Biology subjects

Rocha-Martins, M.

Publications and source records attributed to Rocha-Martins, M..

7 recordsLinked to original sources

bioRxiv2026

Apoptosis-induced self-renewal of neurogenic progenitors safeguards retinal development against extensive cell loss

Developing embryos have a striking ability to buffer external and internal perturbations. A key example of this phenomenon across developing systems is successful organ formation even after substantial cell loss. Although molecular regulators of this robustness are beginning to be understood, it remains unclear whether and how surviving cells can remodel their developmental trajectories to safeguard morphogenesis. To address this question, we use the zebrafish retina, where progenitor behaviour and lineage transitions are highly tractable and extensively characterized. We induce widespread apoptosis via heat stress and genetic approaches and track cellular and tissue-wide responses in vivo over time. We find that retinal development is highly resilient with neurogenesis initiating on time and growth continuing despite extensive apoptosis. Continued growth is supported by neurogenic progenitors that react to apoptosis through a non-cell autonomous switch in behaviour. These cells bypass their canonical differentiation route and undergo self-renewing divisions that expand clonal output and compensate for lost cells. Importantly, self-renewal is transient, progenitors resume lineage progression, generating appropriate neuronal cell types. This adaptive response supports the formation of retinas with proper architecture, connectivity to the brain and visual function. Together, these findings identify latent plasticity in the neurogenic programme as a mechanism that contributes to developmental robustness under stress.

developmental biology↗

bioRxiv2024

Pou4f2/Brn3b Overexpression Promotes The Genesis Of Retinal Ganglion Cell-Like Projection Neurons From Late Progenitors

Retinal ganglion cells (RGCs) are the projection neurons of the retina. In early retinal progenitor cells (RPCs), Atoh7 orchestrates the developmental RGC program and regulates the expression of critical downstream targets, including Pou4f factors. The absence of Pou4f2 or more POU4F family genes results in defects in RGC differentiation, aberrant axonal elaboration and ultimately RGC death, confirming the requirement of POU4F factors for RGC development and survival, with a critical role in regulating RGC axon outgrowth and pathfinding. Here, we investigated in vivo whether ectopic Pou4f2 expression in late retinal progenitor cells (late RPCs) is sufficient to induce the generation of cells with RGC properties, including projecting axons to the brain. Using a strong ubiquitous promoter to induce Pou4f2 overexpression in neonates, we observed a change in targeted cell distribution in the retinal tissue, including the presence of cells in the ganglion cell layer and inner plexiform layer with high density of GFP+ processes along the retina. Similar results on the induction of neuron processes were obtained when we overexpressed Pou4f2 in Atoh7 knockout mice, suggesting that POU4F2 is sufficient to induce them. Single-cell RNA sequencing (scRNA-seq) analysis shows that several RGC-genes (such as Rbpms, Gap-43, Hs6st3, and Foxp2) are upregulated after Pou4f2 overexpression. Additionally, gene ontology analysis indicates the induction of genes related to axonogenesis and neuronal differentiation. Imaging throughout the visual pathway revealed high density of axons projecting toward the optic nerve head and extending to brain regions, such as the superior colliculus and geniculate nucleus. Thus, Pou4f2 induced neurons with specific RGC characteristics that share similarities with resident RGCs and notably project axons that reach brain targets. In conclusion, these results showed that POU4F2 alone was sufficient to promote critical properties of projection neurons from retinal progenitors outside their developmental window.

neuroscience↗

bioRxiv2022

Deterministic and probabilistic fate decisions co-exist in a single retinal lineage

Correct nervous system development depends on the timely differentiation of progenitor cells into neurons. While the output of progenitor differentiation is well investigated at the population and clonal level, the possibilities and constraints for fate decisions of specific progenitors over development are less explored. Particularly little is known about their variability and competence plasticity. To fill this gap, we here use long-term live imaging to follow the outcome of progenitor divisions in the zebrafish retina. We find that neurogenic Atoh7 expressing progenitors produce different neuronal types over development with time-dependent probabilities. Interestingly, deterministic and probabilistic fate decisions co-exist in the same lineage. While interference with the deterministic fate affects lineage progression, interference with fate probabilities of the stochastic lineage branch results in a broader range of fate possibilities than seen in controls. When tissue development is challenged, Atoh7 expressing progenitors can produce any neuronal type, arguing against the concept of fixed competence windows. Stochastic modelling of fate probabilities in challenged conditions revealed a simple gene regulatory network able to recapitulate the observed competence changes during development. Based on our results, we postulate that fate plasticity could be involved in robust retinal development, a concept possibly applicable to other tissues.

developmental biology↗

bioRxiv2021

Bidirectional neuronal migration coordinates retinal morphogenesis bypreventing spatial competition

While the design of industrial products is often optimized for the sequential assembly of single components, organismal development is hallmarked by the concomitant occurrence of tissue growth and organization. Often this means that proliferating and differentiating cells occur at the same time in a shared tissue environment that continuously changes. How cells adapt to architectural changes in order to prevent spatial interference remains unclear. To understand how cell movements important for growth and organization are orchestrated, we here study the emergence of photoreceptor neurons that occur during the peak of retinal growth using zebrafish, human tissue and human organoids. Quantitative imaging reveals that successful retinal morphogenesis depends on active bidirectional photoreceptor translocation. This leads to a transient transfer of the entire cell population away from the apical proliferative zone. This migration pattern is driven by distinct cytoskeletal machineries, depending on direction: microtubules are required for basal translocation, while actomyosin drives apical movement. Blocking photoreceptor translocation leads to apical overcrowding that hampers progenitor movements. Thus, photoreceptor migration is crucial to prevent competition for space and thereby allows concurrent tissue growth and lamination. This shows that neuronal migration, in addition to its canonical role in cell positioning, is involved in coordinating morphogenesis.

developmental biology↗

bioRxiv2020

Asymmetric neurogenic commitment of retinal progenitors is regulated via the Notch endocytic pathway

During brain development, progenitor cells need to balance proliferation and differentiation in order to generate different neurons in the correct numbers and proportions. Currently, the patterns of multipotent progenitors’ division that lead to neurogenic entry and the factors that regulate them are not fully understood. We here use the zebrafish retina to address this gap, exploiting the possibilities for quantitative live-imaging. We show that early neurogenic progenitors arise from asymmetric divisions. Notch regulates this asymmetry as when inhibited, symmetric divisions producing two neurogenic progenitors occur. Surprisingly, Notch does not act through an apicobasal activity gradient as previously suggested but through asymmetric inheritance of Sara-positive endosomes. Further, the resulting neurogenic progenitors show cell biological features different from multipotent progenitors, raising the possibility that an intermediate state of progenitors exists in the retina. Our study thus opens new insights into the regulation of proliferative and differentiative events during central nervous system development.Competing Interest StatementThe authors have declared no competing interest.View Full Text

developmental biology↗

bioRxiv2018

De novo genesis of retinal ganglion cells by targeted expression of KLF4 in vivo

Retinal ganglion cell (RGC) degeneration is a hallmark of glaucoma, the most prevalent cause of irreversible blindness. Thus, innovative therapeutic strategies are needed to protect and replace these projection neurons. It has been shown that endogenous glial cells of the retina, Muller cells, can be directly reprogrammed into late-born retinal interneurons. However, since RGCs are the first neurons born during development, the replacement of damaged RGCs requires the reprograming to an early neurogenic state. Here, we demonstrate that the pluripotency regulator Klf4 is sufficient to reprogram the potency of lineage-restricted retinal progenitor cells (RPCs) to generate RGCs in vivo. Transcriptome analysis disclosed that the overexpression of Klf4 induces crucial regulators of RGC competence and specification, including Atoh7 and Eya2. In contrast, loss-of-function studies in mice and zebrafish demonstrated that Klf4 is not essential for generation or differentiation of RGCs during retinogenesis. Nevertheless, induced RGCs (iRGCs) generated upon Klf4 overexpression migrate to the proper layer and project axons aligned with endogenous fascicles that reach the optic nerve head. Notably, iRGCs survive for up to 30 days after in vivo reprogramming. Finally, we demonstrate that Klf4 converts Muller cells into neurons that express markers of RGCs. Altogether, we identified Klf4 as a promising tool to reprogram retinal cells and regenerate RGCs in the mature retina.\n\nSignificance StatementCell fate determination is a key process for development, regeneration and for the design of therapeutic strategies that involve cellular reprogramming. This work shows that the manipulation of a single pluripotency regulator (Klf4) is sufficient to reprogram restricted progenitor cells in vivo. These reprogrammed progenitors reacquire the potency to generate retinal ganglion cells. Ganglion cell degeneration is the leading cause of irreversible blindness; therefore, manipulation of ganglion cell competence is of relevance for human health. Our findings point to Klf4 as a promising tool to develop therapeutic strategies for the replacement of damaged ganglion cells.

developmental biology↗

bioRxiv2017

Content-Aware Image Restoration: Pushing the Limits of Fluorescence Microscopy

Fluorescence microscopy is a key driver of discoveries in the life-sciences, with observable phenomena being limited by the optics of the microscope, the chemistry of the fluorophores, and the maximum photon exposure tolerated by the sample. These limits necessitate trade-offs between imaging speed, spatial resolution, light exposure, and imaging depth. In this work we show how image restoration based on deep learning extends the range of biological phenomena observable by microscopy. On seven concrete examples we demonstrate how microscopy images can be restored even if 60-fold fewer photons are used during acquisition, how near isotropic resolution can be achieved with up to 10-fold under-sampling along the axial direction, and how tubular and granular structures smaller than the diffraction limit can be resolved at 20-times higher frame-rates compared to state-of-the-art methods. All developed image restoration methods are freely available as open source software in Python, FO_SCPLOWIJIC_SCPLOW, and KO_SCPLOWNIMEC_SCPLOW.

bioinformatics↗