Biology subjects

Gaigneaux, A.

Publications and source records attributed to Gaigneaux, A..

6 recordsLinked to original sources

bioRxiv2026

K-Ras controls asymmetric cell divisions from the primary cilium

The Ras-MAPK pathway drives central cellular processes, including cell proliferation and differentiation. How exactly Ras controls differentiation is however not understood. Supported by mathematical modelling and single-cell RNA sequencing we show that K-Ras4B sustains ciliation during differentiation thus restricting commitment of skeletal muscle stem and progenitor cells during asymmetric cell divisions. Modulation of K-Ras4B abundance or expression of oncogenic K-Ras4B-G12C perturb normal differentiation. K-Ras4B, but not N-Ras and H-Ras, localizes to the primary cilium and its abundance there depends on the ciliary trafficking chaperone PDE6D. The presence of B-Raf and active MEK at the base of and active ERK inside the cilium suggests that K-Ras4B is active there. Conditions that localize a K-Ras4B mutant only to the cilium are sufficient to sustain ciliation and normal differentiation. Finally, in vivo modulation of K-Ras4B activity during zebrafish embryogenesis perturbs ciliation-dependent heart-looping. Our results thus imply a novel fundamental role of K-Ras4B in controlling ciliation and differentiation and suggest an explanation for the phenotypic similarities between RASopathies and ciliopathies.

cell biology↗

bioRxiv2025

ZFHX4 is necessary for dopaminergic neuron differentiation and controls cell cycle by regulating LIN28A

The selective degeneration of midbrain dopaminergic neurons (mDANs) is the main pathological hallmark of Parkinsons disease (PD). Although many transcription factors (TFs) guiding mDAN development have been identified, the details of the underlying regulatory networks remain elusive. We have previously generated time-series transcriptomic and epigenomic profiles of human induced pluripotent stem cell (hiPSC)-derived mDANs. Integrative analysis of the data identified ZFHX4 as a prominent super-enhancer-controlled TF induced in mDAN differentiation. ZFHX4 has been associated with neurodevelopmental processes in several species and shows reduced expression in midbrain of PD patients. Using in vitro knockdown (KD) and overexpression experiments, we show that ZFHX4 is necessary but not sufficient for mDAN differentiation. ZFHX4 binds preferentially at active promoter regions and transcriptomic analysis upon ZFHX4 depletion during mDAN differentiation revealed putative primary target genes to be enriched for targets of cell-cycle-related TFs and pathways. Consistently, ZFHX4-depleted cells accumulated in G2-phase of the cell cycle, preventing normal cell cycle progression and exit. The RNA-binding protein LIN28A, involved in stem-cell maintenance and microRNA (miRNA) maturation, emerged as one of the most upregulated genes upon ZFHX4-KD, in parallel with downregulation of neurogenic miRNA miR-9. Moreover, the LIN28A locus was enriched for ZFHX4 binding in CUT&Tag analysis. Taken together, our analysis indicates a pivotal role for ZFHX4 in regulating the cell cycle, specifically in silencing multipotency and proliferative programs, while maintaining mDANs in a post-mitotic state by controlling LIN28A-miR-9 axis.

molecular biology↗

bioRxiv2023

The response to ketogenic diet in colorectal cancer is mediated by lasting functional alterations of the gut microbiome

Colorectal cancer (CRC) patients have been shown to possess an altered gut microbiome. Diet is a well-established modulator of the microbiome, and thus, dietary interventions might have a beneficial effect on CRC. An attenuating effect of the ketogenic diet (KD) on CRC cell growth has been previously observed, however the role of the gut microbiome in driving this effect remains unknown. Here, we describe a reduced colonic tumor burden upon KD consumption in a CRC mouse model with a humanized microbiome. Importantly, we demonstrate a causal relationship through microbiome transplantation into germ-free mice, whereby alterations in the gut microbiota were maintained in the absence of continued selective pressure from the KD. Specifically, we identify a shift toward bacterial species that produce stearic acid in ketogenic conditions, whereas consumers were depleted, resulting in elevated levels of free stearate in the gut lumen. This microbial product demonstrated tumor-suppressing properties by inducing apoptosis in cancer cells and decreasing colonic Th17 immune cell populations. Taken together, the beneficial effects of the KD are mediated through alterations in the gut microbiome, including, among others, increased stearic acid production, which in turn significantly reduces intestinal tumor growth.

cancer biology↗

bioRxiv2023

Identification of an H-Ras nanocluster disrupting peptide

The Ras-MAPK pathway is critical to regulate cell proliferation and differentiation. Its dysregulation is implicated in the onset and progression of numerous types of cancers. To be active, Ras proteins are membrane anchored and organized into nanoclusters, which realize high-fidelity signal transmission across the plasma membrane. Nanoclusters therefore represent potential drug targets. However, targetable protein components of signalling nanoclusters are poorly established. We previously proposed that the nanocluster scaffold galectin-1 (Gal1) enhances H-Ras nanoclustering by stabilizing stacked dimers of H-Ras and Raf via a direct interaction of dimeric Gal1 with the Ras binding domain (RBD) in particular of B-Raf. Here, we provide further supportive evidence for this model. We establish that the B-Raf preference emerges from divergent regions of the Raf RBDs that were proposed to interact with Gal1. We then identify the L5UR peptide, which disrupts this interaction by binding with low micromolar affinity to the B-Raf-RBD. Its 23-mer core fragment is thus sufficient to interfere with Gal1-enhanced H-Ras nanocluster, reduce MAPK-output and cell viability in HRAS-mutant cancer cell lines. Our data therefore suggest that the interface between Gal1 and the RBD of B-Raf can be targeted to disrupt Gal1-enhanced H-Ras nanoclustering. Collectively, our results support that Raf-proteins are integral components of active Ras nanoclusters.

biochemistry↗

bioRxiv2023

An improved PDE6D inhibitor combines with Sildenafil to synergistically inhibit KRAS mutant cancer cell growth

The trafficking chaperone PDE6D (or PDE) was proposed as a surrogate target for K-Ras, leading to the development of a series of inhibitors that block its prenyl-binding pocket. These inhibitors suffered from low solubility and intracellular potency, preventing their clinical development. Here we developed a highly soluble PDE6D inhibitor (PDE6Di), Deltaflexin3, which has the currently lowest off-target activity, as we demonstrate in dedicated assays. We further increased the K-Ras focus, by exploiting that PKG2-mediated phosphorylation of Ser181 lowers K-Ras binding to PDE6D. Thus, the combination of Deltaflexin3 with the approved PKG2-activator Sildenafil synergistically inhibits cell- and microtumor growth. However, the overall cancer survival of the high PDE6D/ low PKG2 target population is higher than of the group with the opposite signature. Our results therefore suggest re-examining the interplay between PDE6D and K-Ras in cancer, while recommending the development of PDE6Di that plug, rather than stuff the hydrophobic pocket of PDE6D. SignificanceCombinations of a novel PDE6D inhibitor with Sildenafil synergistically focus the inhibition on K-Ras, however, survival data of the target population suggest an interplay of K-Ras and PDE6D that needs further exploration.

biochemistry↗

bioRxiv2023

A multi-omics integrative approach unravels novel genes and pathways associated with senescence escape after targeted therapy in NRAS mutant melanoma

Therapy Induced Senescence (TIS) leads to sustained growth arrest of cancer cells. The associated cytostasis has been shown to be reversible and cells escaping senescence further enhance the aggressiveness of cancers. Together with targeted therapeutics, senolytics, specifically targeting senescent cancer cells, constitute a promising avenue for improved cancer treatments. Understanding how cancer cells evade senescence is needed to optimise the clinical benefits of this therapeutic approach. Here we characterised the response of three different NRAS mutant melanoma cell lines to a combination of CDK4/6 and MEK inhibitors over 33 days. Transcriptomic data show that all cell lines trigger a senescence programme coupled with strong induction of interferons. Kinome profiling revealed the activation of Receptor Tyrosine Kinases (RTKs) and enriched downstream signaling of neurotrophin, ErbB and insulin pathways. Characterisation of the miRNA interactome associates miR-211-5p with resistant phenotypes. Finally, iCELL-based integration of bulk and single-cell RNA-seq data identified biological processes perturbed during senescence, and predicts new genes involved in its escape. Overall, our data associate insulin signaling with persistence of a senescent phenotype and suggest a new role for interferon gamma in senescence escape through the induction of EMT and the activation of ERK5 signaling.

cancer biology↗