Biology subjects

Ben-Sahra, I.

Publications and source records attributed to Ben-Sahra, I..

15 recordsLinked to original sources

bioRxiv2026

MLL3/4 methyltransferases regulate the differentiation of pluripotent stem cells via cellular respiration

Enhancer-regulating epigenetic modifiers play critical roles in normal physiological processes and human pathogenesis. The major enhancer regulator paralogs MLL3 and MLL4 (MLL3/4) belong to the lysine methyltransferase 2 (KMT2) family, which catalyzes the methylation of lysine 4 on histone H3 (H3K4me). MLL3/4 are required for enhancer activation and are essential for mammalian development and stem cell differentiation. Although recent studies have linked MLL3/4 with different metabolic pathways in the regulation of stem cell self-renewal and cancer cell growth, the mechanisms connecting enhancer function to metabolic control remain elusive. Here, using respiration flux assays, stable isotope tracing, transcriptomics, and stem cell biology techniques, we show that the loss of MLL3/4 impairs glycolysis and mitochondrial respiration in murine embryonic stem cells. Mechanistically, MLL3/4 deficiency suppresses the expression of the rate-limiting glycolytic enzyme hexokinase 2 (HK2) and compromises the function of the oxoglutarate dehydrogenase (OGDH) complex, thereby coordinately impairing central carbon metabolism. Remarkably, combined restoration of HK2 and OGDH rescues the metabolic defects caused by MLL3/4 loss and reinstates differentiation capacity. Taken together, our study identifies a direct link between enhancer-regulating epigenetic machineries and metabolic control of cell fate transition, providing a mechanistic framework for understanding how enhancer malfunction contributes to developmental abnormalities and human diseases.

developmental biology↗

bioRxiv2025

MELK controls tumor metabolism to promote resistance to melanoma therapy

Melanoma is the most aggressive form of skin cancer, and despite major advances in targeted and immune therapies, durable responses remain limited due to the emergence of resistance mechanisms. Metabolic reprogramming has emerged as a key driver of therapy resistance, allowing tumor cells to adapt to environmental and therapeutic pressures. Here, we identify the maternal embryonic leucine zipper kinase (MELK) as a critical mediator of resistance in melanoma. We demonstrate that MELK stimulates intracellular accumulation of amino acids, leading to activation of mTORC1 signaling and enhanced mitochondrial metabolism and biogenesis. This metabolic shift promotes resistance to immune checkpoint blockade. Importantly, preclinical MELK inhibition sensitizes resistant melanoma cells to immunotherapy, revealing a potential combinatorial strategy to overcome resistance. Our findings establish MELK as a central regulator of metabolic adaptation and therapeutic resistance in melanoma, highlighting its potential as a promising therapeutic target.

cancer biology↗

bioRxiv2025

Rate Limiting Enzymes in Nucleotide Metabolism Synchronize Nucleotide Biosynthesis and Chromatin Formation

Chromatin formation requires both an adequate nucleotide supply and sufficient availability of histones. Chromatin must be assembled following DNA-based fundamental cellular processes such as replication and transcription to preserve genome integrity. Chromatin assembly is regulated by the orderly engagement of histones with a series of histone chaperones that guide newly synthesized histones from ribosomes to DNA. Although the synthesis of nucleotides and the histone proteins are the two major biosynthetic processes that complete the formation of chromatin, how these processes are coordinated remains unknown. Phosphoribosyl pyrophosphate synthetases (PRPSs) catalyze the first and rate-limiting step in the nucleotide biosynthesis pathway. PRPS enzymes form a complex with PRPS-associated proteins (PRPSAPs). In the present study, we discover that PRPS-PRPSAP enzyme complex are part of histone chaperone network. We show that PRPS enzymes are essential not only for nucleotide biogenesis, but together with PRPSAP also play a key role in the early steps in the process of histone maturation by regulating the interaction of histone chaperones with histone H3 and H4. Importantly, this regulation is separate from PRPS nucleotide biosynthetic activity. Depletion of PRPS proteins leads to limited histone availability and impaired chromatin assembly. Our discovery bridges cellular metabolism and chromatin regulation and provides the evidence of how nucleotide biogenesis and histone deposition are coordinated.

cell biology↗

bioRxiv2025

Succinate dehydrogenase activity supports de novo purine synthesis

The de novo purine synthesis pathway is fundamental for nucleic acid production and cellular energetics, yet the role of mitochondrial metabolism in modulating this process remains underexplored. In many cancers, metabolic reprogramming supports rapid proliferation and survival, but the specific contributions of the tricarboxylic acid (TCA) cycle enzymes to nucleotide biosynthesis are not fully understood. Here, we demonstrate that the TCA cycle enzyme succinate dehydrogenase (SDH) is essential for maintaining optimal de novo purine synthesis in normal and cancer cells. Genetic or pharmacological inhibition of SDH markedly attenuates purine synthesis, leading to a significant reduction in cell proliferation. Mechanistically, SDH inhibition causes an accumulation of succinate, which directly impairs the purine biosynthetic pathway. In response, cancer cells compensate by upregulating the purine salvage pathway, a metabolic adaptation that represents a potential therapeutic vulnerability. Notably, co-inhibition of SDH and the purine salvage pathway induces pronounced antiproliferative and antitumoral effects in preclinical models. These findings not only reveal a signaling role for mitochondrial succinate in regulating nucleotide metabolism but also provide a promising therapeutic strategy for targeting metabolic dependencies in cancer.

biochemistry↗

bioRxiv2025

Parallel multiOMIC analysis reveals glutamine deprivation enhances directed differentiation of renal organoids

Metabolic pathways play a critical role in driving differentiation but remain poorly understood in the development of kidney organoids. In this study, parallel metabolite and transcriptome profiling of differentiating human pluripotent stem cells (hPSCs) to multicellular renal organoids revealed key metabolic drivers of the differentiation process. In the early stage, transitioning from hPSCs to nephron progenitor cells (NPCs), both the glutamine and the alanine-aspartate-glutamate pathways changed significantly, as detected by enrichment and pathway impact analyses. Intriguingly, hPSCs maintained their ability to generate NPCs, even when deprived of both glutamine and glutamate. Surprisingly, single cell RNA-Seq analysis detected enhanced maturation and enrichment for podocytes under glutamine-deprived conditions. Together, these findings illustrate a novel role of glutamine metabolism in regulating podocyte development.

bioengineering↗

bioRxiv2025

The extracellular matrix drives guanylate production and protects pancreatic cancer cells from oxaliplatin-induced DNA damage.

The excessive production of extracellular matrix (ECM) and the metabolic adaptations in pancreatic ductal adenocarcinoma (PDAC) contribute individually to enhanced chemoresistance, dramatic tumor progression and dismal patient survival. However, ECM-driven metabolic alterations that promote chemoresistance in PDAC are so far unexplored. Here, we use in-vitro-generated ECM bio-scaffolds that recapitulate cell-ECM interactions and induce broad metabolic alterations in PDAC cells. High-throughput integration of multi-omics datasets coupled with metabolic tracing showed that the ECM enhances the generation of guanylates in PDAC cells, the accumulation of which alleviates oxaliplatin-induced DNA damage, and boosts PDAC cell proliferation. These events are guided by the guanosine monophosphate (GMP)-producing enzymes Impdh and Gmps, the expression of which correlated with that of matrisomal and DNA repair genes in PDAC patient samples. We propose that targeting ECM-driven metabolic processes, like the enhanced Impdh activity, may be an effective therapeutic approach for PDAC patients that bypasses the negative side effects of direct targeting of the ECM itself.

cancer biology↗

bioRxiv2024

Exploiting De Novo Serine Synthesis as a Metabolic Vulnerability to Overcome Sunitinib Resistance in Advanced Renal Cell Carcinoma

Sunitinib, an oral tyrosine kinase inhibitor used in advanced renal cell carcinoma (RCC), exhibits significant efficacy but faces resistance in 30% of patients. Yet, the molecular mechanisms underlying this therapy resistance remain elusive. Here, we show that sunitinib induces a metabolic shift leading to increased serine synthesis in RCC cells. The activation of the GCN2-ATF4 stress response pathway is identified as the mechanistic link between sunitinib treatment and elevated serine production. Inhibiting key enzymes in the serine synthesis pathway, such as PHGDH and PSAT1, enhances the sensitivity of resistant cells to sunitinib. The study underscores the role of serine biosynthesis in nucleotide synthesis, influencing cell proliferation, migration, and invasion. Beyond RCC, similar activation of serine synthesis occurs in other cancer types, suggesting a shared adaptive response to sunitinib therapy. This research identifies serine synthesis as a potential target to overcome sunitinib resistance, offering insights into therapeutic strategies applicable across diverse cancer contexts. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/586287v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@47d331org.highwire.dtl.DTLVardef@17a4892org.highwire.dtl.DTLVardef@1329e51org.highwire.dtl.DTLVardef@37de9b_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LISunitinib induces an increase in endogenous serine production in metastatic ccRCC. C_LIO_LIThe heightened serine biosynthesis promoted by sunitinib facilitates nucleotide synthesis, thereby sustaining tumor cell proliferation. C_LIO_LISunitinib-induced enhancement of serine biosynthesis enables cell migration and invasion. C_LIO_LIThe stimulation in serine synthesis is also observed in other cancer models treated with sunitinib. C_LI

cancer biology↗

bioRxiv2023

HIF-1 inactivation empowers HIF-2 to drive hypoxia adaptation in aggressive forms of medulloblastoma

Medulloblastoma (MB) is the most prevalent brain cancer in children. Four subgroups of MB have been identified; of these, Group 3 is the most metastatic. Its genetics and biology remain less clear than the other groups, and it has a poor prognosis and few effective treatments available. Tumor hypoxia and the resulting metabolism are known to be important in the growth and survival of tumors but, to date, have been only minimally explored in MB. Here we show that Group 3 MB tumors do not depend on the canonical transcription factor hypoxia-inducible factor-1 (HIF-1) to mount an adaptive response to hypoxia. We discovered that HIF-1 is rendered inactive either through post-translational methylation, preventing its nuclear localization specifically in Group 3 MB, or by a low expression that prevents modulation of HIF-target genes. Strikingly, we found that HIF-2 takes over the role of HIF-1 in the nucleus and promotes the activation of hypoxia-dependent anabolic pathways. The exclusion of HIF-1 from the nucleus in Group 3 MB cells enhances the reliance on HIF-2s transcriptional role, making it a viable target for potential anticancer strategies. By combining pharmacological inhibition of HIF-2 with the use of metformin, a mitochondrial complex I inhibitor to block respiration, we effectively induced Group 3 MB cell death, surpassing the effectiveness observed in Non-Group 3 MB cells. Overall, the unique dependence of MB cells, but not normal cells, on HIF-2-mediated anabolic metabolism presents an appealing therapeutic opportunity for treating Group 3 MB patients with minimal toxicity.

cancer biology↗

bioRxiv2023

ASCT2 is the primary serine transporter in cancer cells

The non-essential amino acid serine is a critical nutrient for cancer cells due to its diverse biosynthetic functions. While some tumors can synthesize serine de novo, others are auxotrophic for serine and therefore reliant on the uptake of exogenous serine. Importantly, however, the transporter(s) that mediate serine uptake in cancer cells are not known. Here, we characterize the amino acid transporter ASCT2 (coded for by the gene SLC1A5) as the primary serine transporter in cancer cells. ASCT2 is well-known as a glutamine transporter in cancer, and our work demonstrates that serine and glutamine compete for uptake through ASCT2. We further show that ASCT2-mediated serine uptake is essential for purine nucleotide biosynthesis and that ER promotes serine uptake by directly activating SLC1A5 transcription. Together, our work defines an additional important role for ASCT2 as a serine transporter in cancer and evaluates ASCT2 as a potential therapeutic target in serine metabolism.

cancer biology↗

bioRxiv2021

Ribonucleotide Reductase Regulatory Subunit M2 as a Driver of Glioblastoma TMZ-Resistance through Modulation of dNTP Production

Glioblastoma (GBM) remains one of the most resistant and fatal forms of cancer. Previous studies have examined primary and recurrent GBM tumors, but it is difficult to study tumor evolution during therapy where resistance develops. To investigate this, we performed an in vivo single-cell RNA sequencing screen in a patient-derived xenograft (PDX) model. Primary GBM was modeled by mice treated with DMSO control, recurrent GBM was modeled by mice treated with temozolomide (TMZ), and during therapy GBM was modeled by mice euthanized after two of five TMZ treatments. Our analysis revealed the cellular population present during therapy to be distinct from primary and recurrent GBM. We found the Ribonucleotide Reductase gene family to exhibit a unique signature in our data due to an observed subunit switch to favor RRM2 during therapy. GBM cells were shown to rely on RRM2 during therapy causing RRM2-knockdown (KD) cells to be TMZ-sensitive. Using targeted metabolomics, we found RRM2-KDs to produce less dGTP and dCTP than control cells in response to TMZ (p<0.0001). Supplementing RRM2-KDs with deoxycytidine and deoxyguanosine rescued TMZ-sensitivity, suggesting an RRM2-driven mechanism of chemoresistance, established by regulating the production of these nucleotides. In vivo, tumor-bearing mice treated with the RRM2-inhibitor, Triapine, in combination with TMZ, survived longer than mice treated with TMZ alone (p<0.01), indicating promising clinical opportunities in targeting RRM2. Our data present a novel understanding of RRM2 activity, and its alteration during therapeutic stress as response to TMZ-induced DNA damage.

cancer biology↗

bioRxiv2021

The hexokinase ''HKDC1'' interaction with the mitochondria is essential for hepatocellular carcinoma progression

Hepatocellular carcinoma (HCC) is a leading cause of death from cancer malignancies. Recently, hexokinase domain containing 1 (HKDC1), was shown to have significant overexpression in HCC compared to healthy tissue. Using in vitro and in vivo tools, we examined the role of HKDC1 in HCC progression. Importantly, HKDC1 ablation stops HCC progression by promoting metabolic reprogramming by shifting glucose flux away from the TCA cycle. Next, HKDC1 ablation leads to mitochondrial dysfunction resulting in less cellular energy which cannot be compensated by enhanced glucose uptake. And finally, we show that the interaction of HKDC1 with the mitochondria is essential for its role in HCC progression, and without this mitochondrial interaction mitochondrial dysfunction occurs. In sum, HKDC1 is highly expressed in HCC cells compared to normal hepatocytes, therefore targeting HKDC1, specifically its interaction with the mitochondria, reveals a highly selective approach to target cancer cells in HCC.

cancer biology↗

bioRxiv2021

Hepatic mTORC1 signaling activates ATF4 as part of its metabolic response to feeding and insulin

ObjectiveThe mechanistic target of rapamycin complex 1 (mTORC1) is dynamically regulated by fasting and feeding cycles in the liver to promote protein and lipid synthesis while suppressing autophagy. However, beyond these functions, the metabolic response of the liver to feeding and insulin signaling orchestrated by mTORC1 remains poorly defined. Here, we determine whether ATF4, a stress responsive transcription factor recently found to be independently regulated by mTORC1 signaling in proliferating cells, is responsive to hepatic mTORC1 signaling to alter hepatocyte metabolism. MethodsATF4 protein levels and expression of canonical gene targets were analyzed in the liver following fasting and physiological feeding in the presence or absence of the mTORC1 inhibitor rapamycin. Primary hepatocytes from wild-type or liver-specific Atf4 knockout (LAtf4KO) mice were used to characterize the effects of insulin-stimulated mTORC1-ATF4 function on hepatocyte gene expression and metabolism. Both unbiased steady-state metabolomics and stable-isotope tracing methods were employed to define mTORC1 and ATF4-dependent metabolic changes. RNA-sequencing was used to determine global changes in feeding-induced transcripts in the livers of wild-type versus LAtf4KO mice. ResultsWe demonstrate that ATF4 and its metabolic gene targets are stimulated by mTORC1 signaling in the liver in response to feeding and in a hepatocyte-intrinsic manner by insulin. While we demonstrate that de novo purine and pyrimidine synthesis is stimulated by insulin through mTORC1 signaling in primary hepatocytes, this regulation was independent of ATF4. Metabolomics and metabolite tracing studies revealed that insulin-mTORC1-ATF4 signaling stimulates pathways of non-essential amino acid synthesis in primary hepatocytes, including those of alanine, aspartate, methionine, and cysteine, but not serine. ConclusionThe results demonstrate that ATF4 is a novel metabolic effector of mTORC1 in liver, extending the molecular consequences of feeding and insulin-induced mTORC1 signaling in this key metabolic tissue to the control of amino acid metabolism.

cell biology↗

bioRxiv2021

Determining pathogenicity of variants of uncertain significance and identification of a founder variant in the epilepsy-associated gene, SZT2

Biallelic pathogenic variants in SZT2 result in a neurodevelopmental disorder with shared features, including early-onset epilepsy, developmental delay, macrocephaly, and corpus callosum abnormalities. SZT2 is as a critical scaffolding protein in the amino acid sensing arm of the mTOR signaling pathway. Due to its large size (3432 amino acids), lack of crystal structure, and absence of functional domains, it is difficult to determine the pathogenicity of SZT2 missense and in-frame deletions. We report a cohort of twelve individuals with biallelic SZT2 variants and phenotypes consistent with SZT2-related neurodevelopmental disorder. The majority of this cohort contained one or more SZT2 variants of uncertain significance (VUS). We developed a novel individualized platform to functionally characterize SZT2 VUSs. We identified a recurrent in-frame deletion (SZT2 p.Val1984del) which was determined to be a loss-of-function variant and therefore likely pathogenic. Haplotype analysis determined this single in-frame deletion is a founder variant in those of Ashkenazi Jewish ancestry. Overall, we present a FACS-based rapid assay to distinguish pathogenic variants from VUSs in SZT2, using an approach that is widely applicable to other mTORopathies including the most common causes of the focal genetic epilepsies, DEPDC5, TSC1/2, MTOR and NPRL2/3.

genetics↗

bioRxiv2019

Coessential Genetic Networks Reveal the Organization and Constituents of a Dynamic Cellular Stress Response

The interrelated programs essential for cellular fitness in the face of stress are critical to understanding tumorigenesis, neurodegeneration, and aging. However, modelling the combinatorial landscape of stresses experienced by diseased cells is challenging, leaving functional relationships within the global stress response network incompletely understood. Here, we leverage genome-scale fitness screening data from 625 cancer cell lines, each representing a unique biological context, to build a network of "coessential" gene relationships centered around master regulators of the response to proteotoxic, oxidative, hypoxic, and genotoxic stress. This approach organizes the stress response into functional modules, identifies genes connecting distinct modules, and reveals mechanisms underlying cellular dependence on individual modules. As an example of the power of this approach, we discover that the previously unannotated HAPSTR (C16orf72) promotes resilience to diverse stressors as a stress-inducible regulator of the E3 ligase HUWE1. Altogether, we present a broadly applicable framework and interactive tool (http://fireworks.mendillolab.org/) to interrogate biological networks using unbiased genetic screens.

genetics↗

bioRxiv2019

Aging-related inflammation driven by cellular senescence enhances NAD consumption via activation of CD38+ macrophages

Decline in tissue NAD levels during aging is linked to aging and its associated diseases. However, the mechanism for aging-associated NAD decline remains unclear. Here we report that pro-inflammatory M1-like macrophages, but not naive or M2 macrophages, accumulate in metabolic tissues including visceral white adipose tissue and the liver during aging. Remarkably, these M1-like macrophages highly express the NAD consuming enzyme CD38 and have enhanced CD38-dependent NADase activity. We also find that senescent cells progressively accumulate in visceral white adipose tissue during aging and that inflammatory cytokines found in the supernatant from senescent cells (Senescence associated secretory proteins, SASP) induce macrophages to proliferate and to express CD38. These results highlight a new causal link between visceral tissue senescence and tissue NAD decline during aging and represent a novel therapeutic opportunity targeting maintenance of NAD levels during aging.

immunology↗