Biology subjects

Gritti, N.

Publications and source records attributed to Gritti, N..

6 recordsLinked to original sources

bioRxiv2025

Axial patterning of gastruloids via diverse cell type compositions

The formation of the germ layers and antero-posterior (AP) axial patterning are interlinked milestones of embryogenesis. Gastruloids, in vitro models from aggregated embryonic stem cells (ESCs), permit the study and deconstruction of these events. Gastruloids are successfully generated from ESCs of variable pluripotent states, but it remains unknown how the initial conditions influence cell type composition and to what degree resulting variations in spatial patterning can converge onto an elongated body axis. To address this, we comparatively study aggregates from varying proportions of primed and naive ESCs. Despite differences in AP symmetry-breaking dynamics and distinct trajectories toward either anterior mesodermal or neuro-ectodermal fates, all conditions produce elongated gastruloids. Furthermore, timed modulation of Activin/Nodal signaling in aggregates equalizes AP polarization modes and mixing of initial ESC pluripotent states generates gastruloids with enhanced tissue type diversity. This work therefore uncovers a previously unappreciated developmental flexibility underlying mammalian AP axial patterning.

developmental biology↗

bioRxiv2024

Desynchronization between timers provokes transient arrest during C. elegans development

In multicellular organisms, development entails the progression of diverse cellular processes that need to be temporally coordinated. During C. elegans postembryonic development, the events of molting and cell divisions progress in parallel during four larval stages and are modulated by external cues such as temperature and nutrient availability. While seam cell divisions occur predominantly before ecdysis of the cuticle, the order of the events can change, suggesting that they are controlled by independent mechanisms. Here, we have analyzed the impact of reduced insulin signaling in molting and in stage-specific cell divisions. We find that reduced insulin signaling in the daf-2(e1370) allele delays both of these events but has a larger impact on the timing of cell divisions, thus increasing the desynchrony between the events. The relative delay in seam cell divisions leads to a delay in the initiation of the subsequent stage of the molting program, providing a mechanism for resynchronization of developmental processes to the beginning of the larval stages.

developmental biology↗

bioRxiv2023

Metabolic control of germ layer proportions through regulation of Nodal and Wnt signalling

During embryonic development, cells exit pluripotency to give rise to the three germ layers. Metabolic pathways influence cell fate decisions by modulating the epigenetic, transcriptional, and signalling states of cells. However, the interplay between metabolism and the major signalling pathways that drive the emergence of ectoderm, mesoderm, and endoderm remains poorly understood. Here, we demonstrate an instructive role of glycolytic activity in activating signalling pathways involved in mesoderm and endoderm induction. Using an in vitro model system for mouse gastrulation, we observed that inhibiting glycolysis prevents the upregulation of primitive streak markers, resulting in a significant increase in ectodermal cell fates at the expense of mesodermal and endodermal lineages. We demonstrate that this relationship is dose-dependent, enabling metabolic control of germ layer proportions through exogenous glucose levels. Mechanistically, we found that glycolysis inhibition leads to the downregulation of Wnt, Nodal, and Fgf signalling. Notably, this metabolic phenotype was rescued by Nodal or Wnt signalling agonists in the absence of glycolytic activity, suggesting that glycolytic activity acts upstream of both signalling pathways. Our work underscores the dependence of specific signalling pathways on metabolic conditions and provides mechanistic insight into the nutritional regulation of cell fate decision making.

developmental biology↗

bioRxiv2022

Image restoration of degraded time-lapse microscopy data mediated by infrared-imaging.

Time-lapse fluorescence microscopy is key to unraveling the processes underpinning biological development and function. However, living systems, by their nature, permit only a limited toolbox for interrogation. Consequently, following time-lapses, expended samples contain untapped information that is typically discarded. Herein we employ convolutional neural networks (CNNs) to augment the live imaging data using this complementary information. In particular, live, deep tissue imaging is limited by the spectral range of live-cell compatible probes/fluorescent proteins. We demonstrate that CNNs may be used to restore deep-tissue contrast in GFP-based time-lapse imaging using paired final-state datasets acquired using infrared dyes and improve information content accordingly. Surprisingly, the networks are remarkably robust over a wide range of developmental times. We employ said network to GFP time-lapse images captured during zebrafish and drosophila embryo/larval development and demonstrate live, deep tissue image contrast.

developmental biology↗

bioRxiv2021

Dynamics of anteroposterior axis establishment in a mammalian embryo-like system

1.In the mammalian embryo, specification of the anteroposterior (AP) axis demarcates one of the first steps of body plan formation. While this process requires interactions with extra-embryonic tissues in the native embryo, minimal in vitro systems from embryonic stem cells (ESCs) undergo initial AP polarization in the absence of any localized, external cues. This self-organizing potential of stem cells remains not well understood. Here, we study such an initial symmetry breaking event in gastruloids, an established in vitro model for mammalian body plan formation, using the mesodermal marker gene Brachyury or T (Bra/T) to denote the onset of AP axis specification and concomitant germ layer formation. Through aggregate fusion experiments and manipulation of initial culture conditions as well as key developmental signalling pathways, we probe the dynamics of Bra/T polarization. We further conduct single-cell (sc) RNA sequencing of gastruloids at early stages to identify incipient molecular signatures of germ layer commitment and differences between Bra/T+ and Bra/T- populations during as well as after symmetry breaking. Moreover, we transcriptionally compare early development of gastruloids to the mouse embryo and conclude that gastruloids reproducibly undergo AP axis and germ layer specification in a parallel, but distinct manner: While their primed pluripotent cell populations adopt a more mesenchymal state in lieu of an epithelial epiblast-like transcriptome, the emerging mesendodermal lineages in vitro are nevertheless similar to their in vivo equivalents. Altogether, this study provides a comprehensive analysis of self-organized body plan establishment in a minimal in vitro system of early mammalian patterning and highlights the regulative capacity of mESCs, thereby shedding light on underlying principles of axial polarity formation.

developmental biology↗

bioRxiv2019

Random birth order and bursty Notch ligand expression drive the stochastic AC/VU cell fate decision in C. elegans

Cells in developing organisms must robustly assume the correct fate in order to fulfill their specific function. At the same time, cells are strongly affected by molecular fluctuations, i.e. noise, leading to inherent variability in individual cells. During development, some cells are thought to exploit such molecular noise to drive stochastic cell fate decisions, with cells randomly picking one cell fate out of several possible ones. Yet, how molecular noise drives such decisions is an open question. We address this question using a novel quantitative approach to study one of the best-understood stochastic cell fate decisions: the AC/VU decision in C. elegans gonad development. Here, two initially equivalent cells, Z1.ppp and Z4.aaa, interact, so that one cell becomes the anchor cell (AC) and the other a ventral uterine precursor cell (VU). It is thought that the symmetry is broken when small molecular fluctuations are amplified into cell fate by positive feedback loops in the Notch signaling pathway. To identify the noise sources that drive the AC/VU decision, we used a novel time-lapse microscopy approach to follow expression dynamics in live animals and single molecule FISH to quantify gene expression with single mRNA resolution. We found not only that random Z1.ppp/Z4.aaa birth order biased the decision outcome, with the first-born cell typically assuming VU fate, but that the strength of this bias and the speed of the decision decreased as the two cells were born closer together in time. Moreover, we find that the Notch ligand lag-2/Delta exhibited strongly stochastic expression already in the two mother cells, Z1.pp/Z4.aa. Combining experiments with mathematical models, we showed that the resulting asymmetry in lag-2/Delta levels inherited by the daughter cells, Z1.ppp/Z4.aaa, stochastic symmetry breaking when both cells are born at similar times. Together, our results suggest that two independent noise sources, birth order and stochastic lag-2/Delta expression, are exploited to amplify noise into cell fate in a manner that ensures a robust decision.

developmental biology↗