Biology subjects

Shen, S.

Publications and source records attributed to Shen, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Choreography of human mitochondrial leaderless mRNA translation initiation

It remains unclear how human mitochondrial ribosomal subunits assemble into an elongation-competent 55S particle on mRNAs devoid of 5 leader sequences. Here, we reconstituted and directly tracked human mitochondrial translation initiation using real-time single-molecule fluorescence spectroscopy. Corroborated with cryo-EM structural analysis, we show that the initiation factor mtIF2 and initiator fMet-tRNAMet are loaded to the 28S subunit to drive mRNAs binding via 5 start codon recognition. This enables sequential loading of the two ribosomal subunits onto the leaderless mRNA to initiate. In parallel, a preassembled 55S monosome can also be loaded with mtIF2 and fMet-tRNAMet to initiate on the mRNA. Both initiation pathways yield active complexes to enter translation elongation, which is gated by mtIF2. The monosome loading pathway can initiate promiscuously with non-formylated Met-tRNAMet, thus its usage may under tight regulation in cells, e.g. by mtIF3. Our work provides a dynamic framework for the distinct human mitochondrial translation initiation.

biochemistry↗

bioRxiv2025

Discovery of Light-Powered Organic Ion Transport by a Natural Protein

Membrane transport proteins play vital roles in living cells by selectively transporting ions and molecules across biological membranes. Among these, microbial rhodopsins are unique in their unparalleled capacity to harness light to drive ion translocation. This distinctive feature has led to their widespread use as optogenetic tools in neuroscience, physiology, and biomed-ical applications. While not all microbial rhodopsins function as ion transporters, many ion-translocating variants have been discov-ered since the identification of the first member--a light-driven H+ pump--in the 1970s. These proteins share a compact structure composed of only seven transmembrane helices and have long been thought to specialize exclusively in transporting small inorganic ions such as H+, Cl-, and Na+. Here, we show that several anion-pumping microbial rhodopsins can also transport organic anions. In particular, a rhodopsin from cyanobacteria is capable of transporting bulky organic anions, including those containing benzene rings, with molecular volumes up to [~]120 [A]3--five times that of Cl-. These organic ions bind to the dark state and are translocated upon photoactivation, following a mechanism similar to that of inorganic anion transport. Mutational analysis indicates that both classes of substrates share a common binding site. Only anions with pKa values below 2 were transported, suggesting that a retained negative charge is essential for binding to the dark state--a prerequisite for transport. This study expands the known sub-strate repertoire of microbial rhodopsins and introduces new possibilities for optogenetic strategies based on light-driven delivery of bioactive organic molecules.

biophysics↗

bioRxiv2025

The titin N2A-MARP signalosome constrains muscle longitudinal hypertrophy in response to stretch

Titin-based mechanosensing is a key driver of trophic signaling in muscle, yet the downstream pathways linking titin sensing to muscle remodeling remain poorly understood. To investigate these signaling mechanisms, we utilized unilateral diaphragm denervation (UDD), an in vivo model that induces titin-stiffness-dependent hypertrophy via mechanical stretch. Using UDD in rats and mice, we characterized the longitudinal hypertrophic response and distinguished stretch-induced signaling from denervation effects by performing global transcriptomic and proteomic analyses following UDD and bilateral diaphragm denervation (BDD) in rats. Our findings identified upregulation of titin-associated muscle ankyrin repeat proteins (MARPs). Subsequent phosphorylation enrichment mass spectrometry in mouse diaphragm highlighted the involvement of the N2A-element. UDD in MARP knockout (KO) mice resulted in enhanced longitudinal hypertrophy, with Western blot analysis revealing activation of the mTOR pathway. Furthermore, pharmacological inhibition of mTORC1 with rapamycin suppressed longitudinal hypertrophy, demonstrating that mTOR signaling regulates titin-mediated hypertrophic growth in a MARP-dependent manner. These findings establish MARPs as key modulators of titin-based mechanotransduction and highlight mTORC1 as a central regulator of longitudinal muscle hypertrophy.

physiology↗

bioRxiv2025

IL-12 Secreting CAR-T Cells Reprogram the Tumor Microenvironment and Improve Efficacy Against Heterogeneous Models of Glioblastoma

BackgroundGlioblastoma (GBM) remains uniformly lethal due to pronounced intratumoral heterogeneity and a highly immunosuppressive microenvironment that limits the efficacy of targeted therapies. MethodsWe engineered chimeric antigen receptor (CAR) T cells targeting EGFRvIII and armored them with a single-chain interleukin-12 (scIL12) payload. These cells were tested in syngeneic, orthotopic GBM mouse models exhibiting heterogeneous EGFRvIII expression. CAR T cells were delivered intracranially without lymphodepletion. ResultsIntracranial administration of of scIL12-secreting CAR-T cells eradicated tumors without requiring lymphodepletion, achieving 50% long-term survival. Survival benefits depended entirely on endogenous CD8 T cells, as efficacy was abolished in CD8-deficient hosts and unaffected by NK cell depletion. Notably, therapeutic efficacy was abrogated by lymphodepletion, underscoring the necessity of an intact endogenous immune response. Mechanistically, scIL12 enhanced CAR-T cell persistence and reprogrammed tumor-associated microglia, indicating potential epitope spreading through polyclonal endogenous CD8+ T-cell responses, which facilitate the elimination of EGFRvIII-negative tumor cells. ConclusionsThis study demonstrates the pleiotropic benefits of IL-12 armored CAR-T cells with improved targeting of antigen-positive tumor cells and simultaneous remodeling of the microenvironment to engage adaptive immunity against antigen-negative clones. This strategy offers a potential clinically actionable approach to improve outcomes in GBM by circumventing the need for toxic lymphodepletion and addressing tumor heterogeneity.

immunology↗

bioRxiv2025

Leveraging Dynamic Stability to Infer Regulation in Protein-Protein Interaction Networks: A Study of Infectious Vulnerability in COPD.

The fourth leading cause of death in the US, Chronic Obstructive Pulmonary Disease (COPD) is punctuated by frequent viral and bacterial infections causing severe acute exacerbations (AECOPD) and increased mortality. In previous work we have shown that altered immune cell signaling may confer increased and persistent susceptibility to infection. Here we continue this investigation by conducting broad-spectrum proteomic profiling of circulating white blood cells to assemble an empirical protein-protein interaction network associated with frequency of infectious exacerbation. In a novel extension of conventional cross-sectional data analyses, we translate these undirected protein-protein interactions into candidate regulatory relationships with both direction and mode of action. The latter are inferred by formulating and solving a constraint satisfaction problem (SAT) whereby predicted dynamic behaviors of any valid regulatory network must support the expected persistent nature of low and high vulnerability phenotypes. Solving this SAT problem produced a set of competing candidate protein regulatory network architectures and signalling rules that unanimously highlighted several novel candidate pathway elements involved in oxidative stress response. Analysis of the overall dynamics supported by these networks, again supported the hypothesis that progression beyond an immune tipping point may confer persistent susceptibility to infection and that this may constitute a stable phenotype or regulatory trap in COPD characterized by a reactive oxygen cascade. HIGHLIGHTSO_LIInfectious vulnerability phenotypes in COPD may represent immune regulatory traps. C_LIO_LIA 15-protein oxidative stress response interactome offers a phenotypic signature. C_LIO_LIHomeostasis in cross-sectional data can support inference of co-regulatory actions. C_LIO_LI8 of 11 documented regulatory actions recovered in candidate model majority vote C_LIO_LI17 new candidate protein co-regulatory actions proposed unanimously by all models. C_LI

systems biology↗

bioRxiv2025

Quantitative RNA pseudouridine landscape reveals dynamic modification patterns and evolutionary conservation across bacterial species

Pseudouridine ({Psi}) modifications are the most abundant RNA modifications; however, their distribution and functional significance in bacteria remain largely unexplored compared to eukaryotic systems. In this study, we present the first transcriptome-wide and quantitative mapping of {Psi} modifications across five diverse bacterial species (Bacillus cereus, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Pseudomonas syringae) at single-base resolution, utilizing the optimized BID-seq method for bacterial RNA. Our analysis revealed growth phase-dependent dynamics of pseudouridylation in bacterial tRNA and mRNA, particularly in genes enriched in core metabolic pathways. Comparative analysis demonstrated evolutionarily conserved features of {Psi} modifications, such as dominant motif contexts, {Psi} clustering within operons, etc. Functional analysis indicated {Psi} modifications influence bacterial mRNA stability, translation, and interactions with specific RNA-binding proteins (RBPs) in response to changing cellular demands during growth phase transitions. The integrated computational analysis on local RNA architecture was conducted to elucidate the structure-dependent {Psi} modifications in bacterial RNA. Furthermore, we developed an integrated deep learning framework, combining Transformer-GNN-based neural networks (pseU_NN) to capture both RNA sequence and structural features for effective prediction of {Psi}-modified sites. Overall, our study provides valuable insights into the landscapes of bacterial RNA {Psi} modifications and establishes a foundation for future mechanistic investigations into the functions of {Psi} in bacterial RNA regulation.

microbiology↗

bioRxiv2025

Proteasome Inhibition Enhances Lysosome-mediated Targeted Protein Degradation

Proteasome inhibitor drugs are currently used in the clinic to treat multiple myeloma and mantle cell lymphoma. These inhibitors cause accumulation of undegraded proteins, thus inducing proteotoxic stress and consequent cell death. However, cancer cells counteract this effect by activating an adaptive response through the transcription factor Nuclear factor erythroid 2-related factor 1 (NRF1, also known as NFE2L1). NRF1 induces transcriptional upregulation of proteasome and autophagy/lysosomal genes, thereby reducing proteotoxic stress and diminishing the effectiveness of proteasome inhibition. While suppressing this protective autophagy response is one potential strategy, here we investigated whether this heightened autophagy could instead be leveraged therapeutically. To this end, we designed an autophagy-targeting chimera (AUTAC) compound to selectively degrade the anti-apoptotic protein Mcl1 via the lysosome. Our results show that this lysosome-mediated targeted degradation is significantly amplified in the presence of proteasome inhibition, in a NRF1-dependent manner. Mechanistically, AUTAC-driven Mcl1 clearance requires K63-linked ubiquitination by UBC13 and TRAF6 and recognition by the cargo receptor p62/SQSTM1. The combination of the proteasome inhibitor carfilzomib and Mcl1 AUTAC synergistically promoted cell death in both in vitro models, including wild-type and proteasome inhibitor-resistant multiple myeloma and lung cancer cells, and in mouse tumor xenografts. Thus, our work offers a novel strategy for enhancing proteasome inhibitor efficacy by exploiting the adaptive autophagy response. More broadly, our study establishes a framework for amplifying lysosome-mediated targeted protein degradation, with potential applications in cancer therapeutics and beyond.

biochemistry↗

bioRxiv2024

A gene co-expression network-based analysis of mesenchymal stromal cells reveals novel genes and molecular pathways underlying heterotopic ossification

BackgroundHeterotopic ossification (HO) represents a frequently seen refractory disease second to musculoskeletal injury, in which bone tissue ectopically exists within soft tissue, resulting in serious extremity loss-of-function. This work focused on identifying regulating factors and gene network associated with HO pathology. Material and MethodsWe randomized the heterotopic ossification dataset GSE94683 as HO and NON-HO group. Weighted gene co-expression network analysis (WGCNA) was applied in identifying HO related modules. We discovered differentially expressed genes (DEGs) between the two groups. Then, we integrated protein-protein interaction (PPI) network, co-expression network, enrichment analysis, gene set variation analysis (GSVA) and gene set enrichment analysis (GSEA) for identifying the HO-related pathways. We also employed GSE126118 test set to investigate hub genes related to HO. Eventually, potential therapeutics for reversing abnormal hub gene levels were predict by DGIdb. ResultsWe discovered twelve HO status modules and 1,483 DEGs in HO samples compared with NON-HO counterparts. Four hub genes were obtained from the overlap of HO related PPI and coexpression networks. Training and test sets were adopted for verification, and three abnormally high hub gene expression in HO were obtained. Functional enrichment of DEGs indicated that these genes were involved in the stem cell pluripotency regulatory pathways, cytokine-cytokine receptor interaction, as well as rap1 pathway. Based on the results from GSEA and GSVA, many hub gene sets were mainly enriched in immune responses. ConclusionWe identified a gene coexpression network associated with regulatory factors in HO, which provided a new perspective on the pathogenesis and provided potential biomarkers or therapeutic targets.

bioinformatics↗

bioRxiv2024

Sirt5 preserves cardiac function in ischemia-reperfusion injury by inhibiting ANT2 lactylation

Myocardial ischemia-reperfusion (IR) injury is the most common type of heart disease. IR-disrupted mitochondrial homeostasis affects heart energy metabolism and function, but the mechanism remains poorly understood. Here we report a cardioprotective role of the mitochondrial protein Sirt5 in a mouse model of cardiac IR injury. The down-regulation of Sirt5 in IR-injured heart is associated with an increased protein lactylation and impaired mitochondrial function. Overexpression of Sirt5 alleviates, whereas conditional knockout of Sirt5 aggravates, mitochondrial damage and cardiac injury. Mechanistically, Sirt5 interacts with the inner mitochondrial membrane protein adenine nucleotide translocase 2 (ANT2), inhibiting its lysine lactylation to promote its association with the outer mitochondrial membrane protein voltage-dependent anion-selective channel 1 (VDAC1). Lactylation-resistant ANT2 efficiently complexes with VDAC1 and improves cardiac function after injury. Therefore, we uncover a Sirt5-regulated interaction of ANT2 and VDAC1 in maintaining mitochondrial homeostasis, highlighting the potential of targeting ANT2 lactylation as a therapeutic strategy in the treatment of cardiac injury. TeaserSirt5 promotes the interaction of mitochondrial proteins to protect mitochondrial homeostasis and improve heart function in ischemia-reperfusion injury.

pathology↗

bioRxiv2024

From movements to words: action monitoring in the medial frontal cortex along a caudal to rostral prediction error gradient

Speech error monitoring recruits the medial frontal cortex (MFC) region in the human brain. Error monitoring-related activity in the MFC has been interpreted both in terms of conflict monitoring and feedback-driven control, but as similar regions of the MFC are implicated in various levels of behavioral control ranging from basic motor movement control to high-level cognitive control functions, a more comprehensive account is needed. Moreover, as speech errors and other actions that involve varying control demands engage a widespread yet partially overlapping set of regions of the MFC, such an account should ideally explain the anatomical distribution of error-related functional activations within the MFC. Here we wanted to assess the hypothesis that the MFC has a similar role in the evaluation of action outcomes for motor and mental actions, operating along a rostral-caudal gradient of higher-lower level control demands involving prediction errors from both sensory and epistemic sources. To this end, we conducted an individual-specific annotation of task-fMRI BOLD activation peaks related to speech errors versus correct productions (i.e. that involve the largest cognitive control demands, Study I and II), tongue movement monitoring (i.e. that involve an intermediate level of cognitive and motor control demands) and tongue movement (i.e. that involve only motor control demands, Study II) in the MFC region. Results revealed overlapping clusters across the three contrasts across the MFC, but importantly both the number of peaks and their relative position along the rostral caudal axis were consistent with a hierarchical rostral caudal processing gradient in the MFC. While tongue movement showed more caudal activation in the MFC, speech errors showed more rostral activation, and tongue movement monitoring patterned in between. Furthermore, the combined results of both studies suggested that activation peaks were located more dorsally for participants that had a paracingulate gyrus, replicating a previously documented effect for movement and further supporting a common functional role of the MFC across very distinct actions.

neuroscience↗

bioRxiv2024

CYP3A5 inhibition causes G1/S blockade and synergizes with CDK4/6 inhibitor to suppress prostate cancer cell growth: Implications in reducing health disparity

Prostate cancer (PC) is a leading cause of death in men because of the high incidence and long-term inefficacy of the existing treatment options. Furthermore, it exhibits significant health disparities that affect African-American (AA) men more adversely than others do. Previously, we established CYP3A5, a highly expressed protein in AAs PC, as a positive regulator of androgen receptor (AR) signaling. We examined the impact of CYP3A5 depletion on genome-wide transcriptional output using RNA sequencing to gain deeper mechanistic insights. The data revealed that 561 genes were downregulated and 263 were upregulated upon silencing of CYP3A5 in PC cells. Furthermore, in silico pathway analyses of differentially expressed genes suggested that the cell cycle regulation pathway was most significantly affected by CYP3A5 inhibition. Cell cycle analysis of CYP3A5-silenced cells and those treated with clobetasol, a specific CYP3A5 pharmacological inhibitor, showed G1/S phase blockade. Both CYP3A5-depletion and pharmacological inhibition resulted in the downregulation of cyclin D, cyclin B, and CDK2, along with the upregulation of p27kip1 but had minimal effects on CDK4/6 levels. Combination treatment with clobetasol and the CDK4/6 inhibitor palbociclib exhibited synergy with combination index (CI) values ranging from 0.28-0.78. Our findings support the utility of CYP3A5 as a druggable therapeutic target that works more effectively in combination with CDK4/6 inhibition to limit the progression of PC, especially for AA patients with AA. This combination addresses CDK4/6 inhibitor resistance, which is often linked to CDK2 overexpression, and can potentially be useful in reducing disparities in the clinical outcomes of PC. SignificanceOur study highlights CYP3A5 as a key regulator of the cell cycle in prostate cancer (PC). Its overexpression in African American (AA) patients may be a key molecular driver of disparities in outcomes. The combination of CYP3A5 and CDK4/6 inhibitors shows a synergistic effect on therapeutic outcomes and addresses CDK2-mediated resistance. Thus, targeting both CYP3A5 and CDK4/6 could improve treatment outcomes, especially in AA PC patients.

cancer biology↗

bioRxiv2024

Multi-omics analysis reveals the mechanism of BnC07MYB3a is involved in seed coat color in Brassica napus L.

In rapeseed (Brassica napus), yellow-seeded varieties accumulate less flavonoid pigments (anthocyanins/proanthocyanidins) in their seed coats compared with black-seeded varieties. The yellow-seeded trait is associated with greatly improved seed oil yield, quality, and commercial value. Many R2R3 MYB activators have been characterized in rapeseed, but how MYB-type repressors affect pigment biosynthesis is not yet fully understood. In this study, we performed transcriptome sequencing and metabolomic analysis of B. napus varieties with extreme differences in seed coat color, combined with weighted gene co-expression network analysis. This analysis identified an R2R3-MYB-type transcription factor, BnC07MYB3a (BnaC07G0178800ZS), as a candidate regulator of the yellow-seeded trait in B. napus. Overexpressing BnC07MYB3a in Arabidopsis thaliana and B. napus downregulated the expression of flavonoid biosynthetic genes, resulting in significantly lower anthocyanin and proanthocyanidin accumulation than in the wild-type and a lighter seed coat color in transgenic plants. BnC07MYB3a directly binds to the promoter of the TRANSPARENT TESTA (TT) gene BnTT6 and represses its expression. BnC07MYB3a also physically interacts with BnA06bHLH92a and the WD40 transcription factor TRANSPARENT TESTA GLABRA1 (BnTTG1), suggesting that they might form a previously unidentified MYB-bHLH-WD40 transcription factor complex. Our results reveal the molecular mechanism and regulatory network of BnC07MYB3a in determining seed coat color in B. napus and provide a genetic resource for breeding yellow-seeded cultivars of B. napus.

molecular biology↗

bioRxiv2024

Expression-based selection identifies a microglia-tropic AAV capsid for direct and CSF routes of administration in mice

Microglia are critical innate immune cells of the brain. In vivo targeting of microglia using gene-delivery systems is crucial for studying brain physiology and developing gene therapies for neurodegenerative diseases and other brain disorders such as NeuroAIDS. Historically, microglia have been extremely resistant to transduction by viral vectors, including adeno-associated virus (AAV) vectors. Recently, there has been some progress demonstrating the feasibility and potential of using AAV to transduce microglia after direct intraparenchymal vector injection. Data suggests that combining specific AAV capsids with microglia-specific gene expression cassettes to reduce neuron off-targeting will be key. However, no groups have developed AAV capsids for microglia transduction after intracerebroventricular (ICV) injection. The ICV route of administration has advantages such as increased brain biodistribution while avoiding issues related to systemic injection. Here, we performed an in vivo selection using an AAV peptide display library that enables recovery of capsids that mediate transgene expression in microglia. Using this approach, we identified a capsid, MC5, which mediated enhanced transduction of microglia after ICV injection compared to AAV9. Furthermore, MC5 enhanced both the efficiency (85%) and specificity (93%) of transduction compared to a recently described evolved AAV9 capsid for microglia targeting after direct injection into the brain parenchyma. Exploration of the use of MC5 in a mouse models of Alzheimers disease revealed transduced microglia surrounding and within plaques. Overall, our results demonstrate that the MC5 capsid is a useful gene transfer tool to target microglia in vivo by direct and ICV routes of administration.

bioengineering↗

bioRxiv2024

Ethograms reveal a fear conditioned visual cue to organize diverse behaviors in rats

Recognizing and responding to threat cues is essential to survival. In rats, freezing is the most common behavior measured. Previously we demonstrated a threat cue can organize diverse behaviors (Chu et al., 2024). However, the experimental design of Chu et al. (2024) was complex and the findings descriptive. Here, we gave female and male Long Evans rats simple paired or unpaired presentations of a light and foot shock (8 total) in a conditioned suppression setting, using a range of shock intensities (0.15, 0.25, 0.35 or 0.5 mA). We found that conditioned suppression was only observed at higher foot shock intensities (0.35 mA and 0.5 mA). We constructed comprehensive, temporal ethograms by scoring 22,272 frames of behavior for 12 mutually exclusive behavior categories in 200 ms intervals around cue presentation. A 0.5 mA and 0.35 mA shock-paired visual cue suppressed reward seeking, rearing and scaling, as well as light-directed rearing and light-directed scaling. The shock-paired visual paired cue further elicited locomotion and freezing. Linear discriminant analyses showed that ethogram data could accurately classify rats into paired and unpaired groups. Considering the complete ethogram data produced superior classification than considering subsets of behaviors. The results demonstrate diverse threat-elicited behaviors - in a simple Pavlovian fear conditioning design - containing sufficient information to distinguish the fear learning status of individual rats.

neuroscience↗

bioRxiv2024

Center-surround inhibition by expectation: a neuro-computational account

Expectation is beneficial for adaptive behavior through quickly deducing plausible interpretations of information. The profile and underlying neural computations of this process, however, remain unclear. When participants expected a grating with a specific orientation, we found a center-surround inhibition profile in orientation space, which was independent from attentional modulations by task-relevance. Using computational modeling, we showed that this center-surround inhibition could be reproduced by either a sharpening of tuning curves of expected orientation or a shift of tuning curves of unexpected orientations. Intriguingly, these two computations were further supported by orientation-adjustment and orientation-discrimination experiments. Finally, the ablation studies in convolutional neural networks revealed that predictive coding feedback played a critical role in the center-surround inhibition in expectation. Altogether, our study reveals for the first time that expectation results in both enhancement and suppression, optimizing plausible interpretations during perception by enhancing expected and attenuating similar but irrelevant and potentially interfering representations.

neuroscience↗

bioRxiv2024

Targeting of SUMOylation leads to cBAF complex stabilization and disruption of the SS18::SSX transcriptome in Synovial Sarcoma

Synovial Sarcoma (SS) is driven by the SS18::SSX fusion oncoprotein. and is ultimately refractory to therapeutic approaches. SS18::SSX alters ATP-dependent chromatin remodeling BAF (mammalian SWI/SNF) complexes, leading to the degradation of canonical (cBAF) complex and amplified presence of an SS18::SSX-containing non-canonical BAF (ncBAF or GBAF) that drives an SS-specific transcription program and tumorigenesis. We demonstrate that SS18::SSX activates the SUMOylation program and SSs are sensitive to the small molecule SAE1/2 inhibitor, TAK-981. Mechanistically, TAK-981 de-SUMOylates the cBAF subunit SMARCE1, stabilizing and restoring cBAF on chromatin, shifting away from SS18::SSX-ncBAF-driven transcription, associated with DNA damage and cell death and resulting in tumor inhibition across both human and mouse SS tumor models. TAK-981 synergized with cytotoxic chemotherapy through increased DNA damage, leading to tumor regression. Targeting the SUMOylation pathway in SS restores cBAF complexes and blocks the SS18::SSX-ncBAF transcriptome, identifying a therapeutic vulnerability in SS, positioning the in-clinic TAK-981 to treat SS.

cancer biology↗

bioRxiv2024

Genome-Wide CRISPR-Cas9 Screening Identifies a Synergy between Hypomethylating Agents and SUMOylation Blockade in MDS/AML

Hypomethylating agents (HMAs) are frontline therapies effective at altering the natural course of Myelodysplastic Neoplasms (MDS) and Acute Myeloid Leukemia (AML). However, acquired resistance and treatment failure are hallmarks of HMA therapy. To address this clinical need, we performed a genome-wide CRISPR-Cas9 screen in a human MDS-derived cell line, MDS-L, and identified TOPORS as a highly ranked loss-of-function target that synergizes with HMAs, reducing leukemic burden and improving survival in xenograft models. We demonstrate that the depletion of TOPORS mediates sensitivity to HMAs by predisposing leukemic blasts to an impaired DNA damage response (DDR) accompanied by an accumulation of SUMOylated DNMT1 in HMA-treated TOPORS-depleted cells. Importantly, the combination of HMAs with targeting of TOPORS did not functionally impair healthy hematopoiesis. While inhibitors of TOPORS are currently unavailable, we show that inhibition of protein SUMOylation (upstream of TOPORS functions) with TAK-981 partially phenocopies HMA-sensitivity and DDR impairment. Overall, our data suggest that the combination of HMAs with the inhibition of SUMOylation or TOPORS demonstrates a favourable therapeutic index and is a rational treatment framework for High-Risk MDS (HR-MDS) or AML.

molecular biology↗

bioRxiv2024

Loss of prohibitin 2 in Schwann cells dysregulates key transcription factors controlling developmental myelination

Schwann cells are critical for the proper development and function of the peripheral nervous system, where they form a mutually beneficial relationship with axons. Past studies have highlighted that a pair of proteins called the prohibitins play major roles in Schwann cell biology. Prohibitins are ubiquitously expressed and versatile proteins. We have previously shown that while prohibitins play a crucial role in Schwann cell mitochondria for long-term myelin maintenance and axon health, they may also be present at the Schwann cell-axon interface during development. Here, we expand on this work, showing that drug-mediated modulation of prohibitins in vitro disrupts myelination and confirming that Schwann cell-specific ablation of prohibitin 2 (Phb2) in vivo results in early and severe defects in peripheral nerve development. Using a proteomic approach in vitro, we identify a pool of candidate PHB2 interactors that change their interaction with PHB2 depending on the presence of axonal signals. Furthermore, we show in vivo that loss of Phb2 in mouse Schwann cells causes ineffective proliferation and dysregulation of transcription factors EGR2 (KROX20), POU3F1 (OCT6) and POU3F2 (BRN2) that are necessary for proper Schwann cell maturation. Schwann cell-specific deletion of Jun, a transcription factor associated with negative regulation of myelination, confers partial rescue of the development defect seen in mice lacking Schwann cell Phb2. This work develops our understanding of Schwann cell biology, revealing that Phb2 may directly or indirectly modulate the timely expression of transcription factors necessary for proper peripheral nervous system development, and proposing candidates that may play a role in PHB2-mediated integration of axon signals in the Schwann cell.

neuroscience↗