Biology subjects

Martinez-Morales, J. R.

Publications and source records attributed to Martinez-Morales, J. R..

12 recordsLinked to original sources

bioRxiv2026

Evolutionary landscapes of zygotic genome activation across animals

During early animal embryogenesis, gene expression control shifts from maternally deposited products to newly transcribed RNA through zygotic genome activation (ZGA). Although essential and universal, ZGA remains poorly understood outside a few model species. Here, we generated a transcriptomic atlas of early embryogenesis across 61 animal species from 13 phyla and used a unified computational framework to infer ZGA timing systematically. We find extensive variation in ZGA onset across animals, but show that the ratio between genome size and egg cytoplasmic volume, a proxy for the nuclear-to-cytoplasmic ratio, robustly predicts when genome activation begins. We also reveal that zygotic genes also differ from maternal transcripts in genomic architecture, function and evolutionary conservation, suggesting that ZGA follows a conserved molecular logic despite flexible transcriptomic outputs.

evolutionary biology↗

bioRxiv2026

Single-cell multiomics identifies key nodes and cis-regulatory elements of the networks specifying the eye domains in zebrafish

Congenital eye malformations result from disruptions in the early developmental programs that specify and regionalize the eye primordium. Human genetics and developmental studies in animal models have identified critical regulatory nodes within the gene regulatory networks (GRNs) patterning the distinct eye domains. However, the fundamental cis-regulatory connectivity of these networks, and their dynamic behaviour, remains poorly understood. To address these gaps, we performed a comprehensive analysis of transcriptome dynamics and chromatin accessibility at single-cell resolution in the developing zebrafish forebrain. Our single-cell multiomic approach enabled us to characterize the regulatory landscape of the main eye derivatives: neural retina, optic stalk, pigmented epithelium and lens; alongside adjacent diencephalic and telencephalic regions. We identified and functionally validated tissue-specific cis-regulatory modules, and predicted highly connected nodes within each domain. Notably, we uncovered the role of foxp1b as a conserved central node within the neural retina network, highlighting the predictive power of our analysis in reconstructing the architecture of the GRNs that specify the vertebrate eye. TeaserIdentification of central nodes and cis-regulatory networks involved in eye patterning by single-cell multiomics in zebrafish.

developmental biology↗

bioRxiv2024

HUMAN SPINAL CORD ORGANOIDS REVEAL CELL INTERCALATION AS A CONSERVED MECHANISM FOR SECONDARY NEURULATION

Human pluripotent stem cells (hPSCs) have enabled major advances in neural organoid research, yet reconstructing spinal cord development in vitro remains challenging, as it requires mimicking the early environment of body axis elongation. Here, by exposing hPSCs to defined extrinsic signals, we directed their self-organizing capacity to generate organoids that recapitulate the transcriptional profile, cellular composition, and tissue architecture of the early human posterior spinal cord. Furthermore, we refined our culture system to more closely model the in vivo morphogenetic events of secondary neurulation. Using this approach, we identified cell intercalation--regulated by Yes-associated protein (YAP) activity--as a key morphogenetic mechanism driving de novo lumen formation and resolution. These biomimetic models provide a powerful platform to investigate the molecular and mechanical processes underlying human spinal cord development and offer new opportunities to elucidate the origins of neural tube defects, among the most common congenital defects.

developmental biology↗

bioRxiv2023

Comparative 3D genome analysis between neural retina and RPE reveals differential cis-regulatory interactions at retinal disease loci

Vision depends on the functional interplay between the photoreceptor cells of the neural retina and the supporting cells of the underlying retinal pigment epithelium (RPE). Most genes involved in inherited retinal diseases (IRD) display highly specific spatiotemporal expression within these interconnected retinal components through the local recruitment of cis-regulatory elements (CREs) in 3D nuclear space. To understand the role of differential chromatin architecture in establishing tissue-specific expression patterns at IRD loci in the human neural retina and the RPE, we mapped genome-wide chromatin interactions by applying in situ Hi-C and H3K4me3 HiChIP to human adult post-mortem donor retinas. A comparative 3D genome analysis between neural retina and RPE/choroid revealed that almost 60% of 290 known IRD genes were marked by differential 3D genome structure and/or cis-regulatory interactions. One of these genes was ABCA4, which is implicated in the most common autosomal recessive IRD. We zoomed in on tissue-specific chromatin interactions at the ABCA4 locus using high-resolution UMI-4C assays. Upon integration with bulk and single-cell epigenomic datasets and in vivo enhancer assays in zebrafish, we revealed tissue-specific CREs interacting with ABCA4. In summary, through extensive comparative 3D genome mapping, based on genome-wide (Hi-C), promoter-centric (HiChIP) and locus-specific (UMI-4C) assays of human neural retina and RPE, we have shown that gene regulation at key IRD loci is likely mediated by tissue-specific chromatin interactions. These findings do not only provide insight into tissue-specific regulatory landscapes of IRD genes, but also delineate the search space for non-coding genomic variation underlying unsolved IRD. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=75 SRC="FIGDIR/small/543842v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@fd0d7dorg.highwire.dtl.DTLVardef@180a5bforg.highwire.dtl.DTLVardef@101aec1org.highwire.dtl.DTLVardef@1781d1d_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2023

Evolutionary conservation of embryonic DNA methylome remodelling in distantly related teleost species

Methylation of cytosines in the CG context (mCG) is the most abundant DNA modification in vertebrates that plays crucial roles in cellular differentiation and identity. After fertilization, DNA methylation patterns inherited from parental gametes are remodelled into a state compatible with embryogenesis. In mammals, this is achieved through the global erasure and re-establishment of DNA methylation patterns. However, in non-mammalian vertebrates like zebrafish, no global erasure has been observed. To investigate the evolutionary conservation and divergence of DNA methylation remodelling in teleosts, we generated base resolution DNA methylome datasets of developing medaka and medaka-zebrafish hybrid embryos. In contrast to previous reports, we show that medaka display comparable DNA methylome dynamics to zebrafish with high gametic mCG levels (sperm: [~]90%; egg: [~]75%), and adoption of a paternal-like methylome during early embryogenesis, with no signs of prior DNA methylation erasure. We also demonstrate that non-canonical DNA methylation (mCH) reprogramming at TGCT tandem repeats is a conserved feature of teleost embryogenesis. Lastly, we find remarkable evolutionary conservation of DNA methylation remodelling patterns in medaka-zebrafish hybrids, indicative of compatible DNA methylation maintenance machinery in far-related teleost species. Overall, these results suggest strong evolutionary conservation of DNA methylation remodelling pathways in teleosts, which is distinct from the global DNA methylome erasure and reestablishment observed in mammals.

developmental biology↗

bioRxiv2022

Multi-omics profiling, in vitro and in vivo enhancer assays dissect the cis -regulatory mechanisms underlying North Carolina macular dystrophy, a retinal enhanceropathy

North Carolina macular dystrophy (NCMD) is a rare autosomal dominant disease affecting macular development. The disease is caused by non-coding single nucleotide variants (SNVs) in two hotspot regions near PRDM13 and by duplications in two distinct chromosomal loci, overlapping DNase I hypersensitive sites near either PRDM13 or IRX1. To unravel the mechanisms by which these variants cause disease, we first established a genome-wide multi-omics retinal database, RegRet. Integration of UMI-4C profiles we generated on adult human retina then allowed fine-mapping of the interactions of the PRDM13 and IRX1 gene promoters, and the identification of eighteen candidate cis-regulatory elements (cCREs), the activity of which was investigated by luciferase and Xenopus enhancer assays. Next, luciferase assays showed that the non-coding SNVs located in the two hotspot regions of PRDM13 affect cCRE activity, including two novel NCMD-associated non-coding SNVs that we identified. Interestingly, the cCRE containing one of these SNVs was shown to interact with the PRDM13 promoter, demonstrated in vivo activity in Xenopus, and is active at the developmental stage when progenitor cells of the central retina exit mitosis, putting forward this region as a PRDM13 enhancer. Finally, mining of single-cell transcriptional data of embryonic and adult retina revealed the highest expression of PRDM13 and IRX1 when amacrine cells start to synapse with retinal ganglion cells, supporting the hypothesis that altered PRDM13 or IRX1 expression impairs interactions between these cells during retinogenesis. Overall, this study gained insight into the cis-regulatory mechanisms of NCMD and supports that this condition is a retinal enhanceropathy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=149 SRC="FIGDIR/small/481329v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@4a85e3org.highwire.dtl.DTLVardef@9bfe55org.highwire.dtl.DTLVardef@156a9d8org.highwire.dtl.DTLVardef@a8cb92_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2022

A Yap-dependent transcriptional program directs cell migration for embryo axis assembly

The condensation of the embryo primary axis is a fundamental landmark in the establishment of the vertebrate body plan. Although the complex morphogenetic movements directing cell convergence towards the midline have been described extensively, little is known on how gastrulating cells interpret mechanical cues. Yap proteins are among the best characterized transcriptional mechanotransducers, yet their role in gastrulation has remained elusive. Here we show that the double knockout of yap and its paralog yap1b in medaka results in an axis assembly failure. Quantitative live imaging reveals that mutant cells display reduced displacement and migratory persistence. By characterizing the Yap-dependent transcriptional program, we identified genes involved in cytoskeletal organization and cell-ECM adhesion, rather than in germ layer specification, as direct Yap targets. Dynamic analysis of Tead sensors and downstream targets reveals Yap is acting in migratory cells, and not as a midline beacon, to direct gastrulating precursors trajectories by promoting cortical actin recruitment and focal adhesions assembly. We propose that Yap is engaged in a mechano-regulatory loop that is essential to maintain the directed cell migration sustaining embryo axis formation.

developmental biology↗

bioRxiv2022

Mutation of Vsx genes in zebrafish highlights the robustness of the retinal specification network.

Genetic studies in human and mice have established a dual role for Vsx genes in retina development: an early function in progenitors specification, and a later requirement for bipolar-cells fate determination. Despite their conserved expression patterns, it is currently unclear to which extent Vsx functions are also conserved across vertebrates, as mutant models are available only in mammals. To gain insight into vsx function in teleosts, we have generated vsx1 and vsx2 CRISPR-Cas9 double knockouts (vsxKO) in zebrafish. Our electrophysiological and histological analyses indicate severe visual impairment and bipolar cells depletion in vsxKO larvae, with retinal precursors being rerouted towards photoreceptor or Muller glia fates. Surprisingly, neural retina is properly specified and maintained in mutant embryos, which do not display microphthalmia. We show that although important cis-regulatory remodelling occurs in vsxKO retinas during early specification, this has little impact at a transcriptomic level. Our observations point to genetic redundancy as an important mechanism sustaining the integrity of the retinal specification network, and to Vsx genes regulatory weight varying substantially among vertebrate species. Brief Summary Statement for use in emailed and online tables of content alertsThe mutation of vsx genes in zebrafish confirms a conserved role in bipolar cells specification across vertebrates, but do not interfere with the specification of the neural retina domain. Our data reveal the unexpected robustness of the genetic network sustaining the identity of the neural retina.

developmental biology↗

bioRxiv2021

Trap-TRAP, a versatile tool for tissue-specific translatomics in zebrafish

Developmental and physiological processes depend on the transcriptional and translational activity of heterogeneous cell populations. A main challenge in gene expression studies is dealing with this intrinsic complexity while keeping sequencing efficiency. Translating ribosome affinity purification (TRAP) methods have allowed cell-specific recovery of polyribosome-associated RNAs by genetic tagging of ribosomes in selected cell populations. Here we combined the TRAP approach with adapted enhancer trap methods (trap-TRAP) to systematically generate zebrafish transgenic lines suitable for tissue-specific translatome interrogation. Through the random integration of the eGFP:rpl10a cassette, we have generated stable lines driving expression in a variety of tissues, including the retina, skeletal muscle, lateral line primordia, rhombomeres, or jaws. To increase the range of applications, a UAS:TRAP transgenic line compatible with available Gal4 lines was also generated and tested. The resulting collection of lines and applications constitutes a resource for the zebrafish community in developmental genetics, organ physiology and disease modelling.

developmental biology↗

bioRxiv2020

The Shh/Gli3 gene regulatory network precedes the origin of paired fins and reveals the deep homology between distal fins and digits.

One of the central problems of vertebrate evolution is understanding the relationship among the distal portions of fins and limbs. Lacking comparable morphological markers of these regions in fish and tetrapods, these relationships have remained uncertain for the past century and a half. Here we show that Gli3 functions in controlling the proliferative expansion of distal progenitors are shared among median and paired fins as well as tetrapod limbs. Mutant knockout gli3 fins in medaka (Oryzias latipes) form multiple radials and rays, in a pattern reminiscent of the polydactyly observed in Gli3 null mutant mice. In limbs, Gli3 controls both anterior-posterior patterning and cell proliferation, two processes that can be genetically uncoupled. In situ hybridization, quantification of proliferation markers, and analysis of regulatory regions reveal that in paired and median fins, gli3 plays a main role in controlling proliferation but not in patterning. Moreover, gli3 downregulation in shh mutant fins rescues fin loss in a manner similar to how Gli3-deficiency restores digits in the limbs of Shh mutant mouse embryos. We hypothesize that the Gli3/Shh pathway preceded the origin of paired appendages and was originally involved in modulating cell proliferation. Accordingly, the distal regions of median fins, paired fins, and limbs retain a deep regulatory and functional homology that predates the origin of paired appendages.

evolutionary biology↗

bioRxiv2020

Genetic Developmental Timing Revealed by Inter-Species Transplantations in Fish

The path from a fertilised egg to an embryo involves the coordinated formation of cell types, tissues and organs. Developmental modules (Raff, 1996) comprise discrete units specified by self-sufficient genetic programs that can interact among each other during embryogenesis. Here we took advantage of the different span of embryonic development between two far related teleosts, zebrafish (Danio rerio) and medaka (Oryzias latipes), of 3 and 9 days respectively, to explore modularity principles. We report that inter-species blastula transplantations result in the ectopic formation of a retina formed by donor cells -- a module. We show that the developmental time of the retina follows a genetic program: an ectopic zebrafish retina in medaka develops with zebrafish dynamics. Heterologous transplantation results in a temporal decoupling between the donor retina and host organism, illustrated by two paradigms that require retina-host interactions: lens recruitment and retino-tectal projections. Our results uncover a new experimental system to address temporal decoupling along embryonic development, and highlight the presence of largely autonomous but yet interconnected developmental modules orchestrating organogenesis.

developmental biology↗

bioRxiv2020

CRISPR-Cas13d induces efficient mRNA knock-down in animal embryos

Early embryonic development is driven exclusively by maternal gene products deposited into the oocyte. Although critical in establishing early developmental programs, maternal gene functions have remained elusive due to a paucity of techniques for their systematic disruption and assessment. CRISPR-Cas13 systems have recently been employed to induce RNA degradation in yeast, plants and mammalian cell lines. However, no systematic study of the potential of Cas13 has been carried out in an animal system. Here, we show that CRISPR-Cas13d is an effective and precise system to deplete specific mRNA transcripts in zebrafish embryos. We demonstrate that both zygotically-expressed and maternally-provided transcripts are efficiently targeted, resulting in an 80% average decrease in transcript level and the recapitulation of well-known embryonic phenotypes. Moreover, we show that this system can be used in medaka, killifish and mouse embryos. Altogether our results demonstrate that CRISPR-Cas13d is an efficient knock-down platform to interrogate gene function in animal embryos.

developmental biology↗