FtMYB22 negatively regulates anthocyanin synthesis by forming an MBW ternary complex in Tartary buckwheat

AbstractAnthocyanins are important flavonoid compounds that are abundant in Tartary buckwheat, providing a variety of bright colors and enhancing abiotic stress resistance. Our study found that FtMYB22 specifically affected anthocyanin anabolism. In this study, overexpression of the FtMYB22 transcription factor in tobacco significantly reduced the accumulation of flavonol in the petals of transgenic tobacco plants. Moreover, it significantly downregulated the expression of the genes involved in anthocyanin biosynthesis in tobacco. Overexpression of the FtMYB22 transcription factor in Tartary buckwheat hairy roots significantly inhibited anthocyanin and total flavonoids biosynthesis, and downregulated the expression of the anthocyanin biosynthesis genes. Comparing wild‐type and transgenic hairy roots, FtMYB22 overexpression significantly inhibited anthocyanin accumulation and downregulated most flavonoid synthesis pathway gene expression. Yeast two‐hybrid assays demonstrated that FtMYB22 could interact with the FtbHLH family member FtTT8 and the FtWD40 family member FtTTG1 to form a ternary MBW complex to inhibit anthocyanin synthesis. Therefore, FtMYB22 can be exploited as a target site for further gene editing and molecular breeding to improve the resilience and quality of Tartary buckwheat.

Effect of Octamer-Binding Transcription Factor 4 Overexpression on the Neural Induction of Human Dental Pulp Stem Cells

Stem cell-based therapy is a potential alternative strategy for brain repair, with neural stem cells (NSC) presenting as the most promising candidates. Obtaining sufficient quantities of NSC for clinical applications is challenging, therefore alternative cell types, such as neural crest-derived dental pulp stem cells (DPSC), may be considered. Human DPSC possess neurogenic potential, exerting positive effects in the damaged brain through paracrine effects. However, a method for conversion of DPSC into NSC has yet to be developed. Here, overexpression of octamer-binding transcription factor 4 (OCT4) in combination with neural inductive conditions was used to reprogram human DPSC along the neural lineage. The reprogrammed DPSC demonstrated a neuronal-like phenotype, with increased expression levels of neural markers, limited capacity for sphere formation, and enhanced neuronal but not glial differentiation. Transcriptomic analysis further highlighted the expression of genes associated with neural and neuronal functions. In vivo analysis using a developmental avian model showed that implanted DPSC survived in the developing central nervous system and respond to endogenous signals, displaying neuronal phenotypes. Therefore, OCT4 enhances the neural potential of DPSC, which exhibited characteristics aligning with neuronal progenitors. This method can be used to standardise DPSC neural induction and provide an alternative source of neural cell types.Graphical

Mycobacterium tuberculosis response to cholesterol is integrated with environmental pH and potassium levels via a lipid metabolism regulator.

Successful colonization of the host requires Mycobacterium tuberculosis (Mtb) to sense and respond coordinately to disparate environmental cues during infection and adapt its physiology. However, how Mtb response to environmental cues and the availability of key carbon sources may be integrated is poorly understood. Here, by exploiting a reporter-based genetic screen, we have unexpectedly found that overexpression of transcription factors involved in Mtb lipid metabolism altered the dampening effect of low environmental potassium concentrations ([K+]) on the pH response of Mtb. Cholesterol is a major carbon source for Mtb during infection, and transcriptional analyses revealed that Mtb response to acidic pH was augmented in the presence of cholesterol and vice versa. Strikingly, deletion of the putative lipid regulator mce3R had little effect on Mtb transcriptional response to acidic pH or cholesterol individually, but resulted specifically in loss of cholesterol response augmentation in the simultaneous presence of acidic pH. Similarly, while mce3R deletion had little effect on Mtb response to low environmental [K+] alone, augmentation of the low [K+] response by the simultaneous presence of cholesterol was lost in the mutant. Finally, a mce3R deletion mutant was attenuated for growth in foamy macrophages and for colonization in a murine infection model that recapitulates caseous necrotic lesions and the presence of foamy macrophages. These findings reveal the critical coordination between Mtb response to environmental cues and cholesterol, a vital carbon source, and establishes Mce3R as a transcription factor that crucially serves to integrate these signals.