bioRxiv : the preprint server for biology最新文献

{"title":"A yeast model reveals the SMAD3 nuclear import mechanism.","authors":"Delfina P González, Morgan C Emokpae, C Patrick Lusk, Mustafa K Khokha","doi":"10.1101/2025.07.09.663975","DOIUrl":"https://doi.org/10.1101/2025.07.09.663975","url":null,"abstract":"<p><p>TGF-β signaling is important for patterning tissues during embryonic development and relies on the nuclear import of SMADs. Basal SMAD nuclear import mechanisms remain to be fully understood and could benefit from advances in our understanding of nuclear transport machinery. To assess the molecular determinants of SMAD nuclear transport, we take advantage of the unicellular <i>S. cerevisiae</i> model that lacks the TGF-β signaling complexity of vertebrates but has conserved nuclear transport machinery. In this minimal system, we find that the steady-state distribution of SMAD2/3, SMAD4, and SMAD1/5 are Ran-dependent and either accumulate in, or are depleted from the nucleus. Conditionally inhibiting each of the karyopherin/importins demonstrates that SMAD3 is imported by orthologues of importin-β, importin-7 and importin-8. The previously defined nuclear localization signal is insufficient to confer nuclear import. Instead, our data suggest the entire MH1 domain may act as an importin binding surface. Deleting a portion of this domain impairs SMAD3 function in <i>Xenopus</i> and SMAD3 nuclear enrichment in HEK293T cells. Thus, the yeast platform provides an efficient strategy to illuminate the nuclear transport mechanisms of embryonic signaling effectors.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265660/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651726","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"On learning functions over biological sequence space: relating Gaussian process priors, regularization, and gauge fixing.","authors":"Samantha Petti, Carlos Martí-Gómez, Justin B Kinney, Juannan Zhou, David M McCandlish","doi":"10.1101/2025.04.26.650699","DOIUrl":"https://doi.org/10.1101/2025.04.26.650699","url":null,"abstract":"<p><p>Mappings from biological sequences (DNA, RNA, protein) to quantitative measures of sequence functionality play an important role in contemporary biology. We are interested in the related tasks of (i) inferring predictive sequence-to-function maps and (ii) decomposing sequence-function maps to elucidate the contributions of individual subsequences. Because each sequence-function map can be written as a weighted sum over subsequences in multiple ways, meaningfully interpreting these weights requires \"gauge-fixing,\" i.e., defining a unique representation for each map. Recent work has established that most existing gauge-fixed representations arise as the unique solutions to$L_2-regularized regression in an overparameterized \"weight space'\" where the choice of regularizer defines the gauge. Here, we establish the relationship between regularized regression in overparameterized weight space and Gaussian process approaches that operate in \"function space,'' i.e. the space of all real-valued functions on a finite set of sequences. We disentangle how weight space regularizers both impose an implicit prior on the learned function and restrict the optimal weights to a particular gauge. We also show how to construct regularizers that correspond to arbitrary explicit Gaussian process priors combined with a wide variety of gauges. Next, we derive the distribution of gauge-fixed weights implied by the Gaussian process posterior and demonstrate that even for long sequences this distribution can be efficiently computed for product-kernel priors using a kernel trick. Finally, we characterize the implicit function space priors associated with the most common weight space regularizers. Overall, our framework unifies and extends our ability to infer and interpret sequence-function relationships.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265701/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651815","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Osteoblast-derived Nerve Growth Factor is Required for Skeletal Adaptation to Mechanical Load and the Osteoanabolic Effect of Gambogic Amide in Mice.","authors":"Ibtesam Rajpar, Eric McLaughlin, Gabriella Fioravanti, Nicholas Ruggiero, Nohael Cherian, Liliana Minichiello, Ryan E Tomlinson","doi":"10.1101/2025.07.08.663521","DOIUrl":"https://doi.org/10.1101/2025.07.08.663521","url":null,"abstract":"<p><p>In adult mice, new bone accrual following mechanical load is mediated by the neurotrophin nerve growth factor (NGF) that is expressed by osteoblasts on the bone surface. NGF can bind to its high affinity receptor, neurotrophic tyrosine kinase receptor type 1 (TrkA), on peripheral sensory nerves resident in bone and support new bone formation. However, the osteoanabolic therapeutic potential of NGF-TrkA signaling to repair bone is limited due to the long-lasting thermal and mechanical hyperalgesia induced by administration of NGF in mice and humans. Here, we investigated whether 1) mature osteoblasts are the primary source of NGF required for bone accrual following loading, and 2) a small molecule TrkA receptor agonist - gambogic amide - can harness the downstream osteoanabolic potential of NGF-TrkA signaling in the absence of endogenous NGF. Loss of <i>Ngf</i> transcription in mature osteoblasts did not appear to affect bone structure or bone mass in adulthood. However, <i>Ngf</i> knockout mice significantly reduced periosteal bone accrual and osteogenic <i>Wnt</i> transcription in response to loading compared to wildtype mice. Intraperitoneal injection of gambogic amide prior to loading was unable to produce its osteoanabolic effects in <i>Ngf</i> knockout mice, suggesting that gambogic amide primarily functions in collaboration with endogenous NGF in bone. In total, our study reveals an important role for osteoblastic NGF in the skeletal adaptation of bone to mechanical forces.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265707/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651817","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Surprising effects of stimulus repetition on neuronal firing rates and gamma-band synchronization in awake macaque V1.","authors":"Martina Pandinelli, Mohsen Parto-Dezfouli, Eleni Psarou, Iris Grothe, Pascal Fries","doi":"10.1101/2025.07.09.663517","DOIUrl":"https://doi.org/10.1101/2025.07.09.663517","url":null,"abstract":"<p><p>Stimulus repetition is abundant, because the environment is redundant and/or because it is redundantly sampled. This offers an opportunity to optimize the processing of repeated stimuli. Indeed, stimulus repetition leads to classically described neuronal response decreases, and to more recently described neuronal gamma synchronization increases (sometimes preceded by decreases for a few trials). Here, we used a full-screen colored background (FSCB) and a area V1 of an awake macaque monkey. We found that the FSCB repetition induced neuronal response increases (sometimes preceded by decreases for a few trials) and gamma synchronization decreases (preceded by increases for a few trials). These effects are largely opposite to the dominant previous findings. Intriguingly, these surprising effects largely reversed when we isolated the responses to the flashed black bar. We discuss these findings, considering differences to previous studies with regards to the subject of the study, the stimuli and the task. We notice that in studies reporting classical results for gamma, sometimes in combination with firing rates, the stimuli were typically (partly) predictive of the reward. Here, we found non-classical results for the FSCB that was not reward predictive, and classical results for the black bar that was reward predictive. Whether this has revealed a general effect of reward predictive versus non-predictive stimuli will require further investigation with stimuli and task designs tailored specifically for this question.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265678/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651953","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Temporal regulation of metabolic processes in the marine diazotroph <i>Crocosphaera watsonii</i> WH 8501.","authors":"Julia M Gauglitz, Keisuke Inomura, Wout Bittremieux, Dawn M Moran, Matthew R McIlvin, Mak A Saito","doi":"10.1101/2025.07.11.664471","DOIUrl":"https://doi.org/10.1101/2025.07.11.664471","url":null,"abstract":"<p><p>Marine diazotrophic cyanobacteria play a crucial role in oceanic nitrogen cycling, supporting primary production and ecosystem balance. <i>Crocosphaera watsonii</i> WH8501 exemplifies this ability by temporally separating photosynthesis and diazotrophy to sustain metabolism. To investigate the regulatory mechanisms underlying this process, we employed LC/MS-MS proteomics in a diel culturing experiment, revealing tightly coordinated protein abundance patterns. Our findings showed a sophisticated temporal regulation of metabolic processes categorized within six distinct protein abundance clusters: (1) nitrogen fixation and amino acid biosynthesis proteins peaked during the night, while (2) glycogen metabolism and dark reactions of photosynthesis were most abundant during the night and day-night transition, likely supporting carbon consumption and energy production. Midday (3 and 4) was dominated by proteins related to photosynthesis, cellular division, and lipid synthesis, whereas late-day peaks (5) in peptide biosynthesis may facilitate nitrogenase complex formation. Notably, the day-night transition (6) exhibited fine-tuned coordination of nitrogenase assembly, with FeS cluster proteins preceding peak nitrogenase iron protein abundance, implying a temporally ordered sequence for functional enzyme formation. Within these categories, sharp temporal patterns emerged in iron trafficking to heme and iron cluster biosynthetic systems, consistent with the need to maintain tight control of iron distribution to metalloproteins at each temporal transition. These results highlight the intricate diel regulation that enables <i>Crocosphaera</i> to balance nitrogen fixation and photosynthesis within a single cell. The observed coordination supports the existence of a complex regulatory system ensuring optimal metabolic performance, reinforcing the critical role of temporal control in sustaining these globally significant biological processes.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265555/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651969","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Context-dependent regulatory variants in Alzheimer's disease.","authors":"Ziheng Chen, Yaxuan Liu, Ashley R Brown, Heather H Sestili, Easwaran Ramamurthy, Xushen Xiong, Dmitry Prokopenko, BaDoi N Phan, Lahari Gadey, Peinan Hu, Li-Huei Tsai, Lars Bertram, Winston Hide, Rudolph E Tanzi, Manolis Kellis, Andreas R Pfenning","doi":"10.1101/2025.07.11.659973","DOIUrl":"https://doi.org/10.1101/2025.07.11.659973","url":null,"abstract":"<p><p>Noncoding genetic variants underlie many complex diseases, yet identifying and interpreting their functional impacts remains challenging. Late-onset Alzheimer's disease (LOAD), a polygenic neurodegenerative disorder, exemplifies this challenge. The disease is strongly associated with noncoding variation, including common variants enriched in microglial enhancers and rare variants that are hypothesized to influence neurodevelopment and synaptic plasticity. These variants often perturb regulatory sequences by disrupting transcription factor (TF) motifs or altering local TF interactions, thereby reshaping gene expression and chromatin accessibility. However, assessing their impact is complicated by the context-dependent functions of regulatory sequences, underscoring the need to systematically examine variant effects across diverse tissues, cell types, and cellular states. Here, we combined <i>in vitro</i> and <i>in vivo</i> massively parallel reporter assays (MPRAs) with interpretable machine-learning models to systematically characterize common and rare variants across myeloid and neural contexts. Parallel profiling of variants in four immune states <i>in vitro</i> and three mouse brain regions <i>in vivo</i> revealed that individual variants can differentially and even oppositely modulate regulatory function depending on cell-type and cell-state contexts. Common variants associated with LOAD tended to exert stronger effects in immune contexts, whereas rare variants showed more pronounced impacts in brain contexts. Interpretable sequence-to-function deep-learning models elucidated how genetic variation leads to cell-type-specific differences in regulatory activity, pinpointing both direct transcription-factor motif disruptions and subtler tuning of motif context. To probe the broader functional consequences of a locus prioritized by our reporter assays and models, we used CRISPR interference to silence an enhancer within the <i>SEC63-OSTM1</i> locus that harbors four functional rare variants, revealing its gatekeeper role in inflammation and amyloidogenesis. These findings underscore the context-dependent nature of noncoding variant effects in LOAD and provide a generalizable framework for the mechanistic interpretation of risk alleles in complex diseases.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265656/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651766","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Fibroblast State Choreographs an Epithelial YAP-dependent Regenerative Program Essential to (Pre)malignancy via ECM-mediated Mechanotransduction.","authors":"Deng Pan, Philippe Gascard, Joseph A Caruso, Chira Chen-Tanyolac, Veena Sangwan, Nicholas Bertos, Sophie Camilleri-Broet, Julie Berube, Spyridon Oikonomopoulos, Michael K Strasser, David L Gibbs, Joanna Bons, Jordan B Burton, Jacob P Rose, Samah Shah, Rosemary Bai, Stuart Lee, Daffolyn Rachael Fels-Elliott, Shoval Miyara, Uri Alon, Anatoly Urisman, Ioannis Ragoussis, Sui Huang, Birgit Schilling, Lorenzo E Ferri, Thea D Tlsty","doi":"10.1101/2025.07.11.661192","DOIUrl":"https://doi.org/10.1101/2025.07.11.661192","url":null,"abstract":"<p><p>Chronic lung injury generates metaplasia which occasionally, but ominously, progresses to squamous dysplasia and squamous lung cancer. To identify mechanisms through which disrupted tissue homeostasis contributes to malignant initiation and progression, we used <i>in vivo</i> and <i>in vitro</i> heterotypic recombinant models of human b ronchial e pithelial c ells (hBECs) and fibroblasts. We demonstrate that injury-associated TGF-β signaling creates a fibroblast state dependent upon HSP47 upregulation. These fibroblasts accumulated collagen, thus elevating tissue stiffness and activating mechanosignaling that sustained YAP-dependent embryonic-like, pro-malignant activities in adjacent hBECs. This S tress/ T ension-Instructive F ibroblast (STIF) state, exhibited by stressed fibroblasts in premalignant and malignant lesions across multiple cancer types, was sufficient to reprogram disease-free hBECs to metaplasia and to drive hBECs with compromised tumor suppressor function to dysplasia, yet could be inhibited and reversed. STIFs suffice to activate epithelial phenotypes reminiscent of oncogene-mediated cell transformation and induce (pre)malignancy via increased force transmission, providing novel targets for prevention.</p><p><strong>Statement of significance: </strong>Tissue injury creates a regenerative pro-tumorigenic S tress/ T ension-Instructive F ibroblast (STIF) state which is sufficient to activate a YAP-dependent, pre-malignant program to induce or unmask pre-cancerous phenotypes in epithelial cells through mechanotransduction. Inhibition of STIF activity or mechanosignaling prevents metaplasia and progression to dysplasia.</p><p><strong>Highlights: </strong>Tissue injury creates a pro-tumorigenic Stress/Tension-Instructive Fibroblast (STIF) state in multiple organs that precedes and persists through cancerSTIF signaling alone, working through fibroblasts and not epithelial cells, is sufficient to activate embryonic-like plasticity and induce epithelial pre-cancerous metaplastic lesionsSTIFs program (pre)malignant phenotypes in adjacent epithelial cells through mechanosignaling by activating YAP prior to tumor formationInhibiting STIFs or mechanosignaling prevents/reverts metaplasia and prevents progression to dysplasia.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265695/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651797","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Active Dissociation of Intracortical Spiking and High Gamma Activity.","authors":"Tianhao Lei, Michael R Scheid, Joshua I Glaser, Marc W Slutzky","doi":"10.1101/2025.07.10.663559","DOIUrl":"https://doi.org/10.1101/2025.07.10.663559","url":null,"abstract":"<p><p>Cortical high gamma activity (HGA) is used in many scientific investigations, yet its biophysical source is a matter of debate. Two leading hypotheses are that HGA predominantly represents summed postsynaptic potentials or-more commonly- predominantly represents summed local spikes. If the latter were true, the nearest neurons to an electrode should contribute most to HGA recorded on that electrode. We trained subjects to decouple spiking from HGA on a single electrode using a brain-machine interface. Their ability to decouple them indicated that HGA is not primarily generated by summed local spiking. Instead, HGA correlated with neuronal population co-firing of neurons that were widely distributed across millimeters. The neuronal spikes that contributed more to this co-firing also contributed more to, and preceded, spike-triggered HGA. These results suggest that HGA arises predominantly from summed postsynaptic potentials triggered by synchronous co-firing of widely distributed neurons.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265708/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651800","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of novel protective loci for executive function using the trail making test part B in the Long Life Family Study.","authors":"Lihua Wang, Katherine Tanner, Stacy L Andersen, Stephanie Cosentino, Vaha Akbary Moghaddam, E Warwick Daw, Jason A Anema, Shiow Jiuan Lin, Acharya Sandeep, Michael Province, Mary K Wojczynski","doi":"10.1101/2025.07.08.663708","DOIUrl":"https://doi.org/10.1101/2025.07.08.663708","url":null,"abstract":"<p><p>The Trail Making Test (TMT) Part B (TMT-B), a well-established assessment of cognitive function, is a frequent component of diagnostic assessments for Mild Cognitive Impairment and dementia in older adults. Identifying the genetic variants associated with the TMT-B will not only gain insights of genetic determinants of cognitive function, but also the molecular mechanisms for dementia. Published GWAS to date for TMT-B suffer from relatively low power due to the use of population level data and imputation methods. To address these deficits, we used a family-based study design to identify the genetic variants associated with the TMT-B incorporating both genome-wide linkage analysis (GWLS) and whole genome sequencing (WGS). As such, we examined the sequenced genetic determinants of TMT-B using GWLS in over 2000 participants from Long Life Family Study (LLFS). In GWLS, the estimated heritability of TMT-B was 0.29. We detected one significant linkage peak at 15q25 (LOD>3.0). Statistical fine-mapping nominated five variants including three SNPs ( <i>NTRK3</i> -rs74031103, protective <i>CEMIP</i> -rs2271159, and protective <i>AGBL1</i> -rs4134376) and two INDELs (protective <i>KLHL25</i> -15:85882445:IND, and protective <i>CEMIP</i> -15:80893381:IND) contributing to the linkage peak. Four out of these five variants are protective for TMT-B. The rs2271159 SNP influences <i>CEMIP</i> expression in cerebellum and hippocampus, while the 15:80893381:IND modulates <i>CEMIP</i> expression in blood. Additionally, the variant rs4134376 is a basal ganglia-specific eQTL for <i>AGBL1</i> . In conclusion, we utilized GWLS, leveraged multi-omics data (whole genome sequence genomic data, transcriptomic data, and lipidomic data), and identified novel protective variants and genes for TMT-B performance.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265606/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651873","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multicellular model of temozolomide resistance in glioblastoma reveals phenotypic shifts in drug response and migratory potential.","authors":"Victoria A Kriuchkovskaia, Ela K Eames, Sydney A McKee, Paul J Hergenrother, Rebecca B Riggins, Brendan A C Harley","doi":"10.1101/2025.07.08.663674","DOIUrl":"https://doi.org/10.1101/2025.07.08.663674","url":null,"abstract":"<p><p>Glioblastoma (GBM) is the most common and aggressive primary malignant brain tumor in adults, with limited survival outcomes due to tumor recurrence, mainly driven by GBM cell invasion and therapy resistance. Although temozolomide (TMZ) remains the standard-of-care chemotherapeutic, its long-term efficacy is often compromised by rapid emergence of acquired resistance, largely mediated by the DNA repair enzyme, methylguanine methyltransferase (MGMT). To investigate the interplay between tumor heterogeneity, drug resistance, and the extracellular matrix (ECM) microenvironment, we adapted a 3D methacrylamide-functionalized gelatin (GelMA) hydrogel model to study the behavior of mixed populations of TMZ-sensitive and TMZ-resistant GBM cells. Using both single-cell distributions and multicellular spheroids, we report the impact of heterogeneous cell populations and TMZ dosing regimens, including physiological, supraphysiological, and metronomic TMZ schedules, on drug response and migration. We show that the combination therapy of TMZ with an MGMT inhibitor, lomeguatrib, can modulate TMZ resistance in vitro. This hydrogel model enables systematic investigation of GBM heterogeneity, go-or-grow phenotypic plasticity, and therapeutic resistance in an ECM-rich microenvironment, offering a valuable platform for future translational research.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12265545/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144651894","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}