Biology subjects

Bobola, N.

Publications and source records attributed to Bobola, N..

12 recordsLinked to original sources

bioRxiv2024

The level of HAND1 controls the specification of multipotent cardiac and extraembryonic progenitors

Diverse sets of progenitors contribute to the development of the embryonic heart, but the mechanisms of their specification have remained elusive. Here, using a human pluripotent stem cell (hPSC) model, we deciphered cardiac and non-cardiac lineage trajectories in differentiation and identified transcription factors underpinning cell specification, identity and function. We discovered a concentration-dependent, fate determining function in mesodermal progenitors for the basic helix-loop-helix transcription factor HAND1 and uncovered its gene regulatory network. At low level, HAND1 directs differentiation towards multipotent juxta-cardiac field progenitors able to make cardiomyocytes and epicardial cells, whereas at high level it promotes the development of extraembryonic mesoderm. Importantly, HAND1-low progenitors can be propagated in their multipotent state. This detailed mechanistic insight into human development will accelerate the delivery of effective disease modelling, including for congenital heart disease, and cell therapy-based regenerative medicine.

developmental biology↗

bioRxiv2024

MicroRNA miR-92b-3p regulation of a cardiovascular gene regulatory network in the developing branchial arches

Vertebrate branchial arches (BAs) are a developmental paradigm, undergoing coordinated differentiation and morphogenesis to form various adult derivative tissues. MicroRNAs can strengthen gene regulatory networks (GRNs) to promote developmental stability. To interrogate the contribution of microRNAs to BA development, we generated a novel microRNA-sequencing dataset from mouse BAs. We identified 550 expressed microRNAs, of which approximately 20% demonstrate significant differential expression across BA domains. The three most posterior BAs and the connecting outflow tract (PBA/OFT) are enriched in biological processes linked to cardiovascular development. We identified enriched predicted microRNA-target interactions with PBA/OFT upregulated cardiovascular genes and validated transcripts encoding for two fundamental cardiac transcription factors (TFs), Gata6 and Tbx20, as targets of miR-92b-3p. Furthermore, we demonstrated that miR-92b-3p can downregulate endogenous GATA6 and TBX20 in human embryonic stem cells (hESCs) undergoing cardiomyocyte differentiation, consistent with conservation of these microRNA-target interactions in a cardiogenic setting. miR-92b-3p has previously been shown to target two other cardiac TFs, Hand2 and Mef2D. Therefore, we hypothesise that miR-92b-3p acts to stabilise cardiovascular GRNs during PBA/OFT development, through acting in multiple microRNA-mediated coherent feedforward loops.

developmental biology↗

bioRxiv2024

The evolutionary ancient MEIS transcription factors actuate lineage-specific transcription to establish cardiac fate

Control of gene expression is commonly mediated by distinct combinations of transcription factors (TFs). This cooperative action allows multiple biological signals to be integrated at specific regulatory elements, resulting in highly specific gene expression patterns in space and time. It is unclear whether combinatorial binding is also necessary to bring together TFs with distinct biochemical functions, which collaborate to effectively recruit and activate RNA polymerase II. Using a cardiac differentiation model, we find that the largely ubiquitous, evolutionary ancient homeodomain proteins MEIS are essential for activating a cardiac-specific gene expression program. MEIS TFs act as actuators, fully activating transcriptional programs selected by lineage-restricted TFs to drive the dynamic progression of cardiac differentiation. Combinatorial binding of MEIS with lineage-enriched TFs, GATA and HOX, provides selectivity, guiding MEIS to function at cardiac-specific enhancers. In turn, MEIS TFs promote accumulation of the methyltransferase KMT2D to initiate lineage-specific enhancer commissioning. MEIS combinatorial binding dynamics, dictated by the changing dosage of its partners, drive cells into progressive stages of cardiac differentiation. Our results uncover tissue-specific transcriptional activation as the result of ubiquitous actuator TFs harnessing general transcriptional coactivators at tissue-specific enhancers, to which they are directed by binding with lineage- and domain-specific TFs.

developmental biology↗

bioRxiv2023

A pipeline to identify TF combinatorial binding uncovers TEAD1 as an antagonist of tissue-specific transcription factors in human organogenesis

Gene expression is largely controlled by transcription factors (TFs), which bind to enhancers in combination with other TFs in a mechanism known as combinatorial binding. While combinatorial binding is well established, a comprehensive view of tissue-specific TF combinations at active enhancers during human embryonic development is still lacking. Using a two-step pipeline to detect co-occurring TF motifs in developmental enhancers across 11 human embryonic tissues, we found that motifs recognized by ubiquitous TF families, including TEAD, TALE, ETS, and STAT, are enriched near tissue-specific sequence signatures in developmental enhancers across multiple tissues. In human heart enhancers, TEAD and GATA motifs frequently co-occur, and in the developing mouse heart TEAD1 and GATA4 co-occupy a set of genomic regions, which are also preferentially bound by CHD4, a component of the NuRD complex involved in transcriptional repression. Consistently, TEAD1 attenuates enhancer activation in vitro, with this repressive effect dependent on tissue-specific activators. Overall, our findings reveal universal patterns of TF connectivity within organ-specific transcriptional networks and highlight a broad, previously unrecognized role for TEAD in coordinating organ growth and differentiation across multiple tissues.

genomics↗

bioRxiv2023

A spatial map of human liver cirrhosis reveals the patho-architecture and gene signatures associated with cell state transitions during liver disease

Liver fibrosis is a major cause of death worldwide. As a progressive step in chronic liver disease, fibrosis is almost always diagnosed too late with limited treatment options. Here, we uncover the spatial transcriptional landscape driving human liver fibrosis using single nuclei RNA and Assay for Transposase-Accessible Chromatin (ATAC) sequencing to deconvolute multi-cell spatial transcriptomic profiling in human liver cirrhosis. Through multi-modal data integration, we define molecular signatures driving cell state transitions in liver disease and define an impaired cellular response and directional trajectory from hepatocytes to cholangiocytes associated with disease remodelling. We identify pro-fibrogenic signatures in non-parenchymal cell subpopulations co-localised within the fibrotic niche and localise transitional cell states at the scar interface. This combined approach provides a spatial atlas of gene regulation and defines molecular signatures associated liver disease for targeted therapeutics or as early diagnostic markers of progressive liver disease.

genomics↗

bioRxiv2023

A cell atlas of the human outflow tract of the heart and its adult derivatives

The outflow tract (OFT) of the heart carries blood away from the heart into the great arteries. During embryogenesis, the OFT divides to form the aorta and pulmonary trunk, creating the double circulation present in mammals. Defects in this area account for one-third of all congenital heart defect cases. Here, we present comprehensive transcriptomic data on the developing OFT at two distinct timepoints (embryonic and fetal) and its adult derivatives, the aortic valves, and use spatial transcriptomics to define the distribution of cell populations. We uncover that distinctive embryonic signatures persist in adult cells and can be used as labels to retrospectively attribute relationships between cells separated by a large time scale. Single-cell regulatory network inference identifies GATA6, a transcription factor linked to common arterial trunk and bicuspid aortic valve, as a key regulator of valve precursor cells. Its downstream network reveals candidate drivers of human cardiac defects and illuminates the molecular mechanisms of both normal and pathological valve development. Our findings define the cellular and molecular signatures of the human OFT and its distinct cell lineages, which is critical for understanding congenital heart defects and developing cardiac tissue for regenerative medicine.

developmental biology↗

bioRxiv2022

Motif2Site: a Bioconductor package to detect accurate transcription factor binding sites from ChIP-seq

AO_SCPLOWBSTRACTC_SCPLOWTranscription factors (TFs) binding are in the core of the Regulatory networks studies. ChIP-seq experiments are available for many TFs in various species. As TFs co-bind in cis-regulatory elements regions to control gene expression, studying the existing relation among co-binding TFs such as distance of binding sites or co-occupancy are highly important to understand the regulatory mechanisms. Currently, to detect binding sites of each TF in cis-regulatory elements, first binding regions of each TF are detected by standard peak calling methods, and at the second step the best candidate binding sites are prioritized by motif detection methods in binding regions. However, it is well-known that the best prioritized candidate motifs are not necessarily the actual binding sites of TFs. Furthermore, motif prioritizing methods that consider more genomic features complexities of TFs bindings are usually computationally expensive methods. Here, we tend to improve the TF binding sites accuracy detection by using the original ChIP-seq signal. The motifs which are located closer to the summits of binding region peaks are more likely to be the actual binding sites. Therefore, We developed a novel post-processing Bioconductor package called Motif2Site to detect TFs binding sites from user provided motif sets and recenter them across experiments. We applied Motif2Site method to detect TF binding sites for major mouse embryonic stem cell (mESC) as well as mouse fetal and birth time (P0) heart TFs. Motif2Site could detect binding regions with comparable accuracy to the existing state-of-the-art while it substantially increased the accuracy of the detected binding sites. Motif2Site could future improve the accuracy of binding sites prediction by recentering binding sites across developmental conditions (fetal/P0 heart) and across homologous TFs (ex. GATA4/GATA6 and MEF2A/MEF2C). Purifying high-confidence binding sites in mouse fetal heart, enabled us to study the co-binding properties of TFs in cis-regulatory elements. We could also traced TFs footprints in selected heart-specific VISTA enhancers chromatin accessible regions.

bioinformatics↗

bioRxiv2022

A spatio-temporally constrained gene regulatory network directed by PBX1/2 acquires limb patterning specificity via HAND2

During development cell fates are specified by tightly controlled gene expression programs. PBX TALE transcription factors control gene regulatory networks (GRN) that direct vertebrate tissue patterning and organ morphogenesis. How PBX1/2 proteins acquire context-specific functions, despite widespread embryonic expression of Pbx1/2, remains elusive. In mouse limb buds, mesenchymal-specific loss of PBX1/2 or of the transcriptional regulator HAND2 results in similar phenotypes, suggesting that PBX1/2- and HAND2-dependent programs converge to control limb development. To investigate this scenario, we combined tissue-specific and temporally-controlled mutagenesis with multi-omics approaches using the murine hindlimb bud as a model. We reconstructed a GRN collaboratively directed by PBX1/2 and HAND2, demonstrating that Pbx1-Hand2 genetically interact in vivo during hindlimb patterning, with PBX1 concomitantly acting as an upstream regulator of Hand2. At organismal-level resolution the GRN is active within restricted subsets of posterior-proximal hindlimb mesenchymal cells, wherein Pbx1/2 and Hand2 are co-expressed with their target genes. Genome-wide profiling of PBX1 binding across multiple tissues further revealed that HAND2 selects a subset of PBX-bound regions to impart limb patterning functionality. This research elucidates mechanisms underlying limb bud-specific functions by PBX1/2, while informing general principles by which promiscuous transcription factors cooperate with select cofactors to instruct distinct developmental programs.

developmental biology↗

bioRxiv2019

HOX paralogs selectively convert binding of ubiquitous transcription factors into tissue-specific patterns of enhancer activation

Gene expression programs determine cell fate in embryonic development and their dysregulation results in disease. Transcription factors (TFs) control gene expression by binding to enhancers, but how TFs select and activate their target enhancers is still unclear. HOX TFs share conserved homeodomains with highly similar sequence recognition properties, yet they impart the identity of different animal body parts. To understand how HOX TFs control their specific transcriptional programs in vivo, we compared HOXA2 and HOXA3 binding profiles in the mouse embryo. HOXA2 and HOXA3 directly cooperate with TALE TFs and selectively target different subsets of a broad TALE chromatin platform. Binding of HOX and tissue-specific TFs convert low affinity TALE binding into high confidence, tissue-specific binding events, which bear the mark of active enhancers. We propose that HOX paralogs, alone and in combination with tissue-specific TFs, generate tissue-specific transcriptional outputs by modulating the activity of TALE TFs at selected enhancers.

developmental biology↗

bioRxiv2019

TALE and NF-Y co-occupancy marks enhancers of developmental control genes during zygotic genome activation in zebrafish

Animal embryogenesis is initiated by maternal factors, but zygotic genome activation (ZGA) shifts control to the embryo at early blastula stages. ZGA is thought to be mediated by specialized maternally deposited transcription factors (TFs), but here we demonstrate that NF-Y and TALE - TFs with known later roles in embryogenesis - co-occupy unique genomic elements at zebrafish ZGA. We show that these elements are selectively associated with early-expressed genes involved in transcriptional regulation and possess enhancer activity in vivo. In contrast, we find that elements individually occupied by either NF-Y or TALE are associated with genes acting later in development - such that NF-Y controls a cilia gene expression program while TALE TFs control expression of hox genes. We conclude that NF-Y and TALE have a shared role at ZGA, but separate roles later during development, demonstrating that combinations of known TFs can regulate subsets of key developmental genes at vertebrate ZGA.

developmental biology↗

bioRxiv2019

The epigenomic landscape regulating organogenesis in human embryos linked to developmental disorders

How the genome activates or silences transcriptional programmes governs organ formation. Little is known in human embryos undermining our ability to benchmark the fidelity of in vitro stem cell differentiation or cell programming, or interpret the pathogenicity of noncoding variation. Here, we studied histone modifications across thirteen tissues during human organogenesis. We integrated the data with transcription to build the first overview of how the human genome differentially regulates alternative organ fates including by repression. Promoters from nearly 20,000 genes partitioned into discrete states without showing bivalency. Key developmental gene sets were actively repressed outside of the appropriate organ. Candidate enhancers, functional in zebrafish, allowed imputation of tissue-specific and shared patterns of transcription factor binding. Overlaying more than 700 noncoding mutations from patients with developmental disorders allowed correlation to unanticipated target genes. Taken together, the data provide a new, comprehensive genomic framework for investigating normal and abnormal human development.

genomics↗

bioRxiv2019

Uncovering tissue-specific binding features from differential deep learning

MotivationTranscription factors (TFs) can bind DNA in a cooperative manner, enabling a mutual increase in occupancy. Through this type of interaction, alternative binding sites can be preferentially bound in different tissues to regulate tissue-specific expression programmes. Recently, deep learning models have become state-of-the-art in various pattern analysis tasks, including applications in the field of genomics. We therefore investigate the application of convolutional neural network (CNN) models to the discovery of sequence features determining cooperative and differential TF binding across tissues. ResultsWe analyse ChIP-seq data from MEIS, TFs which are broadly expressed across mouse branchial arches, and HOXA2, which is expressed in the second and more posterior branchial arches. By developing models predictive of MEIS differential binding in all three tissues we are able to accurately predict HOXA2 co-binding sites. We evaluate transfer-like and multitask approaches to regularising the high-dimensional classification task with a larger regression dataset, allowing for creation of deeper and more accurate models. We test the performance of perturbation and gradient-based attribution methods in identifying the HOXA2 sites from differential MEIS data. Our results show that deep regularised models significantly outperform shallow CNNs as well as k-mer methods in the discovery of tissue-specific sites bound in vivo. AvailabilityFor implementation and models please visit https://doi.org/10.5281/zenodo.2635463.

bioinformatics↗