Biology subjects

Cutillas, P.

Publications and source records attributed to Cutillas, P..

14 recordsLinked to original sources

bioRxiv2026

β3-Integrin controls pericyte metabolic states and shapes tumour-stromal metabolic crosstalk in breast cancer

Pericytes are emerging as dynamic regulators of the tumour microenvironment. Yet, their role in tumour metabolism remains elusive. Here, we investigate whether {beta}3-integrin regulates pericyte metabolic state and shapes stromal-tumour metabolic interactions in breast cancer. By integrating spatial and single-cell transcriptomics from human breast tumours with multi-omics profiling of tumour-derived pericytes in vitro, we identify two {beta}3-integrin-dependent metabolic states. {beta}3-integrin-high pericytes display a metabolically active phenotype characterised by increased glycolysis and enhanced de novo serine/glycine synthesis, supporting collagen production. In contrast, {beta}3-integrin loss induces a lipid-associated state, marked by neutral lipid accumulation and lipid droplets. Mechanistically, {beta}3-integrin regulates this metabolic switch via mTOR signalling. Importantly, these states extend beyond pericytes, with adjacent cancer cells shifting towards fatty acid oxidation and lipid use near {beta}3-integrin-low pericytes. Together, our findings establish {beta}3-integrin as a key metabolic switch in pericytes and highlight their role in driving tumour metabolic plasticity.

cancer biology↗

bioRxiv2026

DUSP6 is transcriptionally upregulated by activated ALK and cooperates with ALK signaling to reduce lorlatinib sensitivity in neuroblastoma cells

Neuroblastoma is a pediatric, sympathoadrenal tumour accounting for 7-10% of childhood malignancies. Some neuroblastomas are driven by activating mutations in ALK kinase and inhibitors show promise in clinical trials. Nevertheless, with resistance an ever-present concern, it remains important to understand better the effectors and modulators of ALK signaling. Wild type ALK promotes ERK activation, raising expression of negative regulators such as the dual-specificity phosphatase (DUSP) DUSP6. DUSP6 though can be pro- or anti-oncogenic in different cancers and its role in neuroblastoma cells remains unclear. We sought to understand its role in cells with either wild type or mutated ALK. Mutated ALK strongly promotes DUSP6 transcription, but apparently not through the ERK pathway. DUSP6 also appears to promote neuroblastoma cell proliferation without affecting ERK. Additionally, when DUSP6 is lost, the cells become more sensitive to ALK inhibitors lorlatinib and crizotinib. Phosphoproteomic analysis of such cells demonstrates that mutated ALK cooperates with DUSP6 to maximise signaling through several potential pathways, but again not through ERK or AKT. Their cooperation may also maintain optimal levels of N-Myc in MYCN-amplified neuroblastoma cells. While key substrates of DUSP6 remain to be determined in neuroblastoma cells, our study defines a novel role for this phosphatase in supporting the action of oncogenic ALK.

Cancer Biology↗

bioRxiv2025

An AGO2 adaptor expands the functional and evolutionary reach of microRNA targeting

Current models of microRNA (miRNA) silencing posit that RNA-sequence rules are sufficient for canonical targeting of mRNAs by Argonaute 2 (AGO2), the central protein of the miRNA-induced silencing complex (miRISC). Using chimeric eCLIP in CRISPR-edited LIMD1+/+, LIMD1+/-, and LIMD1-/- human small airway epithelial cells (hSAECs), we reveal a transcriptome-wide dependency on LIMD1, an AGO2 adaptor, for effective miRNA targeting and repression. In LIMD1-deficient cells, miRNA loading is uncoupled from productive targeting: despite increased AGO2-miRNA interactions, complexes engage fewer transcripts and sites, reducing occupancy and more than halving both the breadth and depth of targeting. We also observe altered AGO2 positional footprints across targets in LIMD1-deficient cells. LIMD1 dependence is most pronounced at defined RNA contexts: weak (GC-poor) seed pairings, interactions involving evolutionarily young miRNAs or sites that nonetheless form thermodynamically stable duplexes, with these losses particularly enriched in coding sequences of rapidly evolving C2H2-zinc-finger genes. Even within canonical seed repertoires of individual AGO2-miRNAs, LIMD1 is most critical at poorly conserved sites, indicating that LIMD1 broadens miRNA regulation beyond ancient, deeply conserved targets. In culture, LIMD1 deficiency de-represses oncogenic proteins that, in vivo, inversely correlate with LIMD1 levels in normal lung and adenocarcinoma, where LIMD1 is characteristically reduced, and whose dysregulation predicts poor survival. Thus, LIMD1 emerges as a key determinant of miRISC architecture, targeting, and potency, challenging RNA-centric models of miRNA function and exemplifying how adaptor proteins diversify post-transcriptional regulation. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=180 HEIGHT=200 SRC="FIGDIR/small/680889v2_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@e0f4f5org.highwire.dtl.DTLVardef@f23fd9org.highwire.dtl.DTLVardef@3de3f2org.highwire.dtl.DTLVardef@1404af2_HPS_FORMAT_FIGEXP M_FIG LIMD1 defines the scope of miRNA-mediated targeting and repression O_LIAGO2-chimeric eCLIP in CRISPR-edited human small airway epithelial cells (hSAECs) shows LIMD1 is required for productive AGO2-miRNA engagement transcriptome-wide. C_LIO_LILIMD1 deficiency reduces the AGO2-miRNA:targetome. Each AGO2-miRNA binds fewer targets, with lower occupancy per site and per transcript and fewer global silencing events. C_LIO_LILIMD1 dependence is strongest for GC-poor seed-sites, less conserved miRNAs and sites, and thermodynamically stronger duplexes. C_LIO_LIDosage-dependent effects of LIMD1 deficiency are broadly observed. C_LIO_LITarget-mRNA decay and translational repression are reduced in LIMD1-deficient hSAECs, increasing protein output. C_LIO_LIIn vivo, LIMD1 LOH-associated deficiency is prevalent and typically clonal in NSCLC. LIMD1 expression inversely correlates with oncogene levels in normal lung and adenocarcinoma, and target dysregulation predicts poor survival. C_LI C_FIG

molecular biology↗

bioRxiv2025

The acidic N-terminus of HHARI and neddylation are essential for the activation and maintenance of RIG-I-mediated type I interferon response

Human homolog of Ariadne (HHARI) is a RING-between-RING ubiquitin E3 ligase which interacts with cullin-RING E3 ligase (CRL) complexes. HHARI has been implicated in the type-I interferon anti-viral response. However, how HHARI drives interferon signalling is not fully understood and the function of the unique, highly conserved acidic N-terminal domain of the protein is unknown. Here, we show that HHARI stimulates interferon-{beta} secretion and autocrine type-I interferon signalling by directly targeting the viral RNA sensor RIG-I (Retinoic Acid-Inducible Gene I) in a neddylation-dependent manner. This suggests that neddylation inhibition could be used to treat interferonopathies and related diseases. Truncated HHARI containing only the N-terminal acidic/UBA-like domains retained the ability to induce interferon signalling in a neddylation-dependent mechanism. HHARI-mediated interferon-{beta} secretion was enhanced by overexpression of cullins 1-5. The N-terminal acidic/UBA-like domain of HHARI is critical for RIG-I activation and interferon signalling, as removal of these domains inactivated the pro-interferon phenotype. We propose a mechanism by which the N-terminus of HHARI interacts with all neddylated cullins leading to endogenous HHARI activation. This suggests a model in which the N-terminus of HHARI unlocks and activates neddylated cullins, which in turn are required for activation of HHARI itself. As cullins typically form modular cullin-RING ligase super-assemblies our findings imply that the HHARI N-terminus domain is a critical regulator of the versatile CRL system, which, through widespread protein ubiquitylation, controls many eukaryotic cell functions.

immunology↗

bioRxiv2025

RAS-PI3K Pathway in CAFs Shapes Physicochemical Properties of Tumor ECM and restrains Tumor progression

Cancer-associated fibroblasts display extensive functional plasticity, yet the signaling mechanisms that stabilize distinct CAF states remain poorly understood. Here, we identify stromal RAS-PI3K signaling as a key regulator of myofibroblastic CAF identity and matrix competence in lung cancer. Genetic disruption of the RAS-PI3K interaction in fibroblasts preserved upstream responsiveness to activating cues but impaired cytoskeletal remodeling, contractile execution and acquisition of a matrix-remodeling CAF program. As a result, RAS-PI3K-deficient CAFs generated extracellular matrices with defective collagen and fibronectin organization, reduced stiffness, altered microstructural properties and broad compositional remodeling of the matrisome, including changes in ECM glycoproteins and glycosylation-associated enzymes. These altered matrices failed to efficiently provide directional, adhesive and proliferative cues to tumor cells, attenuated EMT-associated programs and increased sensitivity to cisplatin plus pemetrexed. In vivo, fibroblast-specific RAS-PI3K disruption reduced tumor burden, decreased tumor cell proliferation, prolonged survival and improved chemotherapy response. Loss of RAS-PI3K signaling did not simply suppress CAF activation, but redirected CAFs toward a STAT3/NF-{kappa}B-linked immunomodulatory state associated with IL6, CXCL12 and PDGFR induction. Functionally, this altered stromal state promoted iNOS-associated macrophage features and was accompanied in vivo by reduced macrophage accumulation and increased CD8 T cell infiltration. These findings establish RAS-PI3K signaling as a stromal state-control mechanism that couples myofibroblastic matrix competence to tumor-supportive microenvironmental function while restraining alternative immunomodulatory CAF programs.

cancer biology↗

bioRxiv2024

The INO80 chromatin remodeling complex regulates histone H2A.Z mobility and the G1-S transition in oligodendrocyte precursors

Chromatin remodelling complexes (CRCs) participate in oligodendrocyte (OL) differentiation, survival and maintenance. We asked whether CRCs also control proliferation of OL precursors (OPs) - focusing on the INO80 complex, which is known to regulate proliferation of a variety of other cell types during development and disease. CRISPR/Cas9-mediated inactivation of Ino80 in vitro, or Cre-mediated deletion in vivo, slowed the OP cell cycle substantially by prolonging G1, without inducing OL differentiation. RNAseq analysis revealed that E2F target genes were dysregulated in OPs from INO80-deficient mice, but correlated RNAseq and ATAC-seq uncovered no general correlation beween gene expression and altered nucleosome positioning at transcription start sites. Fluorescence photobleaching experiments in cultured OPs demonstrated that histone H2A.Z mobility increased following loss of INO80, suggesting that INO80 regulates the cell cycle machinery in OPs through H2A.Z/ H2A exchange. We also present evidence that INO80 associates with OLIG2, a master regulator of OL development.

neuroscience↗

bioRxiv2023

SCD inhibition eradicates AML displaying high de novo fatty acid desaturation and synergizes with chemotherapy

Identification of specific and therapeutically actionable vulnerabilities in acute myeloid leukaemia (AML) is needed to improve patients outcome. These features should be ideally present in many patients independently of mutational background. Here we identify de novo fatty acid (FA) desaturation, specifically stearoyl-CoA desaturase (SCD) inhibition, as a therapeutic vulnerability across multiple AML models in vitro and in vivo. We use the novel clinical grade SCD inhibitor SSI-4 to show that SCD inhibition induces AML cell death via pleiotropic effects, and sensitivity is based on their dependency on FA desaturation regardless of mutational profile. SSI-4 efficacy is enhanced by driving FA biosynthesis in vitro while stroma confers protective effects that extend to in vivo models. SCD inhibition increases DNA damage and its combination with standard DNA damage-inducing chemotherapy prolongs survival in aggressive murine AML models. Our work supports developing FA desaturase inhibitors in AML while stressing the importance of identifying predictive biomarkers of response and biologically validated combination therapies to realize their therapeutic potential. One Sentence SummarySCD inhibition is toxic to AML cells with high rates of fatty acid desaturation and in combination with chemotherapy prolongs survival in murine AML models.

cancer biology↗

bioRxiv2023

Microtubule-associated proteins MAP7 and MAP7D1 promote DNA double strand break repair in the G1 cell cycle phase.

The DNA-damage response is a complex signalling network that guards genomic integrity. The microtubule cytoskeleton is involved in the repair of DNA double-strand breaks; however, little is known about which cytoskeleton-related proteins are involved in DNA repair and how. Using quantitative proteomics, we discovered that microtubule associated proteins MAP7 and MAP7D1 interact with several DNA repair proteins including DNA double-strand break repair proteins RAD50, BRCA1 and 53BP1. We observed that downregulation of MAP7 and MAP7D1 leads to increased phosphorylation of p53 after {gamma}- irradiation. Moreover, we determined that the downregulation of MAP7D1 leads to a strong G1 arrest and that the downregulation of MAP7 and MAP7D1 in cells arrested in G1 negatively affects DNA repair, recruitment of RAD50 to chromatin and localisation of 53BP1 to the sites of damage. These findings describe for the first time a novel function of MAP7 and MAP7D1 in cell cycle regulation and cellular response to DNA double-strand breaks.

cell biology↗

bioRxiv2022

Extracellular matrix educates a tumor macrophage phenotype found in ovarian cancer metastasis

Recent studies have shown the tumor extracellular matrix (ECM) associates with immunosuppression, and that targeting the ECM can improve immune infiltration and immunotherapy response. A question that remains is whether the ECM is directly educating the immune phenotypes seen in cancer. We identified a tumor-associated macrophage (TAM) population correlated with poor prognosis, interruption of the cancer immunity cycle, and tumor ECM composition. To investigate whether ECM was capable of generating the TAM phenotype seen, we developed a decellularized tissue model that retains the native ECM architecture and composition. Macrophages cultured on decellularized ovarian metastasis shared transcriptional profiles with the TAMs found in human tissues. ECM educated macrophages have a tissue remodeling and immunoregulatory phenotype, inducing altered T cell function. We conclude that the tumor ECM is directly educating this macrophage population found in cancer tissues. Therefore, current and emerging cancer therapies that target the tumor ECM may be tailored to improve macrophage phenotype and their downstream regulation of immunity.

cancer biology↗

bioRxiv2021

Proximity interactome of LC3B in normal growth conditions

LC3 (Light Chain 3) is a key player of autophagy, a major stress-responsive proteolysis pathway promoting cellular homeostasis. It coordinates the formation and maturation of autophagosomes and recruits cargo to be further degraded upon autophagosome-lysosome fusion. To orchestrate its functions, LC3 binds to multiple proteins from the autophagosomes inner and outer membranes, but the full extent of these interactions is not known. Moreover, LC3 has been increasingly reported in other cellular locations than the autophagosome, with cellular outcome not fully understood and not all related to autophagy. Furthermore, novel functions of LC3 as well as autophagy can occur in cells growing in a normal medium thus in non-stressed conditions. A better knowledge of the molecule in proximity to LC3 in normal growth conditions will improve the understanding of LC3 function in autophagy and in other cell biology function. Using an APEX2 based proteomic approach, we have detected 407 proteins in proximity to the well-characterised LC3B isoform in non-stress conditions. These include known and novel LC3B proximity proteins, associated with various cell localisation and biological functions. Sixty-nine of these proteins contain a putative LIR (LC3 Interacting Region) including 41 not reported associated to autophagy. Several APEX2 hits were validated by co-immunoprecipitation and co-immunofluorescence. This study uncovers the LC3B global interactome and reveals novel LC3B interactors, irrespective of LC3B localisation and function. This knowledge could be exploited to better understand the role of LC3B in autophagy and non-autophagy cellular processes.

cell biology↗

bioRxiv2020

CKS1-dependent proteostatic regulation has dual roles combating acute myeloid leukemia whilst protecting normal hematopoiesis

Acute myeloid leukemia (AML) is an aggressive hematological disorder comprising a hierarchy of quiescent leukemic stem cells (LSCs) and proliferating blasts with limited self-renewal ability. AML has a dismal prognosis, with extremely low two-year survival rates in the poorest cytogenetic risk patients, primarily due to the failure of intensive chemotherapy protocols unable to deplete LSCs, which reconstitute the disease in vivo, and the significant toxicity towards healthy hematopoietic cells. Whilst much work has been done to identify genetic and epigenetic vulnerabilities in AML LSCs, little is known about protein dynamics and the role of protein degradation in drug resistance and relapse. Here, using a highly specific inhibitor of the SCFSKP2-CKS1 complex, we report a dual role for CKS1-dependent protein degradation in reducing AML blasts in vivo, and importantly depleting LSCs. Whilst many AML LSC targeted therapies show significant toxicity to healthy hematopoiesis, inhibition of CKS1-dependent protein degradation has the opposite effect, protecting normal hematopoietic cells from chemotherapeutic toxicity. Together these findings demonstrate CKS1-dependent proteostasis is key for normal and malignant hematopoiesis. SignificanceCKS1-dependent protein degradation is a specific vulnerability in AML LSCs. Specific inhibition of SCFSKP2-CKS1 is lethal to CKS1Bhigh AML blasts and all AML LSCs. Normal hematopoiesis is protected from chemotherapeutic toxicity by inhibition of CKS1-dependent protein degradation, substantiating a dual role for CKS1-dependent protein degradation in clinical treatment of AML.

cancer biology↗

bioRxiv2019

The breast cancer oncogene IKKε coordinates mitochondrial function and serine metabolism

The I{kappa}B kinase {varepsilon} (IKK{varepsilon}) is a key molecule at the crossroads of inflammation and cancer. Known for its role as an activator of NF{kappa}B and IRF3 signalling leading to cytokine secretion, the kinase is also a breast cancer oncogene, overexpressed in a variety of tumours. However, to what extent IKK{varepsilon} remodels cellular metabolism is currently unknown. Here we used a combination of metabolomics and phosphoproteomics to show that IKK{varepsilon} orchestrates a complex metabolic reprogramming that affects mitochondrial metabolism and serine biosynthesis. Acting independently of its canonical signalling role, IKK{varepsilon} upregulates the serine biosynthesis pathway (SBP) mainly by limiting glucose and pyruvate derived anaplerosis of the TCA cycle. In turn, this elicits activation of the transcription factor ATF4 and upregulation of the SBP genes. Importantly, pharmacological inhibition of the IKK{varepsilon}-induced metabolic phenotype reduces proliferation of breast cancer cells. Finally, we show that in a set of basal ER negative and highly proliferative human breast cancer tumours, IKK{varepsilon} and PSAT1 expression levels are positively correlated corroborating the link between IKK{varepsilon} and the SBP in the clinical context.

cancer biology↗

bioRxiv2019

Dynamic trafficking and turnover of Jam-C is essential for endothelial cell migration

Junctional complexes between endothelial cells form a dynamic barrier that hinder passive diffusion of blood constituents into interstitial tissues. Re-modelling of junctions is an essential process during leukocyte trafficking, vascular permeability and angiogenesis. However, for many junctional proteins the mechanisms of junctional remodelling have yet to be determined. Here we used receptor mutagenesis, HRP and APEX-2 proximity labelling, alongside light and electron microscopy, to map the intracellular trafficking routes of junctional adhesion molecule-C (Jam-C). We found that Jam-C co-traffics with receptors associated with changes in permeability, such as VE-Cadherin, NRP-1 and 2, but not with junctional proteins associated with the transmigration of leukocytes. Dynamic Jam-C trafficking and degradation is necessary for junctional remodelling during cell migration and angiogenesis. By identifying new trafficking machinery we show that a key point of regulation is the ubiquitylation of Jam-C by the E3 ligase CBL, this controls the rate of recycling versus lysosomal degradation.

cell biology↗

bioRxiv2019

Spatial Regulation of Polo-like Kinase Activity during C. elegans Meiosis by the Nucleoplasmic HAL-2/HAL-3 Complex

Proper partitioning of homologous chromosomes during meiosis relies on the coordinated execution of multiple interconnected events: Homologs must locate, recognize and align with their correct pairing partners. Further, homolog pairing must be coupled to assembly of the synaptonemal complex (SC), a meiosis-specific tripartite structure that maintains stable associations between the axes of aligned homologs and regulates formation of crossovers between their DNA molecules to create linkages that enable their segregation. Here we identify HAL-3 (Homolog Alignment 3) as an important player in coordinating these key events during C. elegans meiosis. HAL-3 and the previously-identified HAL-2 are interacting and interdependent components of a protein complex that localizes to the nucleoplasm of germ cells. hal-3 (or hal-2) mutants exhibit multiple meiotic prophase defects including failure to establish homolog pairing, inappropriate loading of SC subunits onto unpaired chromosome axes, and premature loss of synapsis checkpoint protein PCH-2. Further, loss of hal function results in misregulation of the subcellular localization and activity of polo-like kinases (PLK-1 and PLK-2), which dynamically localize to different defined subnuclear sites during wild-type prophase progression to regulate distinct cellular events. Moreover, loss of PLK-2 activity partially restores tripartite SC structure in a hal mutant background, suggesting that the defect in pairwise SC assembly in hal mutants reflects inappropriate PLK activity. Together our data support a model in which the nucleoplasmic HAL-2/HAL-3 protein complex constrains both localization and activity of meiotic Polo-like kinases, thereby preventing premature interaction with stage-inappropriate targets.

genetics↗