Biology subjects

Crist, C.

Publications and source records attributed to Crist, C..

7 recordsLinked to original sources

bioRxiv2026

Ctcf deficiency in myofibers induces pathological genome reprogramming toward the spontaneous development of myopathy

How perennial, postmitotic multinucleated tissues, such as skeletal myofibers, maintain their identity and transcriptional adaptation to homeostatic perturbations through adult life is an outstanding question. To address this issue, we investigated the consequences of loss of 3D-genome architecture in skeletal muscles by generating myofiber-specific Ctcf-deficient (CtcfmKO) mice. CtcfmKO mice did not exhibit muscular phenotype at birth but spontaneously developed a severe myopathy. Integrated analysis of snRNAseq, ATACseq and promoter-capture Hi-C revealed both common and fiber-type specific patterns of dysregulated gene expression associated with alterations in chromatin accessibility and promoter-based interactions in Ctcf-deficient myonuclei at distinct stages of myopathy development. Decreased chromatin accessibility at promoters and changes in their connectivity with distal elements were observed across all myonuclei as a direct consequence of Ctcf deficiency at early stages and associated with downregulation of genes implicated in myofiber contraction and anabolism, metabolism, adhesion and neuromuscular transmission. Conversely, at later stages, upregulation of genes leading to persistent activation of ER stress/UPR and catabolism resulted from global reconfiguration of chromatin structure and connectivity, partly as indirect consequence of Ctcf deficiency. Notably, type-IIB myonuclei exhibited specific alterations in gene expression that culminated in loss of fiber-type identity and ectopic expression of inflammatory genes. These results reveal a requirement of Ctcf for maintenance of fiber-type identity and transcriptional adaptation in vivo, through multilayered control of 3D genome integrity. They also indicate an unprecedented association between Ctcf deficiency in myofibers and susceptibility to develop myopathies, whereby Ctcf dispensability for developmental myogenesis confers vulnerability to develop myopathic syndromes.

molecular biology↗

bioRxiv2025

The Short Isoform of the Mouse Actin Adaptor Protein Synaptopodin-2 Activates Actin-Responsive Transcription Factors and Enhances Myoblast Fusion

Expression of synaptopodin-2 (SYNPO2), an actin cytoskeleton adaptor protein, is up-regulated following myoblast differentiation into myocytes and subsequently myotubes but no functional role in muscle development has been attributed to SYNPO2. We now show that of the three known mouse SYNPO2 isoforms, only the shortest isoform, SYNPO2As, is upregulated following differentiation of C2C12 myoblasts and primary mouse muscle satellite cells. Consistent with the differentiation-dependent expression pattern of these different isoforms, ectopic expression of SYNPO2As significantly increased myotube formation while the two large SYNPO2 isoforms (SYNPO2A or SYNPO2B) inhibited myocyte fusion into multinucleated myotubes. Irrespective of the fusion phenotype, all three isoforms enhanced migration of differentiating myoblasts. Knockdown studies using shRNA confirmed a pro-myogenic role for the short SYNPO2As isoform. Interestingly, SYNPO2As-transduced cells increased the transcript levels of late myogenic differentiation markers in the STARS (striated muscle activator of Rho signaling) and SRF (serum response factor) pathways that correlated with the enhanced fusion phenotype. These results identify the actin adaptor protein SYNPO2As as a new pro-myogenic factor that upregulates late myogenic differentiation markers and myotube formation.

developmental biology↗

bioRxiv2024

Modular small RNA drives the emergence of virulence traits and environmental trade-offs in Vibrio cholerae

The sole gain of laterally acquired virulence genes does not fully explain the transition of environmental strains into human pathogens. To date, the specific molecular drivers and fitness trade-offs that enable some strains within a population to undergo this process remain enigmatic. Here, we describe a small RNA (sRNA) with a unique modular structure that shapes the evolution of toxigenic Vibrio cholerae, the agent of cholera. The sRNA comprises of a highly variable 5 module located within the ompU ORF and a conserved 3 one downstream from the gene. This atypical location confers a distinct bimodular structure to the OmpU-encoded sRNA (OueS), generating allelic variants that differentially contribute to the emergence of virulence potential in some strains and associated fitness trade-offs between human infection and environmental survival. Unlike environmental counterparts, the OueS allele from toxigenic strains controls phenotypes essential during host colonization: a) stringently inhibits biofilm formation via a novel sRNA-sRNA interaction by suppressing the iron-responsive sRNA RyhB, and b) confers resistance against intestinal bacteriophages by activating the recently discovered CBASS phage defense system. Toxigenic OueS is also required for successful intestinal colonization and, as the first known example of its kind, acts as a functional surrogate of the master virulence regulator ToxR, controlling over 84% of its regulome. On the other hand, isogenic strains encoding environmental alleles of OueS exhibit higher competitive fitness than those harboring toxigenic ones during colonization of natural reservoirs such as crustaceans and phytoplankton. Our findings provide critical insights into the evolution of toxigenic V. cholerae and reveal specific molecular mechanisms and fitness costs associated with the emergence of pathogenic traits in bacteria.

microbiology↗

bioRxiv2021

Transcriptional Reprogramming of Skeletal Muscle Stem Cells by the Niche Environment

Adult stem cells are indispensable for tissue regeneration. Tissue-specific stem cells reside in a specialized location called their niche, where they are in constant cross talk with neighboring niche cells and circulatory signals from their environment. Aging has a detrimental effect on the number and the regenerative function of various stem cells. However, whether the loss of stem cell function is a cause or consequence of their aging niche is unclear. Using skeletal muscle stem cells (MuSCs) as a model, we decouple cell-intrinsic from niche-mediated extrinsic effects of aging on their transcriptome. By combining in vivo MuSC heterochronic transplantation models and computational methods, we show that on a genome-wide scale, age-related altered genes fall into two distinct categories regarding their response to the niche environment. Genes that are inelastic in their response to the niche exhibit altered chromatin accessibility and are associated with differentially methylated regions (DMRs) between young and aged cells. On the other hand, genes that are restorable by niche exposure exhibit altered transcriptome but show no change in chromatin accessibility or DMRs. Taken together, our data suggest that the niche environment plays a decisive role in controlling the transcriptional activity of MuSCs, and exposure to a young niche can reverse approximately half of all age-associated changes that are not epigenetically encoded. The muscle niche therefore serves as an important therapeutic venue to mitigate the negative consequence of aging on tissue regeneration.

genetics↗

bioRxiv2021

ERK3-MK5 signaling regulates myogenic differentiation and muscle regeneration by promoting FoxO3 degradation

The physiological functions and downstream effectors of the atypical mitogen-activated protein kinase ERK3 remain to be characterized. We recently reported that mice expressing catalytically-inactive ERK3 (Mapk6KD/KD) exhibit a reduced post-natal growth rate as compared to control mice. Here, we show that genetic inactivation of ERK3 impairs post-natal skeletal muscle growth and adult muscle regeneration after injury. Loss of MK5 phenocopies the muscle phenotypes of Mapk6KD/KD mice. At the cellular level, genetic or pharmacological inactivation of ERK3 or MK5 induces precocious differentiation of C2C12 or primary myoblasts, concomitant with MyoD activation. Reciprocally, ectopic expression of activated MK5 inhibits myogenic differentiation. Mechanistically, we show that MK5 directly phosphorylates FoxO3, promoting its degradation and reducing its association with MyoD. Depletion of FoxO3 rescues in part the premature differentiation of C2C12 myoblasts observed upon inactivation of ERK3 or MK5. Our findings reveal that ERK3 and its substrate MK5 act in a linear signaling pathway to control post-natal myogenic differentiation.

cell biology↗

bioRxiv2020

Satellite cell expansion is mediated by P-eIF2α dependent Tacc3 translation

Translational control of gene expression is an important regulator of adult stem cell quiescence, activation and self-renewal. In skeletal muscle, quiescent satellite cells maintain low levels of protein synthesis, mediated in part through the phosphorylation of eIF2 (P-eIF2). Pharmacological inhibition of the eIF2 phosphatase with the small molecule sal003 maintains P-eIF2 and permits the expansion of satellite cells ex vivo. Paradoxically, P-eIF2 also increases the translation of specific mRNAs, which is mediated by P-eIF2 dependent readthrough of inhibitory upstream open reading frames (uORFs). Here, we ask whether P-eIF2 dependent mRNA translation enables expansion of satellite cells. Using transcriptomic and proteomic analyses, we show a number of genes associated with the assembly of the spindle pole to be upregulated at the level of protein, without corresponding change in mRNA levels, in satellite cells expanded in the presence of sal003. We show that uORFs in the 5UTR of mRNA for the mitotic spindle stability gene Tacc3 direct P-eIF2 dependent translation. Satellite cells deficient for TACC3 exhibit defects in expansion, self-renewal and regeneration of skeletal muscle. SignificanceTranslational control of gene expression has emerged as an important regulator of adult stem cell populations, which maintain low levels of protein synthesis. In adult muscle stem cells, or satellite cells, a portrait of translational control has emerged whereby multiple repression mechanisms prevent the translation of specific mRNAs. It remains unclear how other mRNAs escape repression and are efficiently translated. We show that within the context of low global rates of protein synthesis, satellite cell expansion occurs through the selective translation of Tacc3 mRNA. Tacc3 deficient satellite cells expand poorly, leading to defects in skeletal muscle regeneration. Our study provides a more complete picture of translational control of gene expression in adult stem cell populations.

cell biology↗

bioRxiv2019

Ex vivo expansion of skeletal muscle stem cells with a novel small compound inhibitor of eIF2α dephosphorylation

BACKGROUNDRegeneration of adult tissues requires the activity of rare, mitotically quiescent somatic stem cells. These features are illustrated by the muscle stem cell (MuSC), also known as the satellite cell for its satellite position underneath the basal lamina of the myofiber. Isolation of MuSCs results in their rapid activation of the myogenic program and their subsequent culture ex vivo leads to loss of stem cell regenerative capacity. These shortcomings make MuSCs difficult to study, manipulate and prevent cell based therapies. We have previously shown that muscle stem cells (MuSCs) require tightly regulated protein synthesis through the phosphorylation of eIF2. Sal003, an analog of salubrinal that blocks eIF2 dephosphorylation, promotes ex vivo expansion of MuSCs retaining regenerative capacity after engraftment into the Dmdmdx mouse model of Duchenne muscular dystrophy.\n\nMETHODSSince micromolar concentrations of sal003 (10M) are required to expand MuSCs ex vivo, we undertook a structure relations study to identify novel sal003 analogs with efficacy at lower concentrations. We demonstrate ex vivo expansion of MuSCs isolated from wild-type and mdx mice using new compounds, and use CRISPR/Cas9 genome editing tools to restore dystrophin expression in cultured MuSCs.\n\nRESULTSHere, we have synthesized and screened chemical analogs of sal003 to identify a novel compound promoting the ex vivo expansion of MuSCs. The novel compound expands wild-type and mdx MuSCs more efficiently than sal003 and also prolongs culture of primary myoblasts from isolated MuSCs.\n\nCONCLUSIONSWe identify a novel sal003 analog, C10, with increased potency at lower concentrations. Culture conditions including sal003 or C10 can extend culture of primary myoblasts from isolated MuSCs, which we predict will enable their further study, genetic manipulation and cell based therapies.

cell biology↗