Biology subjects

Franco, S. J.

Publications and source records attributed to Franco, S. J..

9 recordsLinked to original sources

bioRxiv2025

Optimizing in utero electroporation of the developing mouse pons to model diffuse midline glioma

The leading cause of brain cancer-related death in children is diffuse midline glioma (DMG). A particularly aggressive DMG subtype is pediatric diffuse intrinsic pontine glioma (DIPG), which is caused by the histone mutation H3.3K27M. Because of its diffuse growth and location in a critical brainstem structure, therapeutic options are limited and DIPG is considered universally fatal. Lack of appropriate animal models has hindered our understanding of the developmental origins and progression of DIPG, which in turn has limited development of effective therapeutics. To address this barrier, we optimized an in utero electroporation method to model DIPG in vivo in the developing mouse pons. In this protocol, we use in utero electroporation to express canonical DIPG mutant oncogenes in neural progenitors lining the 4th ventricle, which give rise to cells in the pons. As the embryos continue to develop in utero and then postnatally, they develop large diffuse brainstem tumors with molecular characteristics of pediatric DIPG, allowing us to model the formation and progression of this deadly pediatric brain cancer.

cancer biology↗

bioRxiv2025

Epigenetic priming of neural progenitors by Notch enhances Sonic hedgehog signaling and establishes gliogenic competence

The remarkable cell diversity of multicellular organisms relies on the ability of multipotent progenitor cells to generate distinct cell types at the right times and locations during embryogenesis. A key question is how progenitors establish competence to respond to the different environmental signals required to produce specific cell types at critical developmental timepoints. We addressed this in the mouse developing forebrain, where neural progenitor cells must switch from producing neurons to making oligodendrocytes in response to increased Sonic Hedgehog (SHH) signaling during late embryogenesis. We show that progenitor responses to SHH are regulated by Notch signaling, thus permitting proper timing of the neuron-oligodendrocyte switch. Notch activity epigenetically primes genes associated with the oligodendrocyte lineage and SHH pathway, enabling amplified transcriptional responses to endogenous SHH and robust oligodendrogenesis. These results reveal a critical role for Notch in facilitating progenitor competence states and influencing cell fate transitions at the epigenetic level.

developmental biology↗

bioRxiv2024

Meningeal-derived retinoic acid regulates neurogenesis via suppression of Notch and Sox2

The meninges act as a regulator of brain development by secreting ligands that act on neural cells to regulate neurogenesis and neuronal migration. Meningeal-derived retinoic acid (RA) promotes neocortical neural progenitor cell cycle exit; however, the underlying molecular mechanism is unknown. Here, we used spatial transcriptomics and profiling of retinoic-acid receptor- (RAR) DNA binding in Foxc1-mutant embryos that lack meninges-derived ligands to identify the neurogenic transcriptional mechanisms of RA signaling in telencephalic neural progenitors. We determined that meningeal-derived RA controls neurogenesis by suppressing progenitor self-renewal pathways Notch signaling and the transcription factor Sox2. We show that RAR binds in the Sox2ot promoter, a long non-coding RNA that regulates Sox2 expression, and RA promotes Sox2ot expression in neocortical progenitors. Our findings elucidate a previously unknown mechanism of how meningeal RA coordinates neocortical development and insight into how defects in meningeal development may cause neurodevelopmental disorders.

neuroscience↗

bioRxiv2021

Tubulinopathy mutations in TUBA1A that disrupt neuronal morphogenesis and migration override XMAP215/Stu2 regulation of microtubule dynamics

Heterozygous, missense mutations in - or {beta}-tubulin genes are associated with a wide range of human brain malformations, known as tubulinopathies. We seek to understand whether a mutations impact at the molecular and cellular levels scale with the severity of brain malformation. Here we focus on two mutations at the valine 409 residue of TUBA1A, V409I and V409A, identified in patients with pachygyria or lissencephaly, respectively. We find that ectopic expression of TUBA1A-V409I/A mutants disrupt neuronal migration in mice and promote excessive neurite branching and delayed retraction in primary neuronal cultures, accompanied by increased microtubule acetylation. To determine the molecular mechanisms, we modeled the V409I/A mutants in budding yeast and found that they promote intrinsically faster microtubule polymerization rates in cells and in reconstitution experiments with purified tubulin. In addition, V409I/A mutants decrease the recruitment of XMAP215/Stu2 to plus ends and ablate tubulin binding to TOG domains. In each assay tested, the TUBA1A-V409I mutant exhibits an intermediate phenotype between wild type and the more severe TUBA1A-V409A, reflecting the severity observed in brain malformations. Together, our data support a model in which the V409I/A mutations may limit tubulin conformational states and thereby disrupt microtubule regulation during neuronal morphogenesis and migration.

cell biology↗

bioRxiv2020

Unraveling the transcriptional networks that drive oligodendrocyte fate specification in Sonic hedgehog-responsive neocortical progenitors

In the developing nervous system, progenitors first generate neurons before making astrocytes and oligodendrocytes. We previously showed that increased Sonic hedgehog (Shh) signaling in dorsal forebrain progenitors is important for their production of oligodendrocytes as neurogenesis winds down. Here, we analyzed single-cell RNA sequencing datasets to better understand how Shh controls this neuron-to-oligodendrocyte switch in the neocortex. We first identified Shh-responding progenitors using a dataset in which Shh was overexpressed in the mouse dorsal forebrain. Pseudotime trajectory inferences revealed a subpopulation committed to the oligodendrocyte precursor cell (OPC) lineage. Genes upregulated along this lineage defined a pre-OPC state, as cells transitioned from progenitors to OPCs. Using several datasets from wild-type mouse and human embryos at different ages, we confirmed a pre-OPC state preceding OPC emergence during normal development. Finally, we show that pre-OPCs are enriched for a gene regulatory network involving the transcription factor Ascl1. Genetic lineage-tracing demonstrated Ascl1+ dorsal progenitors primarily make oligodendrocytes. We propose a model in which Shh shifts the balance between opposing transcriptional networks toward an Ascl1 lineage, thereby facilitating the switch between neurogenesis and oligodendrogenesis.

developmental biology↗

bioRxiv2020

Csmd2 interacts with Dab1 and is Required in Reelin-Mediated Neuronal Maturation

Reelin is a glycoprotein secreted by Cajal-Retzius cells to regulate development of the cerebral cortex. Reelin binding to its receptors on immature neurons initiates a signaling cascade through the downstream adaptor protein, Dab1. Defects in this signaling mechanism result in perturbed neuronal migration, reductions in dendrite complexity, and deficits in synapse development and function. How Reelin controls neuronal migration and brain lamination have been extensively investigated over the years, but the pathways that regulate dendrite and spine development downstream of Reelin and Dab1 have yet to be fully elucidated. Here, we have identified a novel interaction between Dab1 and Csmd2, a synaptic transmembrane protein required for dendrite and dendritic spine development in forebrain excitatory neurons. We demonstrate that Csmd2 contains an NPxY motif on its intracellular region, through which Dab1 interacts with Csmd2. Interestingly, we find that this NPxY consensus motif is not required for Csmd2 to localize at the postsynaptic densities of spiny neurons. Rather, the introduction of an NPxY mutant form of Csmd2 results in a significant overproduction of immature, filopodia-like dendritic spines in maturing neurons. Moreover, we show that knockdown of Csmd2 mRNA expression in immature developing neurons abolishes the ability of Reelin to promote dendrite elaboration and dendritic spine maturation. This suggests that the Csmd2-Dab1 interaction may be a requirement of Reelin/Dab1 signaling to mediate the structural maturation of neurons. Together, these results point toward a role of Csmd2 in the Reelin/Dab1 signaling axis that promotes the development of dendrites and dendritic spines in maturing neurons. Summary StatementHow Reelin controls neuronal maturation remains to be understood. We demonstrate that the synaptic protein Csmd2 interacts with the Reelin-associated adaptor protein Dab1. We also determine that Reelin requires Csmd2 to regulate structural development and maturation of forebrain neurons.

neuroscience↗

bioRxiv2019

Loss of Shh signaling in the neocortex reveals heterogeneous cell recovery responses from distinct oligodendrocyte populations

The majority of oligodendrocytes in the neocortex originate from neural progenitors that reside in the dorsal forebrain. We recently showed that Sonic Hedgehog (Shh) signaling in these dorsal progenitors is required to produce normal numbers of neocortical oligodendrocytes during embryonic development. Conditional deletion of the Shh signaling effector, Smo, in dorsal progenitors caused a dramatic reduction in oligodendrocyte numbers in the embryonic neocortex. In the current study, we show that the depleted oligodendrocyte lineage in Smo conditional mutants is able to recover to control numbers over time. This eventual recovery is achieved in part by expansion of the ventrally-derived wild-type lineage that normally makes up a minority of the total oligodendrocyte population. However, we find that the remaining dorsally-derived mutant cells also increase in numbers over time to contribute equally to the recovery of the total population. Additionally, we found that the ways in which the dorsal and ventral sources cooperate to achieve recovery is different for distinct subclasses of oligodendrocyte-lineage cells. Oligodendrocyte precursor cells (OPCs) in the neocortical white matter recover completely by expansion of the remaining dorsally-derived Smo mutant cells. On the other hand, mature oligodendrocytes in the white and gray matter recover through an equal contribution from dorsal mutant and ventral wild-type lineages. Interestingly, the only population that did not make a full recovery was OPCs in the gray matter. We find that gray matter OPCs are less proliferative in Smo cKO mutants compared to controls, which may explain their inability to fully recover. These studies shed light on the nature of competition and cooperation between dorsal and ventral sources of oligodendrocytes in the developing neocortex. Furthermore, our data point towards differential use of Shh signaling in the development of distinct subclasses of the oligodendrocyte lineage.

neuroscience↗

bioRxiv2018

Lmx1a drives Cux2 expression in the cortical hem through activation of a conserved intronic enhancer.

During neocortical development, neurons are produced by a diverse pool of neural progenitors. A subset of progenitors express the Cux2 gene and are fate-restricted to produce certain neuronal subtypes, but the upstream pathways that specify these progenitor fates remain unknown. To uncover the transcriptional networks that regulate Cux2 expression in the forebrain, we characterized a conserved Cux2 enhancer that we find recapitulates Cux2 expression specifically in the cortical hem. Using a bioinformatic approach, we found several potential transcription factor (TF) binding sites for cortical hem-patterning TFs. We found that the homeobox transcription factor, Lmx1a, can activate the Cux2 enhancer in vitro. Furthermore, we show that multiple Lmx1a binding sites required for enhancer activity in the cortical hem in vivo. Mis-expression of Lmx1a in neocortical progenitors caused an increase in Cux2+-lineage cells. Finally, we compared several conserved human enhancers with cortical hem-restricted activity and found that recurrent Lmx1a binding sites are a top shared feature. Uncovering the network of TFs involved in regulating Cux2 expression will increase our understanding of the mechanisms pivotal in establishing Cux2-lineage fates in the developing forebrain.\n\nSummary StatementAnalysis of a cortical hem-specific Cux2 enhancer reveals role for Lmx1a as a critical upstream regulator of Cux2 expression patterns in neural progenitors during early forebrain development.

neuroscience↗

bioRxiv2018

Csmd2 is a Synaptic Transmembrane Protein that Interacts with PSD-95

Mutations and copy number variants of the Cub and Sushi Multiple Domains 2 (CSMD2) gene are associated with schizophrenia and autism spectrum disorder. CSMD2 is a single-pass transmembrane protein with a large extracellular domain comprising repeats of Cub and Sushi domains. Although the biological functions of CSMD2 have not been studied, the association between CSMD2 variants and cognitive function suggest that it may have a role in brain development or function. In this study, we show that mouse Csmd2 is expressed in excitatory and inhibitory neurons in the brain. Csmd2 protein exhibits a somatodendritic localization in the neocortex and hippocampus, with smaller puncta localizing further out in the neuropil. We show that many of these Csmd2 puncta co-localize with the synaptic protein PSD-95. Using immunohistochemical and biochemical methods, we further demonstrate that Csmd2 localizes to dendritic spines and is enriched in the postsynaptic density. We also find Csmd2 at ribbon synapses of the inner plexiform layer of the retina, suggesting a broader synaptic function of Csmd2 in the central nervous system. Finally, we show that the cytoplasmic tail domain of Csmd2 interacts with synaptic scaffolding proteins of the membrane-associated guanylate kinase (MAGUK) family. The association between Csmd2 and MAGUK member PSD-95 is dependent on a PDZ-binding domain on the Csmd2 tail, which is also required for synaptic targeting of Csmd2. Together, these results point toward a function for Csmd2 in dendrites and synapses, which may account for its association with several psychiatric disorders.

cell biology↗