Transcriptional Profiling Research Articles

Sink strength, nutrient allocation, cannabinoid yield and associated transcript profiles vary in two drug-type Cannabis chemovars.

Cannabis sativa L. is one of the oldest domesticated crops. Hemp-type cultivars, which predominantly produce non-intoxicating cannabidiol (CBD), have been selected for their fast growth, seed, and fibre production, while drug-type chemovars were bred for high accumulation of tetrahydrocannabinol (THC). We investigated how the generation of CBD-dominant chemovars by introgression of hemp- into drug-type Cannabis impacted plant performance. The THC-dominant chemovar showed superior sink strength, higher flower biomass and demand-driven control of nutrient uptake. By contrast, the CBD-dominant chemovar hyperaccumulated phosphate in sink organs leading to reduced carbon and nitrogen assimilation in leaves, which limited flower biomass and cannabinoid yield. RNA-seq analyses determined organ- and chemovar-specific differences in expression of genes associated with nitrate and phosphate homeostasis as well as growth-regulating transcription factors that were correlated with measured traits. Among these were genes positively selected for during Cannabis domestication encoding an inhibitor of the phosphate starvation response SPX DOMAIN GENE3, nitrate reductase and two nitrate transporters. Altered nutrient sensing, acquisition or distribution are likely a consequence of adaption to growth on marginal, low-nutrient input lands in hemp. Our data provide evidence that such ancestral traits may become detrimental for female flower development and consequently overall CBD yield in protected cropping environments.

Apple vacuolar sugar transporters regulated by MdDREB2A enhance drought resistance by promoting accumulation of soluble sugars and activating ABA signaling

Abstract Soluble sugars are not only an important contributor to fruit quality, but also serve as the osmotic regulators in response to abiotic stresses. Early drought stress promotes sugar accumulation, while specific sugar transporters govern the cellular distribution of the sugars. Here, we show that apple plantlets accumulate soluble sugars in leaf tissues under drought stress. Transcriptional profiling of stressed and control plantlets revealed differential expression of several plasma membrane- or vacuolar membrane-localized sugar transporter genes. Among these, four previously identified vacuolar sugar transporter (VST) genes (MdERDL6–1, MdERDL6–2, MdTST1 and MdTST2) showed higher expression under drought, suggesting their roles in response to drought stress. Promoter cis-elements analyses, yeast one-hybrid and dual-luciferase tests confirmed that the drought-induced transcription factor MdDREB2A could promote the expression of MdERDL6–1/−2 and MdTST1/2 by binding to their promoter regions. Moreover, overexpressing of each of these four MdVSTs alone in transgenic apple or Arabidopsis plants accumulated more soluble sugars and abscisic acid, and enhanced drought resistance. Furthermore, apple plants overexpressing MdERDL6–1 also showed reduced water potential, facilitated stomatal closure and reactive oxygen species scavenging under drought condition compared to control plants. Overall, our results suggest a potential strategy to enhance drought resistance and sugar accumulation in fruits through manipulating the genes involved in vacuolar sugar transport.

Genome Engineering of Primary and Pluripotent Stem Cell-Derived Hepatocytes for Modeling Liver Tumor Formation

Genome editing has demonstrated its utility in generating isogenic cell-based disease models, enabling the precise introduction of genetic alterations into wild-type cells to mimic disease phenotypes and explore underlying mechanisms. However, its application in liver-related diseases has been limited by challenges in genetic modification of mature hepatocytes in a dish. Here, we conducted a systematic comparison of various methods for primary hepatocyte culture and gene delivery to achieve robust genome editing of hepatocytes ex vivo. Our efforts yielded editing efficiencies of up to 80% in primary murine hepatocytes cultured in monolayer and 20% in organoids. To model human hepatic tumorigenesis, we utilized hepatocytes differentiated from human pluripotent stem cells (hPSCs) as an alternative human hepatocyte source. We developed a series of cellular models by introducing various single or combined oncogenic alterations into hPSC-derived hepatocytes. Our findings demonstrated that distinct mutational patterns led to phenotypic variances, affecting both overgrowth and transcriptional profiles. Notably, we discovered that the PI3KCA E542K mutant, whether alone or in combination with exogenous c-MYC, significantly impaired hepatocyte functions and facilitated cancer metabolic reprogramming, highlighting the critical roles of these frequently mutated genes in driving liver neoplasia. In conclusion, our study demonstrates genome-engineered hepatocytes as valuable cellular models of hepatocarcinoma, providing insights into early tumorigenesis mechanisms.

Refined home-brew media for cost-effective, weekend-free hiPSC culture and genetic engineering

Background Cost-effective, practical, and reproducible culture of human induced pluripotent stem cells (hiPSCs) is required for both basic and translational research. This is especially crucial for large-scale expansion of hiPSCs for cell therapy, which should be made accessible to many patients regardless of their socioeconomic background. Basal 8 (B8) has emerged as a cost-effective solution for weekend-free and chemically-defined hiPSC culture. However, homebrewing of some recombinant growth factors for B8 can be a bottleneck towards both access and reproducibility of this technology. Moreover, we found the published B8 formulation to be suboptimal in normoxic hiPSC culture, which is widely used. Lastly, the suitability of B8 for applications such as genome editing or organoid differentiation remains to be assessed. Methods We formulated B8 with commercially available, animal-free growth factors, refined its composition to support normoxic culture of the widely-used WTC11 hiPSC line, and compared it to commercial Essential 8 (E8) and a home-made, weekend-free E8 formulation (hE8). We measured pluripotency marker expression and cell cycle with flow cytometry, and investigated the transcriptional profiles by bulk RNA sequencing. We also assessed the efficiency of gene editing, single-cell sorting, and cardiac differentiation in both monolayer and organoids. Results hE8 performed similarly to commercial E8 in all the assays. Despite morphological changes, cells in B8+, our optimised variant of B8, expressed the pluripotency marker NANOG at the highest level. At the same time, cells grown in B8+ were primed towards a mesendodermal fate. B8+ outperformed other media with regard to genome editing via homology directed recombination, and was on par with other media in other assays. Conclusions Overall, optimised weekend-free media formulations promise to democratise the generation of engineered cells for a wide range of applications.

Multi-omics approach for understanding the response of Bacteroides fragilis to carbapenems

BackgroundThe prevalence of Bacteroides fragilis isolates resistant to first-line beta-lactam drugs is increasing, resulting in reduced treatment efficacy. Investigating the bacterial transcriptome and proteome can uncover links between bacterial genes and resistance mechanisms. In this study, we experimentally assessed in vitro the transcriptional and proteomic profiles of B. fragilis exposed to SICs of meropenem, an effective antimicrobial agent, collected from patients with intra-abdominal diseases at Astana City Hospital, Kazakhstan. MethodsB. fragilis was cultured in brain heart infusion broth and sub-cultured every 48 h for 8 days in media with and without meropenem. Total RNA was extracted from the liquid cultures using a commercial RNeasy mini kit, and strand-specific RNA sequencing (RNA-seq) was performed on the DNBSEQ platform. Raw RNA-seq data were retrieved from BioProject No. PRJNA531645 and uploaded to the NCBI Sequence Read Archive (accession no. SRX22081155). Proteins of B. fragilis were extracted and separated using sodium dodecyl sulphate–polyacrylamide gel electrophoresis, followed by analysis of the eluted peptides using liquid chromatography–tandem mass spectrometry. Cluster analysis utilised the Database for Annotation, Visualization, and Integrated Discovery. ResultsThe subinhibitory concentration (SIC) of meropenem was determined to be 0.5 μg/L (minimum inhibitory concentration: 1). Mapping of reads to the reference genome identified 2,477 expressed genes in all B. fragilis BFR KZ01 samples. Ten differentially expressed genes (DEGs) were common across comparison groups during and post-antibiotic exposure (wMEM vs. MEM2 and MEM2 vs. rMEM8); however, no substantially enriched Gene Ontology terms were identified. The cluster analysis highlighted a significant enrichment cluster (W-0560 oxidoreductase) of DEGs following antibiotic withdrawal. In total, 859 B. fragilis proteins were identified, with the expressions of three proteins, 3-oxoacyl-[acyl carrier protein] reductase, acetyl-CoA carboxylase biotin carboxylase subunit, and beta-ketoacyl-ACP synthase III, being upregulated in the enriched protein folding category. Notably, chaperone proteins such as FKBP-type peptidyl-prolyl cis-trans isomerases (involved in the cis-trans isomerisation of prolyl peptide bonds) and GroES (a co-chaperone functioning with GroEL) were also identified. ConclusionsUnder the influence of low doses of antibiotics defense mechanisms are activated which contribute to the emergence of resistance. These results provide insight into the response of B. fragilis to meropenem exposure, mainly at the SIC, contributing to the understanding bacterial survival strategies under stress conditions.

Thymic epithelial organoids mediate T-cell development.

Although the advent of organoids has opened unprecedented perspectives for basic and translational research, immune system-related organoids remain largely underdeveloped. Here, we established organoids from the thymus, the lymphoid organ responsible for T-cell development. We identified conditions enabling mouse thymic epithelial progenitor cell proliferation and development into organoids with diverse cell populations and transcriptional profiles resembling in vivo thymic epithelial cells (TECs) more closely than traditional TEC cultures. In contrast to these two-dimensional cultures, thymic epithelial organoids maintained thymus functionality in vitro and mediated physiological T-cell development upon reaggregation with T-cell progenitors. The reaggregates showed in vivo-like epithelial diversity and the ability to attract T-cell progenitors. Thymic epithelial organoids are the first organoids originating from the stromal compartment of a lymphoid organ. They provide new opportunities to study TEC biology and T-cell development in vitro, paving the way for future thymic regeneration strategies in ageing or acute injuries.

A ferroptosis-associated prognostic model correlated with immune landscape and radiotherapy response in low-grade gliomas (LGGs)

Despite receiving comprehensive treatment, the prognosis for low-grade gliomas (LGGs) patients varies considerably. Recent studies have focused extensively on ferroptosis, across a range of tumor types. Nevertheless, methodologies to evaluate the efficacy of radiotherapy for LGGs, from the perspective of ferroptosis-related genes (FRGs), remain strikingly rare. In this study, we conducted a retrospective study on the transcriptional profiles of LGG patients from the public databases and a local cohort. An FRG model was developed and validated, exhibits heightened robustness when contrasted with the traditional ssGSEA model. Patients demonstrating higher FRG scores were identified as a high-risk group, displaying a worse prognosis. By incorporating the FRG score alongside other prognosis-associated clinical indicators, we formulated an enhanced nomogram to achieve a higher level of prediction performance. Additionally, among LGG patients receiving radiotherapy, a poorer prognosis was observed in the high-risk group. Further investigation revealed that samples from the high-risk group generally exhibit a TME in an immuno-suppressive state. Collectively, we developed an FRG model and a robust nomogram for LGG prognostication. This study suggests that a high FRG score, indicative of an immunosuppressive TME, could potentially lead to a less favorable prognosis for certain LGG patients receiving radiotherapy.

Advancing towards a unified eosinophil signature from transcriptional profiling.

Eosinophils are granulocytes that can accumulate in increased numbers in tissues and/or peripheral blood in disease. Phenotyping of eosinophils in health and disease has the potential to improve the precision of diagnosis and choice of therapies for eosinophilic-associated diseases. Transcriptional profiling of eosinophils has been plagued by cell fragility and difficulty isolating high quality RNA. With several technological advances, single-cell RNA sequencing (scRNA-seq) has become possible with eosinophils, at least from mice, while bulk RNA sequencing and microarrays have been performed in both murine and human samples. Anticipating more eosinophil transcriptional profiles in the coming years, we provide a summary of prior studies conducted on mouse and human eosinophils in blood and tissue, with a discussion of the advantages and potential pitfalls of various approaches. Common technical standards in studying eosinophil biology would help advance the field and make cross-study comparisons possible. Knowledge gaps and opportunities include identifying a minimal set of genes that define the eosinophil lineage, comparative studies between active disease and remission vs. homeostasis or development, especially in humans, and a comprehensive comparison between murine and human eosinophils at the transcriptional level. Characterizing such transcriptional patterns will be important to understanding the complex and diverse roles of eosinophils in both health and disease.

Pilot study: Understanding Canine Transmissible Venereal Tumor through its Transcriptional Profile

Canine transmissible venereal tumor (CTVT) is transmitted through the implantation of tumor cells. CTVT was the first tumor described with contagious characteristics and remains one of the few tumors with this capability. This study aimed to map the transcriptomic profile of CTVT to elucidate the potential mechanisms through which this tumor implants and evades host immune surveillance. For this study, 11 dogs aged ≥ 2 years diagnosed with CTVT were selected. Tumor biopsies were performed, RNA was extracted and converted into complementary DNA, followed by RT-qPCR analysis. The transcriptomic profile of CTVT revealed a wide array of differentially expressed genes. However, only the most relevant genes from an oncological perspective were discussed. IL-8, CXCL13, NCAM1, RNASEL, COROA1, and CBLB demonstrated potential associations with immune system evasion and transmission via implantation. Therefore, studying these genes may contribute to the development of targeted therapies that prevent contagion and immune evasion.

Modeling "Two-Hit" Severe Pneumonia in Mice: Pathological Characteristics and Mechanistic Studies.

Severe pneumonia is one of the most common critical diseases in clinical practice. Existing models for severe pneumonia have limitations, leading to limited clinical translation. In this study, a two-hit severe pneumonia mouse model was established by inducing primary pneumonia through intratracheal instillation of 800μg lipopolysaccharide (LPS), followed by intraperitoneal injection of 10mg/kg LPS. The effectiveness of various inflammatory indicators and the lung tissue damage during the time course of this model were confirmed and evaluated. At 3h post two-hit, the IL-6, TNF-α levels in peripheral blood and bronchoalveolar lavage fluid (BALF), and the white blood cells, neutrophils, and lymphocytes in BALF notably exhibited the most pronounced elevation. At 12h post two-hit, the white blood cells and neutrophils in peripheral blood significantly increased, accompanied by notable alterations in splenic immune cells and worsened pulmonary histopathological damage. Transcriptomics of lung tissue, microbiota analysis of lung and gut, as well as plasma metabolomics analyses further indicated changes in transcriptional profiles, microbial composition, and metabolites due to the two-hit modeling. These results validate that the two-hit model mimics the clinical presentation of severe pneumonia and serves as a robust experimental tool for studying the pathogenesis of severe pneumonia and developing and assessing treatment strategies.

Activation of ATF3 via the integrated stress response pathway regulates innate immune response to restrict Zika virus.

Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus that co-opts cellular mechanisms to support viral processes that can reprogram the host transcriptional profile. Such viral-directed transcriptional changes and the pro- or anti-viral outcomes remain understudied. We previously showed that ATF3, a stress-induced transcription factor, is significantly upregulated in ZIKV-infected mammalian cells, along with other cellular and immune response genes. We now define the intracellular pathway responsible for ATF3 activation and elucidate the impact of ATF3 expression on ZIKV infection. We show that during ZIKV infection, the integrated stress response pathway stimulates ATF3 which enhances the innate immune response to antagonize ZIKV infection. This study establishes a link between viral-induced stress response and transcriptional regulation of host defense pathways and thus expands our knowledge of virus-mediated transcriptional mechanisms and transcriptional control of interferon-stimulated genes during ZIKV infection.

Phase separation of phospho-HDAC6 drives aberrant chromatin architecture in triple-negative breast cancer.

How dysregulated liquid-liquid phase separation (LLPS) contributes to the oncogenesis of female triple-negative breast cancer (TNBC) remains unknown. Here we demonstrate that phosphorylated histone deacetylase 6 (phospho-HDAC6) forms LLPS condensates in the nuclei of TNBC cells that are essential for establishing aberrant chromatin architecture. The disordered N-terminal domain and phosphorylated residue of HDAC6 facilitate effective LLPS, whereas nuclear export regions exert a negative dominant effect. Through phase-separation-based screening, we identified Nexturastat A as a specific disruptor of phospho-HDAC6 condensates, which effectively suppresses tumor growth. Mechanistically, importin-β interacts with phospho-HDAC6, promoting its translocation to the nucleus, where 14-3-3θ mediates the condensate formation. Disruption of phospho-HDAC6 LLPS re-established chromatin compartments and topologically associating domain boundaries, leading to disturbed chromatin loops. The phospho-HDAC6-induced aberrant chromatin architecture affects chromatin accessibility, histone acetylation, RNA polymerase II elongation and transcriptional profiles in TNBC. This study demonstrates phospho-HDAC6 LLPS as an emerging mechanism underlying the dysregulation of chromatin architecture in TNBC.

Advances in spatial multi-omics in tumors.

Single-cell techniques have convincingly demonstrated that tumor tissue usually contains multiple genetically defined cell subclones with different gene mutation sets as well as various transcriptional profiles, but the spatial heterogeneity of the microenvironment and the macrobiological characteristics of the tumor ecosystem have not been described. For the past few years, spatial multi-omics technologies have revealed the cellular interactions, microenvironment, and even systemic tumor-host interactions in the tumor ecosystem at the spatial level, which can not only improve classical therapies such as surgery, radiotherapy, and chemotherapy but also promote the development of emerging targeted therapies in immunotherapy. Here, we review some emerging spatial omics techniques in cancer research and therapeutic applications and propose prospects for their future development.

Patient mutations in DRP1 perturb synaptic maturation of cortical neurons.

With the advent of exome sequencing, a growing number of children are being identified with de novo loss of function mutations in the dynamin 1 like ( DNM1L) gene encoding the large GTPase essential for mitochondrial fission, dynamin-related protein 1 (DRP1); these mutations result in severe neurodevelopmental phenotypes, such as developmental delay, optic atrophy, and epileptic encephalopathies. Though it is established that mitochondrial fission is an essential precursor to the rapidly changing metabolic needs of the developing cortex, it is not understood how identified mutations in different domains of DRP1 uniquely disrupt cortical development and synaptic maturation. We leveraged the power of induced pluripotent stem cells (iPSCs) harboring DRP1 mutations in either the GTPase or stalk domains to model early stages of cortical development in vitro . High-resolution time-lapse imaging of axonal transport in mutant DRP1 cortical neurons reveals mutation-specific changes in mitochondrial motility of severely hyperfused mitochondrial structures. Transcriptional profiling of mutant DRP1 cortical neurons during maturation also implicates mutation dependent alterations in synaptic development and calcium regulation gene expression. Disruptions in calcium dynamics were confirmed using live functional recordings of 100 DIV (days in vitro) mutant DRP1 cortical neurons. These findings and deficits in pre- and post-synaptic marker colocalization using super resolution microscopy, strongly suggest that altered mitochondrial morphology of DRP1 mutant neurons leads to pathogenic dysregulation of synaptic development and activity.

A single-cell atlas of spatial and temporal gene expression in the mouse cranial neural plate.

The formation of the mammalian brain requires regionalization and morphogenesis of the cranial neural plate, which transforms from an epithelial sheet into a closed tube that provides the structural foundation for neural patterning and circuit formation. Sonic hedgehog (SHH) signaling is important for cranial neural plate patterning and closure, but the transcriptional changes that give rise to the spatially regulated cell fates and behaviors that build the cranial neural tube have not been systematically analyzed. Here we used single-cell RNA sequencing to generate an atlas of gene expression at six consecutive stages of cranial neural tube closure in the mouse embryo. Ordering transcriptional profiles relative to the major axes of gene expression predicted spatially regulated expression of 870 genes along the anterior-posterior and mediolateral axes of the cranial neural plate and reproduced known expression patterns with over 85% accuracy. Single-cell RNA sequencing of embryos with activated SHH signaling revealed distinct SHH-regulated transcriptional programs in the developing forebrain, midbrain, and hindbrain, suggesting a complex interplay between anterior-posterior and mediolateral patterning systems. These results define a spatiotemporally resolved map of gene expression during cranial neural tube closure and provide a resource for investigating the transcriptional events that drive early mammalian brain development.

Individualized gray matter morphological abnormalities uncover two robust transdiagnostic biotypes

Psychiatric disorders exhibit a shared neuropathology, yet the diverse presentations among patients necessitate the identification of transdiagnostic subtypes to enhance diagnostic and treatment strategies. This study aims to unveil potential transdiagnostic subtypes based on personalized gray matter morphological abnormalities. A total of 496 patients with psychiatric disorders and 255 healthy controls (HCs) from three distinct datasets (one for discovery and two for validation) were enrolled. Individualized gray matter morphological abnormalities were determined using normative modeling to identify transdiagnostic subtypes. In the discovery dataset, two transdiagnostic subtypes with contrasting patterns of structural abnormalities compared to HCs were identified. Reproducibility and generalizability analyses demonstrated that these subtypes could be generalized to new patients and even to new disorders in the validation datasets. These subtypes were characterized by distinct disease epicenters. The gray matter abnormal pattern in subtype 1 was mainly linked to excitatory receptors, whereas subtype 2 showed a predominant association with inhibitory receptors. Furthermore, we observed that the gray matter abnormal pattern in subtype 2 was correlated with transcriptional profiles of inflammation-related genes, while subtype 1 did not show this association. Our findings reveal two robust transdiagnostic biotypes, offering novel insights into psychiatric nosology.

The Impact of Growing Area on the Expression of Fruit Traits Related to Sensory Perception in Two Tomato Cultivars.

Environmental conditions greatly influence the quality of tomato fruit by affecting the expression of genes, the abundance of metabolites, and the perception of sensorial attributes. In this study, a fruit transcriptome investigation, a sensory test, and a metabolomic analysis were performed to evaluate the impact of the environment on two popular tomato cultivars grown in two Italian regions. The transcriptional profile of each cultivar, cultivated in two different areas, highlighted differential expression in genes involved in pathways related to cell wall components such as pectin, lignin, and hemicellulose and sugars as well as in amino acids, phenylpropanoids, and pigment synthesis. The cultivation area mainly affects sensory attributes related to texture and flavor and the metabolic pattern of cell wall precursors, sugars, glutamate, aspartate, and carotenoids. In the two genotypes cultivated in the same environment, some attributes and fruit-related quality processes are similarly affected, while others are differently influenced based on the specific genetic makeup of the tomato. A combination of transcriptomic, sensory, and metabolomic data obtained from the two tomato genotypes revealed that the environment has a profound effect on specific sensory traits, providing information on factors that shape the specific characteristics and genetic targets for improving tomato fruit characteristics.

Scupa: Single-cell unified polarization assessment of immune cells using the single-cell foundation model.

Immune cells undergo cytokine-driven polarization in respond to diverse stimuli. This process significantly modulates their transcriptional profiles and functional states. Although single-cell RNA sequencing (scRNA-seq) has advanced our understanding of immune responses across various diseases or conditions, currently there lacks a method to systematically examine cytokine effects and immune cell polarization. To address this gap, we developed Single-cell unified polarization assessment (Scupa), the first computational method for comprehensive immune cell polarization analysis. Scupa is trained on data from the Immune Dictionary, which characterizes 66 cytokine-driven polarization states across 14 immune cell types. By leveraging the cell embeddings from the Universal Cell Embeddings model, Scupa effectively identifies polarized cells in new datasets generated from different species and experimental conditions. Applications of Scupa in independent datasets demonstrated its accuracy in classifying polarized cells and further revealed distinct polarization profiles in tumor-infiltrating myeloid cells across cancers. Scupa complements conventional single-cell data analysis by providing new insights into immune cell polarization, and it holds promise for assessing molecular effects or identifying therapeutic targets in cytokine-based therapies.

Characterization of a Novel Oxidative Stress Responsive Transcription Regulator in Mycobacterium bovis.

The antioxidant defense is critical for the survival of intracellular pathogens such as Mycobacterium tuberculosis complex (MTBC) species, including Mycobacterium bovis, which are often exposed to an oxidative environment caused by reactive oxygen species (ROS) in hosts. However, the signaling pathway in mycobacteria for sensing and responding to oxidative stress remains largely unclear. In this study, we characterize a TetR-type transcription regulator BCG_3893c, designated AotM, as a novel redox sensor in Mycobacterium bovis that increases mycobacterial tolerance to oxidative stress. AotM is required for the growth of M. bovis in the presence of 1 mM hydrogen peroxide. Loss of the aotM gene leads to altered transcriptional profiles with 352 genes significantly up-regulated and 25 genes significantly down-regulated. AotM recognizes a 14-bp palindrome sequence motif and negatively regulates the expression of a FAD-dependent oxidoreductase encoded by bcg_3892c. Overexpression of BCG_3892c increases intracellular ROS production and reduces the growth of M. bovis. In summary, we propose that AotM enhances the mycobacterial resistance against oxidative stress probably by inhibiting intracellular ROS production. Our findings reveal a novel underlying regulatory mechanism behind mycobacterial oxidative stress adaptation.

First Peek into the Transcriptomic Response in Heat-Stressed Tomato Inoculated with Septoglomus constrictum.

In this study, we report the interaction between an arbuscular mycorrhizal fungus, Septoglomus constrictum, and tomato plants under heat stress. For the first time, this interaction was studied by Illumina RNA-seq, followed by a comprehensive bioinformatic analysis that investigated root and leaf tissue samples. The genome-wide transcriptional profiling displayed fewer transcriptomic changes in the root under heat-stress conditions caused by S. constrictum. The top 50 DEGs suggested significant changes in the expression of genes encoding heat-shock proteins, transporter proteins, and genes of phytohormone metabolism involving jasmonic acid signalling. S. constrictum induced the upregulation of genes associated with pathways such as 'drought-responsive' and the 'development of root hair' in the root, as well as 'glycolipid desaturation', 'intracellular auxin transport', and 'ethylene biosynthesis' in the leaf. The pathways 'biotin biosynthesis' and 'threonine degradation' were found in both investigated tissue types. Expression analysis of transcription factors showed 2 and 11 upregulated transcription factors in heat-stressed root and leaf tissues, respectively. However, we did not find shared transcription factors. Heat-stressed arbuscular mycorrhizal plants suffered less oxidative stress when exposed to high temperatures. Colorimetric tests demonstrated less accumulation of H2O2 and MDA in heat-stressed mycorrhizal plants. This phenomenon was accompanied by the higher expression of six stress genes that encode peroxidases, glutathione S-transferase and ubiquitin carboxyl-terminal hydrolase in roots and leaves. Our findings provide a new perspective on elucidating the functional metabolic processes of tomato plants under mycorrhizal-heat stressed conditions.