Biology subjects

Charron, F.

Publications and source records attributed to Charron, F..

5 recordsLinked to original sources

bioRxiv2024

Targeting synthetic lethality between non-homologous end joining and radiation in very high-risk SHH medulloblastoma

Specific and biologically informed treatments for medulloblastoma, especially the highly lethal TP53 mutant SHH subgroup, remain elusive, where radiotherapy is the primary treatment option. Applying genome-wide CRISPR-Cas9 screening in combination with lethal doses of radiotherapy, we identified the main driver of radiation resistance in SHH medulloblastoma is loss of p53. A negative selection CRISPR-Cas9 screen across multiple models of Trp53-deficient SHH medulloblastoma revealed a strong synthetic lethal interaction between components of the non-homologous end-joining pathway and radiation, particularly DNA protein kinase (DNA-PK) and its binding partners. Both genetic and pharmacological perturbation of DNA-PK enhanced radiosensitivity in TP53-deficient SHH medulloblastoma, leading to cell death. In vivo treatment of somatic and germline TP53-mutant SHH medulloblastoma models with peposertib, a small-molecule inhibitor of DNA-PK, significantly improved survival when combined with radiotherapy, strongly supporting further clinical investigation.

cancer biology↗

bioRxiv2024

A central role for Numb/Nbl in multiple Shh-mediated axon repulsion processes

Sonic hedgehog (Shh) is an axon guidance molecule that can act as either a chemorepellent or a chemoattractant, depending on the neuron type and their developmental stage. In the developing spinal cord, Shh initially attracts commissural axons to the floor plate and later induces their repulsion after they cross the midline. In the developing visual system, Shh repels ipsilateral retinal ganglion cell (iRGC) axons at the optic chiasm. Although Shh requires the endocytic adaptor Numb for attraction of commissural neurons, the molecular mechanisms underlying Shh dual function in attraction and repulsion are still unclear. In this study, we investigate whether Numb also regulates repulsive axon guidance. We show that Numb is essential for two Shh-mediated repulsion processes: iRGC axon repulsion at the optic chiasm and antero-posterior commissural axon repulsion in the spinal cord. Therefore, Numb is required for Shh-mediated attraction and repulsion. These results position Numb as a central player in the non-canonical Shh signalling pathway mediating axon repulsion. Summary statementHere, we show that Numb is required for Shh-mediated midline repulsion of ipsilateral retinal ganglion cell axons and post-crossing commissural axons of the spinal cord.

neuroscience↗

bioRxiv2023

Eta-secretase-like processing of the amyloid precursor protein (APP) by RHBDL4

The amyloid precursor protein (APP) has been extensively studied with regards to its contribution to the pathology of Alzheimers disease. APP is an ubiquitously expressed type I transmembrane protein synthesized in the endoplasmic reticulum (ER) and translocated to the plasma membrane where it undergoes proteolytic cleavages by several identified proteases. Conversely to other known proteases, we previously elucidated human rhomboid protease RHBDL4 as a novel APP processing enzyme where several cleavages likely occur already in the ER. Interestingly, the pattern of RHBDL4-derived large APP C-terminal fragments resemble those generated by the {eta}-secretase or MT5-MMP, which was described to generate so called A{eta} fragments. The similarity in large APP C-terminal fragments between both proteases raised the question whether RHBDL4 may contribute to {eta}-secretase activity and A{eta}-like fragments. Here, we identified two cleavage sites of RHBDL4 in APP by mass spectrometry, which, intriguingly, lie in close proximity to the cleavage sites of MT5-MMP. Indeed, we observed that RHBDL4 generates A{eta}-like fragments in vitro without contributions of -, {beta}-, or {gamma}-secretases. Such A{eta}-like fragments are likely generated in the ER since RHBDL4-derived APP-C-terminal fragments do not reach the cell surface. Inherited, familial APP mutations appear to not affect this processing pathway. In RHBDL4 knockout mice, we observed increased cerebral full length APP levels in comparison to WT brains in support of RHBDL4 being a physiologically relevant protease for APP. Furthermore, we found secreted A{eta} fragments in dissociated mixed cortical cultures from wild type mice, however significantly less A{eta} fragments in cultures from RHBDL4 knockout mice. Our data underscores that RHBDL4 contributes to {eta}-secretease-like processing of APP and that RHBDL4 is a physiologically relevant protease for APP.

biochemistry↗

bioRxiv2022

A human DCC variant causing mirror movement disorder reveals an essential role for the Wave regulatory complex in Netrin/DCC signaling

The axon guidance cue, Netrin-1, signals through its receptor DCC to attract commissural axons to the midline. Pathogenic variants in DCC frequently lead to congenital mirror movements (CMM), but how these variants impact DCC function is largely unknown. Screening of DCC in individuals with CMM recently revealed a novel variant located in a conserved motif in the cytoplasmic tail of DCC that is predicted to bind to a central actin nucleation promoting factor, the WAVE regulatory complex (WRC). Here, we use biochemical and axon guidance assays to show that this CMM-associated DCC variant is pathogenic by disrupting the interaction between DCC and the WRC. This DCC-WRC interaction is evolutionarily conserved and is required for Netrin-1 mediated commissural axon outgrowth and guidance. Together, we identify the WRC as a pivotal component of Netrin-1/DCC signaling and further provide a molecular mechanism explaining how genetic variants in DCC may lead to CMM.

neuroscience↗

bioRxiv2022

Cilium proteomics reveals Numb as a positive regulator of the Hedgehog signaling pathway

The transduction of Hedgehog (Hh) signaling relies on the primary cilium, a cell surface organelle serving as a signaling hub for the cell. Using proximity labeling and quantitative proteomics, we identified Numb as a new ciliary protein that positively regulates Hh signaling. Numb localizes to the ciliary pocket and acts as an endocytic adaptor to incorporate Ptch1 into clathrin-coated vesicles, thereby promoting Ptch1 exit from the cilium, a key step in Hh signaling activation. Numb loss hampers Sonic Hedgehog (Shh)-induced Ptch1 departure from the cilium, resulting in reduced activation of Hh signaling. Numb loss in spinal neural progenitors reduces Shh-induced differentiation into Nkx2.2-positive progenitors, a process reliant on high Hh signaling activity. Genetic ablation of Numb in the developing cerebellum impaired the proliferation of granule cell precursors, a Hh-dependent process, resulting in reduced cerebellar size. This study highlights Numb as a critical regulator of Ptch1 levels in the cilium during Hh signal activation and demonstrates the key role of ciliary pocket-mediated endocytosis in modulating the transduction of cell signaling.

cell biology↗