Genome-wide Transcription Research Articles

Genome-wide identification, transcript profiling and functional analyses of PCP gene family in Wucai (Brassica campestris)

Pollen coat proteins (PCPs) are cysteine-rich small-molecule proteins, which exhibit high levels of polymorphism and are expressed in gametocytes. Previous investigations have revealed that PCP genes are involved in pollen wall synthesis, pollen-stigma recognition, and pollen development and germination. However, gene expression and function of PCP family in pollen development is not well understood in Wucai (Brassica campestris L.). In this study, genome-wide identification and expression analysis of the BcPCP gene family members were conducted, including their physical and chemical properties, chromosome localization, phylogenetic relationships, gene structure, and tertiary structure. A total of 20 BcPCP genes were identified and classified into three subfamilies showing high homology to Arabidopsis thaliana. Expression pattern analysis indicated that the BcPCP gene family exhibits higher expression levels in reproductive organs, suggesting their potential involvement in the reproductive development. Notably, BraA02g002400.3 C, potentially associated with male sterility, was identified through multiple transcriptomic and proteomic datasets. Subsequently, sequence analysis revealed its homology with the Arabidopsis GRP20 gene, and thus it was named BcGRP20. Functional analysis of this gene showed that overexpression of BcGRP20 gene in the Arabidopsis grp20 mutant could restore anther fertility. Overall, our findings indicate that BcGRP20 plays a critical role in pollen development and may be the causative gene for male sterility in Wucai. This study provides candidate genes for further functional identification of BcPCP genes in Wucai, which are crucial for anther development.

KLF13 promotes SLE pathogenesis by modifying chromatin accessibility of key proinflammatory cytokine genes

Although significant progress has been achieved in elucidating the genetic architecture of systemic lupus erythematosus (SLE), identifying genes underlying the pathogenesis has been challenging. The NZM2410-derived lupus susceptibility Sle3 locus is associated with T cell hyperactivity and activated myeloid cells. However, candidate genes associated with these phenotypes have not been identified. Here, we narrow the Sle3 locus to a smaller genomic segment (Sle3k) and show that mice carrying Sle3k and Sle1 loci developed lupus nephritis. We identify Klf13 as the primary candidate gene that is associated with genome-wide transcription changes resulting in higher levels of proinflammatory cytokines, enhanced T cell activation, and hyperresponsiveness of myeloid cells. Correspondingly, Klf13–/– mice display repression of genes involved in mediating immune activation, including key proinflammatory cytokines/chemokines in T cells and dysregulation in cytokine signaling pathways in myeloid cells in response to toll receptor ligands. Klf13 upregulation is associated with increased production of RANTES, a key chemokine in lupus nephritis, in activated T cells and the kidneys of lupus-prone mice. In sum, our findings reveal Klf13 as a key gene in the Sle3 interval in mediating lupus pathogenesis that may have implications in the rational design of new therapies for SLE.

Genome-wide DNA methylation and mRNA transcription analysis revealed aberrant gene regulation pathways in dermatomyositis and polymyositis patients.

Genome-wide DNA methylation and mRNA transcription analysis revealed aberrant gene regulation pathways in dermatomyositis and polymyositis patients.

Integrating Microarray Data and Single-Cell RNA-Seq Reveals Key Gene Involved in Spermatogonia Stem Cell Aging.

The in vitro generation of spermatogonial stem cells (SSCs) from embryonic stem cells (ESCs) offers a viable approach for addressing male infertility. A multitude of molecules participate in this intricate process, which requires additional elucidation. Despite the decline in SSCs in aged testes, SSCs are deemed immortal since they can multiply for three years with repeated transplantation. Nonetheless, the examination of aging is challenging due to the limited quantity and absence of precise indicators. Using a microarray, we assessed genome-wide transcripts (about 55,000 transcripts) of fibroblasts and SSCs. The WGCNA approach was then used to look for SSC-specific transcription factors (TFs) and hub SSC-specific genes based on ATAC-seq, DNase-seq, RNA-seq, and microarray data from the GEO databases as well as gene expression data (RNA-seq and microarray data). The microarray analysis of three human cases with different SSCs revealed that 6 genes were upregulated, and the expression of 23 genes was downregulated compared to the normal case in relation to aging genes. To reach these results, online assessments of Enrich Shiny GO, STRING, and Cytoscape were used to forecast the molecular and functional connections of proteins before identifying the master routes. The biological process and molecular function keywords of cell-matrix adhesion, telomerase activity, and telomere cap complex were shown to be significantly altered in upregulated differentially expressed genes (DEGs) by the functional enrichment analysis. According to our preliminary research, cell-specific TFs and TF-mediated GRNs are involved in the creation of SSCs. In order to maximize the induction efficiency of ESC differentiation into SSCs in vitro, hub SSC-specific genes and important SSC-specific TFs were identified, and sophisticated network regulation was proposed. According to our research, these genes and the hub proteins that they interact with may be able to shine a light on the pathophysiologies of infertility and aberrant germ cells.

Stabilization of RRBP1 mRNA via an m6A-dependent manner in prostate cancer constitutes a therapeutic vulnerability amenable to small-peptide inhibition of METTL3

Mounting evidence has implicated the RNA m6A methylation catalyzed by METTL3 in a wide range of physiological and pathological processes, including tumorigenesis. The detailed m6A landscape and molecular mechanism of METTL3 in prostate cancer (PCa) remains ill-defined. We find that METTL3 is overexpressed in PCa and correlates with worse patient survival. Functional studies establish METTL3 as an oncoprotein dependent on its m6A enzymatic activity in both AR+ and AR− PCa cells. To dissect the regulatory network of m6A pathway in PCa, we map the m6A landscape in clinical tumor samples using m6A-seq and identify genome-wide METTL3-binding transcripts via RIP-seq. Mechanistically, we discover RRBP1 as a direct METTL3 target in which METTL3 stabilizes RRBP1 mRNA in an m6A-dependent manner. RRBP1 positively correlates with METTL3 expression in PCa cohorts and exerts an oncogenic role in aggressive PCa cells. Leveraging the 3D structural protein-protein interaction between METTL3 and METTL14, we successfully develop two potential METTL3 peptide inhibitors (RM3 and RSM3) that effectively suppress cancer cell proliferation in vitro and tumor growth in vivo. Collectively, our study reveals a novel METTL3/m6A/RRBP1 axis in enhancing aggressive traits of PCa, which can be therapeutically targeted by small-peptide METTL3 antagonists.

Differential gene expression and chromatin accessibility in the medial prefrontal cortex associated with individual differences in rat behavioral models of opioid use disorder.

Opioid use disorder (OUD) is a neuropsychological disease that has a devastating impact on public health. Substantial individual differences in vulnerability exist, the neurobiological substrates of which remain unclear. To address this question, we investigated genome-wide gene transcription (RNA-seq) and chromatin accessibility (ATAC-seq) in the medial prefrontal cortex (mPFC) of male and female rats exhibiting differential vulnerability in behavioral paradigms modeling different phases of OUD: Withdrawal-Induced Anhedonia (WIA), Demand, and Reinstatement. Ingenuity Pathway Analysis (IPA) of RNA-seq revealed greater changes in canonical pathways in Resilient (vs. Saline) rats in comparison to Vulnerable (vs. Saline) rats across 3 paradigms, suggesting brain adaptations that might contribute to resilience to OUD across its trajectory. Analyses of gene networks and upstream regulators implicated processes involved in oligodendrocyte maturation and myelination in WIA, neuroinflammation in Demand, and metabolism in Reinstatement. Motif analysis of ATAC-seq showed changes in chromatin accessibility to a small set of transcription factor (TF) binding sites as a function either of opioid exposure (i.e., morphine versus saline) generally or of individual vulnerability specifically. Some of these were shared across the 3 paradigms and others were unique to each. In conclusion, we have identified changes in biological pathways, TFs, and their binding motifs that vary with paradigm and OUD vulnerability. These findings point to the involvement of distinct transcriptional and epigenetic mechanisms in response to opioid exposure, vulnerability to OUD, and different stages of the disorder.

Post-transcriptional reprogramming by thousands of mRNA untranslated regions in trypanosomes

Although genome-wide polycistronic transcription places major emphasis on post-transcriptional controls in trypanosomatids, messenger RNA cis-regulatory untranslated regions (UTRs) have remained largely uncharacterised. Here, we describe a genome-scale massive parallel reporter assay coupled with 3’-UTR-seq profiling in the African trypanosome and identify thousands of regulatory UTRs. Increased translation efficiency was associated with dosage of adenine-rich poly-purine tracts (pPuTs). An independent assessment of native UTRs using machine learning based predictions confirmed the robust correspondence between pPuTs and positive control, as did an assessment of synthetic UTRs. Those 3’-UTRs associated with upregulated expression in bloodstream-stage cells were also enriched in uracil-rich poly-pyrimidine tracts, suggesting a mechanism for developmental activation through pPuT ‘unmasking’. Thus, we describe a cis-regulatory UTR sequence ‘code’ that underpins gene expression control in the context of a constitutively transcribed genome. We conclude that thousands of UTRs post-transcriptionally reprogram gene expression profiles in trypanosomes.

Single-nucleus multi-omics identifies shared and distinct pathways in Pick's and Alzheimer's disease.

The study of neurodegenerative diseases, particularly tauopathies like Pick's disease (PiD) and Alzheimer's disease (AD), offers insights into the underlying regulatory mechanisms. By investigating epigenomic variations in these conditions, we identified critical regulatory changes driving disease progression, revealing potential therapeutic targets. Our comparative analyses uncovered disease-enriched non-coding regions and genome-wide transcription factor (TF) binding differences, linking them to target genes. Notably, we identified a distal human-gained enhancer (HGE) associated with E3 ubiquitin ligase (UBE3A), highlighting disease-specific regulatory alterations. Additionally, fine-mapping of AD risk genes uncovered loci enriched in microglial enhancers and accessible in other cell types. Shared and distinct TF binding patterns were observed in neurons and glial cells across PiD and AD. We validated our findings using CRISPR to excise a predicted enhancer region in UBE3A and developed an interactive database (http://swaruplab.bio.uci.edu/scROAD) to visualize predicted single-cell TF occupancy and regulatory networks.

Prion-like domain mediated phase separation of ARID1A promotes oncogenic potential of Ewing’s sarcoma

Liquid-liquid phase separation (LLPS) facilitates the formation of membraneless organelles within cells, with implications in various biological processes and disease states. AT-rich interactive domain-containing protein 1A (ARID1A) is a chromatin remodeling factor frequently associated with cancer mutations, yet its functional mechanism remains largely unknown. Here, we find that ARID1A harbors a prion-like domain (PrLD), which facilitates the formation of liquid condensates through PrLD-mediated LLPS. The nuclear condensates formed by ARID1A LLPS are significantly elevated in Ewing’s sarcoma patient specimen. Disruption of ARID1A LLPS results in diminished proliferative and invasive abilities in Ewing’s sarcoma cells. Through genome-wide chromatin structure and transcription profiling, we identify that the ARID1A condensate localizes to EWS/FLI1 target enhancers and induces long-range chromatin architectural changes by forming functional chromatin remodeling hubs at oncogenic target genes. Collectively, our findings demonstrate that ARID1A promotes oncogenic potential through PrLD-mediated LLPS, offering a potential therapeutic approach for treating Ewing’s sarcoma.

Quantifying genome-wide transcription factor binding affinities for chromatin using BANC-seq.

Transcription factors (TFs) bind specific DNA sequences to regulate transcription. Apart from DNA sequences, local factors such as DNA accessibility and chromatin structure determine the affinity of a TF for any given locus. Including these factors when measuring TF-DNA affinities has proven difficult. To address this challenge, we recently developed a method called binding affinities in native chromatin by sequencing (BANC-seq). In BANC-seq, intact mammalian nuclei are incubated with a concentration range of epitope-tagged TF, followed by either chromatin immunoprecipitation or cleavage under target and release using nuclease with spike-in DNA. This allows determination of apparent dissociation constant (KdApp) values, defined by the concentration of TF at which half-maximum binding occurs, across the genome. Here we present a detailed stepwise protocol for BANC-seq, including downstream data analysis. In principle, any molecular biologist should be able to perform a BANC-seq experiment in as little as 1.5 d (excluding analysis). However, preprocessing and analysis of the sequencing data does require some experience in command-line shell and R programming.

Genome-wide transcription response of Staphylococcus epidermidis to heat shock and medically relevant glucose levels.

Skin serves as both barrier and interface between body and environment. Skin microbes are intermediaries evolved to respond, transduce, or act in response to changing environmental or physiological conditions. We quantified genome-wide changes in gene expression levels for one abundant skin commensal, Staphylococcus epidermidis, in response to an internal physiological signal, glucose levels, and an external environmental signal, temperature. We found 85 of 2,354 genes change up to ~34-fold in response to medically relevant changes in glucose concentration (0-17 mM; adj p ≤0.05). We observed carbon catabolite repression in response to a range of glucose spikes, as well as upregulation of genes involved in glucose utilization in response to persistent glucose. We observed 366 differentially expressed genes in response to a physiologically relevant change in temperature (37-45°C; adj p ≤ 0.05) and an S. epidermidis heat-shock response that mostly resembles the heat-shock response of related staphylococcal species. DNA motif analysis revealed CtsR and CIRCE operator sequences arranged in tandem upstream of dnaK and groESL operons. We identified and curated 38 glucose-responsive genes as candidate ON or OFF switches for use in controlling synthetic genetic systems. Such systems might be used to instrument the in-situ skin microbiome or help control microbes bioengineered to serve as embedded diagnostics, monitoring, or treatment platforms.

Genome-wide transcript expression analysis reveals major chickpea and lentil genes associated with plant branching.

The search for elite cultivars with better architecture has been a demand by farmers of the chickpea and lentil crops, which aims to systematize their mechanized planting and harvesting on a large scale. Therefore, the identification of genes associated with the regulation of the branching and architecture of these plants has currently gained great importance. Herein, this work aimed to gain insight into transcriptomic changes of two contrasting chickpea and lentil cultivars in terms of branching pattern (little versus highly branched cultivars). In addition, we aimed to identify candidate genes involved in the regulation of shoot branching that could be used as future targets for molecular breeding. The axillary and apical buds of chickpea cultivars Blanco lechoso and FLIP07-318C, and lentil cultivars Castellana and Campisi, considered as little and highly branched, respectively, were harvested. A total of 1,624 and 2,512 transcripts were identified as differentially expressed among different tissues and contrasting cultivars of chickpea and lentil, respectively. Several gene categories were significantly modulated such as cell cycle, DNA transcription, energy metabolism, hormonal biosynthesis and signaling, proteolysis, and vegetative development between apical and axillary tissues and contrasting cultivars of chickpea and lentil. Based on differential expression and branching-associated biological function, ten chickpea genes and seven lentil genes were considered the main players involved in differentially regulating the plant branching between contrasting cultivars. These collective data putatively revealed the general mechanism and high-effect genes associated with the regulation of branching in chickpea and lentil, which are potential targets for manipulation through genome editing and transgenesis aiming to improve plant architecture.

Genome-wide nucleosome and transcription factor responses to genetic perturbations reveal chromatin-mediated mechanisms of transcriptional regulation.

Epigenetic mechanisms contribute to gene regulation by altering chromatin accessibility through changes in transcription factor (TF) and nucleosome occupancy throughout the genome. Despite numerous studies focusing on changes in gene expression, the intricate chromatin-mediated regulatory code remains largely unexplored on a comprehensive scale. We address this by employing a factor-agnostic, reverse-genetics approach that uses MNase-seq to capture genome-wide TF and nucleosome occupancies in response to the individual deletion of 201 transcriptional regulators in Saccharomyces cerevisiae, thereby assaying nearly one million mutant-gene interactions. We develop a principled approach to identify and quantify chromatin changes genome-wide, observing differences in TF and nucleosome occupancy that recapitulate well-established pathways identified by gene expression data. We also discover distinct chromatin signatures associated with the up- and downregulation of genes, and use these signatures to reveal regulatory mechanisms previously unexplored in expression-based studies. Finally, we demonstrate that chromatin features are predictive of transcriptional activity and leverage these features to reconstruct chromatin-based transcriptional regulatory networks. Overall, these results illustrate the power of an approach combining genetic perturbation with high-resolution epigenomic profiling; the latter enables a close examination of the interplay between TFs and nucleosomes genome-wide, providing a deeper, more mechanistic understanding of the complex relationship between chromatin organization and transcription.

The survival, gene expression, and DNA methylation of Paralichthys olivaceus impacted by the decay of green tide and bacterial infection in both laboratory and field simulation experiments

The recurring appearance of Ulva prolifera green tides has become a pressing environmental issue, especially for marine transportation, tourism, and aquaculture in the stage of decomposition. An abundance of decaying U. prolifera leads to water acidification, hypoxia and pathogenic microorganism proliferation, threatening marine germplasm resources, particularly benthic organisms with weak escape ability. Epigenetic modification is considered to be one of the molecular mechanisms involved in the plastic adaptive response to environmental changes. However, few studies concerning the specific impact of decaying green tide on benthic animals at the epigenetic level. In this study, decomposing algal effluents of U. prolifera, sediments containing uncorrupted U. prolifera, pathogenic microorganism were considered as impact factors, to reveal the effect of decaying U. prolifera on marine economic benthic species, Paralichthys olivaceus, using both field and laboratory simulation experiments. Field simulation experiment showed higher mortality rates and serious histopathological damage than the laboratory simulation experiment. And both the decaying U. prolifera and the sediment containing U. prolifera were harmful to P. olivaceus. Genome-wide DNA methylation and transcription correlation analyses showed that the response of P. olivaceus to green tide stress and bacterial infection was mainly mediated by immune signaling pathways such as PI3K-Akt signaling pathway. DNA methylation regulates the expression of immune-related genes involved in the PI3K-Akt signaling pathway, which enables P. olivaceus to adapt to the adverse environmental stresses by resisting apoptosis. In summary, this research analyzed the potential role of P. olivaceus in decaying U. prolifera, which is of great significance for understanding the impact of decaying green tide on marine commercial fish and also provides some theoretical guidance for the proliferation and release of fish seedlings.

Genome-wide identification and transcription factor regulation of monolignol biosynthetic genes in Ginkgo biloba L.

BackgroundGinkgo biloba L. is a widely cultivated Mesozoic relict gymnosperm, and the wood is resource-rich. As a main component of plant cell walls, lignin serves for plant water transport and mechanical support, as well as directly defining the properties of wood-based products. The lignin biosynthesis pathway in gymnosperms is simpler than that in angiosperms, but has been poorly studied due to the lack of model plants. ResultsIn the present study, we systematically identified a total of 157 monolignol biosynthetic genes from 10 families in G. biloba, including 12 Gb4CLs, 34 GbC3Hs, 7 GbC4Hs, 34 GbCADs, 8 GbCCoAOMTs, 19 GbCCRs, 13 GbCOMTs, 14 GbCSEs, 11 GbHCTs and 5 GbPALs. The chromosome localization, gene structure, gene promoter and other analyses were also conducted. We established a regulatory network between the identified monolignol biosynthetic genes and TFs generated by TGMI algorithm using 133 public RNA-seq datasets of various G. biloba tissues. We then examined 2 gene pairs from the network of their regulatory relationship via yeast one-hybrid and dual-luciferase reporter assay, and the results were positive confirming interactions existing between the GbMYB-025 protein and GbCAD31 promoter, GbERF-174 protein and GbCSE13 promoter in vivo. ConclusionsOur results provided a new effective approach to find transcriptional regulatory relationships and established an important foundation for molecular mechanisms and genetic regulation of monolignol biosynthesis in G. biloba.

Novel oncogenic transcriptional targets of mutant p53 in esophageal squamous cell carcinoma.

Missense mutations in the DNA binding domain of p53 are observed frequently in esophageal squamous cell carcinoma (ESCC). Recent studies have revealed the potentially oncogenic transcriptional networks regulated by mutant p53 proteins. However, majority of these studies have focused on common "hotspot" p53 mutations while rarer mutations are poorly characterized. In this study, we report the characterization of rare, "non-hotspot" p53 mutations from ESCC. In vitro tumorigenic assays performed following ectopic-expression of certain "non-hotspot" mutant p53 proteins caused enhancement of oncogenic properties in squamous carcinoma cell lines. Genome-wide transcript profiling of ESCC tumor samples stratified for p53 status, revealed several genes exhibiting elevated transcript levels in tumors harboring mutant p53. Of these, ARF6, C1QBP, and TRIM23 were studied further. Reverse transcription-quantitative PCR (RT-qPCR) performed on RNA isolated from ESCC tumors revealed significant correlation of TP53 transcript levels with those of the three target genes. Ectopic expression of wild-type and several mutant p53 forms followed by RT-qPCR, chromatin affinity-purification (ChAP), and promoter-luciferase assays indicated the exclusive recruitment of p53 mutants-P190T and P278L, to the target genes leading to the activation of expression. Several functional assays following knockdown of the target genes revealed a significant suppression of tumorigenicity in squamous carcinoma cell lines. Rescue experiments confirmed the specificity of the knockdown. The tumorigenic effects of the genes were confirmed in nude mice xenograft assays. This study has therefore identified novel oncogenic targets of "non-hotspot" mutant p53 proteins relevant for ESCC besides validating the functional heterogeneity of the spectrum of tumor-specific p53 mutations.

DDIT4L regulates mitochondrial and innate immune activities in early life.

Pattern recognition receptor responses are profoundly attenuated before the third trimester of gestation in the relatively low-oxygen human fetal environment. However, the mechanisms regulating these responses are uncharacterized. Herein, genome-wide transcription and functional metabolic experiments in primary neonatal monocytes linked the negative mTOR regulator DDIT4L to metabolic stress, cellular bioenergetics, and innate immune activity. Using genetically engineered monocytic U937 cells, we confirmed that DDIT4L overexpression altered mitochondrial dynamics, suppressing their activity, and blunted LPS-induced cytokine responses. We also showed that monocyte mitochondrial function is more restrictive in earlier gestation, resembling the phenotype of DDIT4L-overexpressing U937 cells. Gene expression analyses in neonatal granulocytes and lung macrophages in preterm infants confirmed upregulation of the DDIT4L gene in the early postnatal period and also suggested a potential protective role against inflammation-associated chronic neonatal lung disease. Taken together, these data show that DDIT4L regulates mitochondrial activity and provide what we believe to be the first direct evidence for its potential role supressing innate immune activity in myeloid cells during development.

Morphological change and genome-wide transcript analysis of Haloxylon ammodendron leaf development reveals morphological characteristics and genes associated with the different C3 and C4 photosynthetic metabolic pathways.

C4 photosynthesis outperforms C3 photosynthesis in natural ecosystems by maintaining a high photosynthetic rate and affording higher water-use and nitrogen-use efficiencies. C4 plants can survive in environments with poor living conditions, such as high temperatures and arid regions, and will be crucial to ecological and agricultural security in the face of global climate change in the future. However, the genetic architecture of C4 photosynthesis remains largely unclear, especially the genetic regulation of C4 Kranz anatomy. Haloxylon ammodendron is an important afforestation tree species and a valuable C4 wood plant in the desert region. The unique characteristic of H. ammodendron is that, during the seedling stage, it utilizes C3 photosynthesis, while in mature assimilating shoots (maAS), it switches to the C4 pathway. This makes an exceptional opportunity for studying the development of the C4 Kranz anatomy and metabolic pathways within individual plants (identical genome). To provide broader insight into the regulation of Kranz anatomy and non-Kranz leaves of the C4 plant H. ammodendron, carbon isotope values, anatomical sections and transcriptome analyses were used to better understand the molecular and cellular processes related to the development of C4 Kranz anatomy. This study revealed that H. ammodendron conducts C3 in the cotyledon before it switches to C4 in AS. However, the switching requires a developmental process. Stable carbon isotope discrimination measurements on three different developmental stages showed that young AS have a C3-like δ13C even though C4 Kranz anatomy is found, which is inconsistent with the anatomical findings. A C4-like δ13C can be measured in AS until they are mature. The expression analysis of C4 key genes also showed that the maAS exhibited higher expression than the young AS. In addition, many genes that may be related to the development of Kranz anatomy were screened. Comparison of gene expression patterns with respect to anatomy during leaf ontogeny provided insight into the genetic features of Kranz anatomy. This study helps with our understanding of the development of Kranz anatomy and provides future directions for studies on key C4 regulatory genes.

Genome-wide transcript analysis of the root tip of 7-day-old seedlings treated with or without HSAL for 12 h using RNA-seq (Oryza sativa L. japonica)

Genome-wide transcript analysis of the root tip of 7-day-old seedlings treated with or without HSAL for 12 h using RNA-seq (Oryza sativa L. japonica)

Structure-primed embedding on the transcription factor manifold enables transparent model architectures for gene regulatory network and latent activity inference

BackgroundModeling of gene regulatory networks (GRNs) is limited due to a lack of direct measurements of genome-wide transcription factor activity (TFA) making it difficult to separate covariance and regulatory interactions. Inference of regulatory interactions and TFA requires aggregation of complementary evidence. Estimating TFA explicitly is problematic as it disconnects GRN inference and TFA estimation and is unable to account for, for example, contextual transcription factor-transcription factor interactions, and other higher order features. Deep-learning offers a potential solution, as it can model complex interactions and higher-order latent features, although does not provide interpretable models and latent features.ResultsWe propose a novel autoencoder-based framework, StrUcture Primed Inference of Regulation using latent Factor ACTivity (SupirFactor) for modeling, and a metric, explained relative variance (ERV), for interpretation of GRNs. We evaluate SupirFactor with ERV in a wide set of contexts. Compared to current state-of-the-art GRN inference methods, SupirFactor performs favorably. We evaluate latent feature activity as an estimate of TFA and biological function in S. cerevisiae as well as in peripheral blood mononuclear cells (PBMC).ConclusionHere we present a framework for structure-primed inference and interpretation of GRNs, SupirFactor, demonstrating interpretability using ERV in multiple biological and experimental settings. SupirFactor enables TFA estimation and pathway analysis using latent factor activity, demonstrated here on two large-scale single-cell datasets, modeling S. cerevisiae and PBMC. We find that the SupirFactor model facilitates biological analysis acquiring novel functional and regulatory insight.