Imed-Eddine Gallouzi

The RNA-binding protein HuR has been shown to promote the differentiation of cultured muscle cells into muscle fibers. HuR mediates this process by differentially regulating, at different stages of this process, mRNA targets encoding pro-myogenic factors. Despite these advancements, the role of HuR, in vivo, at various stages of the myogenic process and its impact on muscle formation and function remain elusive. Towards this end, we used the Myf5-Cre loxP system to knock out HuR at a stage where muscle precursor cells (satellite cells, SCs) commit to myogenesis. Using these mice, we found that the muscle-specific depletion of HuR impairs, physiologically, its formation during embryogenesis and in response to injury. These mice exhibited smaller skeletal muscles and reduced exercise endurance. We demonstrate, using these mice, that this effect is due, in part, to the HuR-mediated regulation of mTOR (mechanistic target of rapamycin) mRNA expression. Using primary and cultured muscle cells, we show that HuR associates with this message, regulating its stability. In doing so, HuR facilitates the commitment of satellite cells toward myogenesis, thus preventing their transdifferentiation toward adipogenesis. These findings thus identify HuR as a master regulator of SCs' commitment to myogenesis and uncover a potential target for manipulating muscle myogenic capacity in both normal and pathological conditions.

Radiotherapy remains a primary treatment for brain cancers but induces neuronal damage through the generation of reactive oxygen species (ROS), particularly hydrogen peroxide (H2O2). Astrocytes, which provide essential neuroprotective support, are highly sensitive to oxidative stress. Thus, monitoring H2O2 dynamics in these cells is critical for understanding radiation-induced neurotoxicity. Conventional fluorescence-based ROS probes, such as DCFH-DA and optogenetic sensors, face limitations in selectivity, signal stability, and multiplexing. To address these challenges, we developed an electrochemical sensing platform for real-time monitoring of H2O2 in astrocyte cultures. The system employs laser-scribed graphene (LSG) interdigitated electrodes with platinum (Pt) and Prussian blue (PB) functioning as generator and collector electrodes, respectively. PB’s molecular selectivity toward H2O2 enables high specificity and mitigates interference from electroactive species in complex media. The sensor exhibited a limit of detection of 4.58 µM and a sensitivity of 579 mA•M⁻¹•cm⁻² in cell culture medium. Its capability was validated by detecting extracellular H2O2 released from X-ray-irradiated astrocytes across varying doses and time points. Overall, this graphene-based electrochemical system provides a robust and scalable approach for real-time analysis of radiation-induced oxidative stress, offering a valuable tool for studying neuroprotection, radiotherapy response, and brain-on-chip disease modeling.

Low-temperature PECVD growth of TiO₂@C core–shell nanowires with enhanced electrochemical performance, stability, and biocompatibility for cellular interface applications.

The third Sustainable Development Goal (SDG 3—Good Health and Wellbeing) is one of the goals the United Nations created to make sure people live healthy and happy lives. Biology, the study of living organisms, can help us achieve SDG 3. One important example of how studying biology can keep people healthy is how science rapidly developed the vaccines that put an end to the coronavirus pandemic, a global health crisis. This was made possible by the efforts of scientists who studied how proteins are made by cells. We, along with other scientists, are studying the steps involved in making proteins to find ways to prevent and treat muscle diseases. This article highlights why health is important, some of the things everyone can do to stay healthy, and how studying diseases—and solving them by understanding biology—can help many people around the world live healthier, longer lives.

The inherently low thermal stability of RNA poses operational, accessibility, and financial challenges for RNA research and RNA-based technologies. Natural deep eutectic solvents (NADES) have been shown to enhance the stability of various macromolecules, including DNA and protein. This study explores NADES as alternative storage media for RNA preservation and evaluates their biocompatibility in human cells. In vitro -transcribed mRNA was stored in various NADES at temperatures ranging from 4−50 °C, and the integrity was assessed by quantifying its translatability, represented by protein-expressing cells. NADES effectively preserved RNA integrity, retaining over 50% of its translatability for at least four months at room temperature (21 °C) and 48 hours at 50 °C, in comparison to −80 °C storage. Notably, the preservation efficacy remained unaffected by temperature fluctuations. Furthermore, the selected NADES concentrations maintained ∼ 99% cell viability, demonstrating their biocompatibility. These findings establish NADES as efficient, biocompatible RNA storage media, enabling stable storage and transport at ambient and extreme temperatures while withstanding sudden fluctuations. This enhanced stability simplifies and expands the accessibility of RNA-based applications. Additionally, the biocompatibility of NADES supports their potential use in RNA-based biomedical applications.

The RNA-binding protein HuR regulates various cellular processes, such as proliferation, differentiation, and cell fate. Moreover, recent studies have shown that HuR modulates the expression of factors important for tumor growth and progression. Despite its prominent role in tumorigenesis, until recently, there have been no reported mutations in HuR that have been associated to cancer. Here, we show that a HuR mutation, HuR-V225I, previously identified in a patient with Adult T-cell Leukemia/Lymphoma, interferes with the pro-apoptotic function of HuR. In response to apoptosis, HuR translocates to the cytoplasm and is cleaved in a caspase-dependent manner. In cervical cancer cells, neuroblastoma cells, and T-lymphocytes, we observed a decrease in cleavage of the HuR-V225I mutant under apoptotic conditions. This effect was shown to be mediated by the nuclear retention of HuR-V225I. Finally, expression of the HuR-V225I mutant decreases the cell’s response to apoptotic stimuli through the increased expression of mRNAs encoding anti-apoptotic factors, such as XIAP and BCL-2. Therefore, our data establishes that the absence of HuR cytoplasmic translocation and cleavage promotes cell viability, and that acquiring this mutation during tumorigenesis may thus reduce the efficacy of cancer therapy.

Translation is one of the main gene expression steps targeted by cellular stress, commonly referred to as translational stress, which includes treatment with anticancer drugs. While translational stress blocks the translation initiation of bulk mRNAs, it nonetheless activates the translation of specific mRNAs known as short upstream open reading frames (uORFs)-mRNAs. Among these, the ATF4 mRNA encodes a transcription factor that reprograms gene expression in cells responding to various stresses. Although the stress-induced translation of the ATF4 mRNA relies on the presence of uORFs (upstream to the main ATF4 ORF), the mechanisms mediating this effect, particularly during chemoresistance, remain elusive. Here, we report that ALKBH5 (AlkB Homolog 5) and FTO (FTO: Fat mass and obesity-associated protein), the two RNA demethylating enzymes, promote the translation of ATF4 mRNA in a transformed liver cell line (Hep3B) treated with the chemotherapeutic drug sorafenib. Using the in vitro luciferase reporter translational assay, we found that depletion of both enzymes reduced the translation of the reporter ATF4 mRNA upon drug treatment. Consistently, depletion of either protein abrogates the loading of the ATF3 mRNA into translating ribosomes as assessed by polyribosome assays coupled to RT-qPCR. Collectively, these results indicate that the ALKBH5 and FTO-mediated translation of the ATF4 mRNA is regulated at its initiation step. Using in vitro methylation assays, we found that ALKBH5 is required for the inhibition of the methylation of a reporter ATF4 mRNA at a conserved adenosine (A235) site located at its uORF2, suggesting that ALKBH5-mediated translation of ATF4 mRNA involves demethylation of its A235. Preventing methylation of A235 by introducing an A/G mutation into an ATF4 mRNA reporter renders its translation insensitive to ALKBH5 depletion, supporting the role of ALKBH5 demethylation activity in translation. Finally, targeting either ALKBH5 or FTO sensitizes Hep3B to sorafenib-induced cell death, contributing to their resistance. In summary, our data show that ALKBH5 and FTO are novel factors that promote resistance to sorafenib treatment, in part by mediating the translation of ATF4 mRNA.

HuR (ElavL1) is one of the main post-transcriptional regulators that determines cell fate. Although the role of HuR in apoptosis is well established, the post-translational modifications that govern this function remain elusive. In this study, we show that PARP1/2-mediated poly(ADP)-ribosylation (PARylation) is instrumental in the pro-apoptotic function of HuR. During apoptosis, a substantial reduction in HuR PARylation is observed. This results in the cytoplasmic accumulation and the cleavage of HuR, both of which are essential events for apoptosis. These effects are mediated by a pADP-ribose–binding motif within the HuR-HNS region (HuR PAR-binding site). Under normal conditions, the association of the HuR PAR-binding site with pADP-ribose is responsible for the nuclear retention of HuR. Mutations within this motif prevent the binding of HuR to its import factor TRN2, leading to its cytoplasmic accumulation and cleavage. Collectively, our findings underscore the role of PARylation in controlling the pro-apoptotic function of HuR, offering insight into the mechanism by which PARP1/2 enzymes regulate cell fate and adaptation to various assaults.

Poly(ADP-ribosylation) (PARylation) is a post-translational modification mediated by a subset of ADP-ribosyl transferases (ARTs). Although PARylation-inhibition based therapies are considered as an avenue to combat debilitating diseases such as cancer and myopathies, the role of this modification in physiological processes such as cell differentiation remains unclear. Here, we show that Tankyrase1 (TNKS1), a PARylating ART, plays a major role in myogenesis, a vital process known to drive muscle fiber formation and regeneration. Although all bona fide PARPs are expressed in muscle cells, experiments using siRNA-mediated knockdown or pharmacological inhibition show that TNKS1 is the enzyme responsible of catalyzing PARylation during myogenesis. Via this activity, TNKS1 controls the turnover of mRNAs encoding myogenic regulatory factors such as nucleophosmin (NPM) and myogenin. TNKS1 mediates these effects by targeting RNA-binding proteins such as Human Antigen R (HuR). HuR harbors a conserved TNKS-binding motif (TBM), the mutation of which not only prevents the association of HuR with TNKS1 and its PARylation, but also precludes HuR from regulating the turnover of NPM and myogenin mRNAs as well as from promoting myogenesis. Therefore, our data uncover a new role for TNKS1 as a key modulator of RBP-mediated post-transcriptional events required for vital processes such as myogenesis.