Biology subjects

Shah, S.

Publications and source records attributed to Shah, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2024

Measuring the burden of hundreds of BioBricks defines an evolutionary limit on constructability in synthetic biology

Engineered DNA will slow the growth of a host cell if it redirects limiting resources or otherwise interferes with homeostasis. Populations of engineered cells can rapidly become dominated by "escape mutants" that evolve to alleviate this burden by inactivating the intended function. Synthetic biologists working with bacteria rely on genetic parts and devices encoded on plasmids, but the burden of different engineered DNA sequences is rarely characterized. We measured how 301 BioBricks on high-copy plasmids affected the growth rate of Escherichia coli. Of these, 59 (19.6%) negatively impacted growth. The burden imposed by engineered DNA is commonly associated with diverting ribosomes or other gene expression factors away from producing endogenous genes that are essential for cellular replication. In line with this expectation, BioBricks exhibiting burden were more likely to contain highly active constitutive promoters and strong ribosome binding sites. By monitoring how much each BioBrick reduced expression of a chromosomal GFP reporter, we found that the burden of most, but not all, BioBricks could be wholly explained by diversion of gene expression resources. Overall, no BioBricks reduced the growth rate of E. coli by >45%, which agreed with a population genetic model that predicts such plasmids should be "unclonable" because escape mutants will take over during growth of a bacterial colony or small laboratory culture from a transformed cell. We made this model available as an interactive web tool for synthetic biology education and added our burden measurements to the iGEM Registry descriptions of each BioBrick.

synthetic biology↗

bioRxiv2024

Thermal tolerance of mosquito eggs is associated with urban adaptation and human interactions

Climate change is expected to profoundly affect mosquito distributions and their ability to serve as vectors for disease, specifically with the anticipated increase in heat waves. The rising temperature and frequent heat waves can accelerate mosquito life cycles, facilitating higher disease transmission. Conversely, higher temperatures could increase mosquito mortality as a negative consequence. Warmer temperatures are associated with increased human density, suggesting a need for anthropophilic mosquitoes to adapt to be more hardy to heat stress. Mosquito eggs provide an opportunity to study the biological impact of climate warming as this stage is stationary and must tolerate temperatures at the site of female oviposition. As such, egg thermotolerance is critical for survival in a specific habitat. In nature, Aedes mosquitoes exhibit different behavioral phenotypes, where specific populations prefer depositing eggs in tree holes and prefer feeding non-human vertebrates. In contrast, others, particularly human-biting specialists, favor laying eggs in artificial containers near human dwellings. This study examined the thermotolerance of eggs, along with adult stages, for Aedes aegypti and Ae. albopictus lineages associated with known ancestry and shifts in their relationship with humans. Mosquitoes collected from areas with higher human population density, displaying increased human preference, and having a human-associated ancestry profile have increased egg viability following high-temperature stress. Unlike eggs, thermal tolerance among adults showed no significant correlation based on the area of collection or human-associated ancestry. This study highlights that the egg stage is likely critical to mosquito survival when associated with humans and needs to be accounted when predicting future mosquito distribution.

ecology↗

bioRxiv2024

Androgen receptor-negative prostate cancer is vulnerable to SWI/SNF-targeting degrader molecules

Proteolysis targeting chimera (PROTAC) therapies degrading SWI/SNF ATPases offer a novel approach to interfere with androgen receptor (AR) signaling in AR-dependent castration-resistant prostate cancer (CRPC-AR). To explore the utility of SWI/SNF therapy beyond AR-sensitive CRPC, we investigated SWI/SNF-targeting agents in AR-negative CRPC. SWI/SNF targeting PROTAC treatment of cell lines and organoid models reduced the viability of not only CRPC-AR but also WNT-signaling dependent AR- negative CRPC (CRPC-WNT). The CRPC-WNT subgroup represents 11% of around 400,000 cases of CRPC worldwide who die yearly of CRPC. We discovered that SWI/SNF ATPase SMARCA4 depletion interfered with the master transcriptional regulator TCF7L2 (TCF4) in CRPC-WNT. Functionally, TCF7L2 maintains proliferation via the MAPK signaling axis in this subtype of CRPC. These data suggest a mechanistic rationale for interventions that perturb the DNA binding of the pro-proliferative TCF7L2 transcription factor (TF) and/or direct MAPK signaling inhibition in the CRPC-WNT subclass of advanced prostate cancer. Statement of significanceAndrogen receptor (AR)-negative prostate cancer (PCa) remains a clinical challenge due to the lack of targeted therapeutic options. Here, we identified a lineage-defining molecular axis in a subtype of AR- negative PCa, accounting for around 10% of castration-resistant PCa (CRPC) that can be interfered with by SWI/SNF-targeting agents.

cancer biology↗

bioRxiv2024

Adaptation of Pseudomonas aeruginosa to repeated invasion into a commensal competitor

The host-associated microbiome is an important barrier to bacterial pathogen colonization and can mediate protection through a variety of mechanisms. We wanted to investigate the potential consequences of selection imposed by commensal bacterial competitors on an invading bacterial pathogen. To do this, we tested the ability of the opportunistic pathogen Pseudomonas aeruginosa to invade pre-established communities of an abundant commensal bacterium in the human microbiome, Staphylococcus epidermidis. We passaged ten independent lines of P. aeruginosa through daily invasion into a pre-established S. epidermidis population (coculture evolved lines), alongside daily passage through monoculture conditions (monoculture evolved lines). The monoculture evolved lines showed strong parallel evolution in the Wsp (Wrinkly spreader phenotype) signal transducing system involved in biofilm formation, and significantly elevated biofilm formation. On the other hand, adaptation to S. epidermidis occurred via mutations in a diverse set of genes, and the coculture evolved lines showed much weaker evidence for parallel evolution, suggesting that the selective pressure imposed by competition with S. epidermidis is more complex than the pressure imposed by culture conditions. Interestingly, the elevated biofilm formation phenotype seen in the monoculture evolved lines was not observed in the lines evolved in the presence of S. epidermidis, raising the question of whether enhanced biofilm formation did not evolve with S. epidermidis present because it was not beneficial, or because S. epidermidis may be able to restrict this evolutionary path by inhibiting biofilm formation.

microbiology↗

bioRxiv2024

Structural insights into CXCR4 modulation and oligomerization

Activation of the chemokine receptor CXCR4 by its chemokine ligand CXCL12 regulates diverse cellular processes. CXCR4 also serves as a key target for diseases such as cancer and HIV. Previously reported crystal structures of CXCR4 bound to antagonists revealed the architecture of an inactive, homodimeric receptor. However, many structural aspects of CXCR4 remain poorly understood, including its activation by CXCL12, as well as its assembly into higher-order oligomers. Here, we use cryoelectron microscopy (cryoEM) to investigate various modes of CXCR4 regulation in the presence and absence of Gi protein. CXCL12 activates CXCR4 by inserting its N-terminus deep into the CXCR4 orthosteric pocket. The binding of FDA-approved antagonist AMD3100 is stabilized by electrostatic interactions with acidic residues in the 7 transmembrane helix bundle. A potent antibody blocker, REGN7663, binds across the extracellular face of CXCR4 and inserts its CDR-H3 loop into the orthosteric pocket. Trimeric and tetrameric structures of CXCR4 reveal, to our knowledge, previously undescribed modes of GPCR oligomerization. Remarkably, CXCR4 adopts distinct subunit conformations in trimeric and tetrameric assemblies, highlighting how oligomerization could allosterically regulate chemokine receptor function.

biochemistry↗

bioRxiv2024

Homologous recombination deficiency and tumor suppressor heterozygosity mediate resistance to front-line therapy in breast cancer

The co-occurrence of germline and somatic oncogenic alterations is frequently observed in breast cancer, but their combined biologic and clinical significance has not been evaluated. To assess the role of germline-somatic interactions on outcomes in routine practice, we developed an integrated clinicogenomic pipeline to analyze the genomes of over 4,500 patients with breast cancer. We find that germline (g)BRCA2-associated tumors are enriched for RB1 loss-of-function mutations and manifest poor outcomes on standard-of-care, front-line CDK4/6 inhibitor (CDK4/6i) combinations. Amongst these tumors, gBRCA2-related homologous recombination deficiency (HRD) as well as baseline RB1 LOH status promote acquisition of RB1 loss-of- function mutations under the selective pressure of CDK4/6i, causing therapy resistance. These findings suggest an alternative therapeutic strategy using sequential targeting of HRD in gBRCA- associated breast cancers through PARP inhibitors prior to CDK4/6i therapy to intercept deleterious RB1-loss trajectories and thus suppress the emergence of CDK4/6 inhibitor resistance. More broadly, our findings demonstrate how germline-somatic driven genomic configurations shape response to systemic therapy and can be exploited therapeutically as part of biomarker-directed clinical strategies.

genomics↗

bioRxiv2024

A large-scale cancer-specific protein-DNA interaction network

Cancer development and progression are generally associated with dysregulation of gene expression, often resulting from changes in transcription factor (TF) sequence or expression. Identifying key TFs involved in cancer gene regulation provides a framework for potential new therapeutics. This study presents a large-scale cancer gene TF-DNA interaction network as well as an extensive promoter clone resource for future studies. Most highly connected TFs do not show a preference for binding to promoters of genes associated with either good or poor cancer prognosis, suggesting that emerging strategies aimed at shifting gene expression balance between these two prognostic groups may be inherently complex. However, we identified potential for oncogene targeted therapeutics, with half of the tested oncogenes being potentially repressed by influencing specific activator or bifunctional TFs. Finally, we investigate the role of intrinsically disordered regions within the key cancer-related TF estrogen receptor [a] (ESR1) on DNA binding and transcriptional activity, and found that these regions can have complex trade-offs in TF function. Altogether, our study not only broadens our knowledge of TFs involved in the cancer gene regulatory network but also provides a valuable resource for future studies, laying a foundation for potential therapeutic strategies targeting TFs in cancer.

systems biology↗

bioRxiv2024

Heparan sulfate modified proteins affect cellular processes central to neurodegeneration and modulate presenilin function

Mutations in presenilin-1 (PSEN1) are the most common cause of familial, early-onset Alzheimers disease (AD), typically producing cognitive deficits in the fourth decade. A variant of APOE, APOE3 Christchurch (APOE3ch), was found associated with protection from both cognitive decline and Tau accumulation in a 70-year-old bearing the disease-causing PSEN1-E280A mutation. The amino acid change in ApoE3ch is within the heparan sulfate (HS) binding domain of APOE, and purified APOEch showed dramatically reduced affinity for heparin, a highly sulfated form of HS. The physiological significance of ApoE3ch is supported by studies of a mouse bearing a knock-in of this human variant and its effects on microglia reactivity and A{beta}-induced Tau deposition. The studies reported here examine the function of heparan sulfate-modified proteoglycans (HSPGs) in cellular and molecular pathways affecting AD-related cell pathology in human cell lines and mouse astrocytes. The mechanisms of HSPG influences on presenilin-dependent cell loss and pathology were evaluated in Drosophila using knockdown of the presenilin homolog, Psn, together with partial loss of function of sulfateless (sfl), a homolog of NDST1, a gene specifically affecting HS sulfation. HSPG modulation of autophagy, mitochondrial function, and lipid metabolism were shown to be conserved in cultured human cell lines, Drosophila, and mouse astrocytes. RNAi of Ndst1 reduced intracellular lipid levels in wild-type mouse astrocytes or those expressing humanized variants of APOE, APOE3, and APOE4. RNA-sequence analysis of human cells deficient in HS synthesis demonstrated effects on the transcriptome governing lipid metabolism, autophagy, and mitochondrial biogenesis and showed significant enrichment in AD susceptibility genes identified by GWAS. Neuron-directed knockdown of Psn in Drosophila produced cell loss in the brain and behavioral phenotypes, both suppressed by simultaneous reductions in sfl mRNA levels. Abnormalities in mitochondria, liposome morphology, and autophagosome-derived structures in animals with Psn knockdown were also rescued by simultaneous reduction of sfl. sfl knockdown reversed Psn-dependent transcript changes in genes affecting lipid transport, metabolism, and monocarboxylate carriers. These findings support the direct involvement of HSPGs in AD pathogenesis.

neuroscience↗

bioRxiv2023

Precise single-cell transcriptomic mapping of normal and leukemic cell states reveals unconventional lineage priming in acute myeloid leukemia

In acute myeloid leukemia (AML), genetic mutations distort hematopoietic differentiation, resulting in the accumulation of leukemic blasts. Yet, it remains unclear how these mutations intersect with cellular origins and whether they converge upon similar differentiation patterns. Single-cell RNA sequencing (scRNA-seq) has enabled high-resolution mapping of the relationship between leukemia and normal cell states, yet this application is hampered by imprecise reference maps of normal hematopoiesis and small sample sizes among patient cohorts. As a first step we constructed a reference atlas of human bone marrow hematopoiesis from 263,519 single-cell transcriptomes spanning 55 cellular states, that was benchmarked against independent datasets of immunophenotypically pure hematopoietic stem and progenitor cells. Using this reference atlas, we mapped over 1.2 million single-cell transcriptomes spanning 318 AML, mixed phenotype acute leukemia (MPAL), and acute erythroid leukemia (AEL) samples. This large-scale analysis, together with systematic mapping of genotype-to-phenotype associations between driver mutations and differentiation landscapes, revealed convergence of diverse genetic alterations on twelve recurrent patterns of aberrant differentiation in AML. This included unconventional lymphoid and erythroid priming linked to RUNX1 and TP53 mutations, respectively. We also identified non-genetic determinants of AML differentiation such as two subgroups of KMT2A-rearranged AML that differ in the identity of their leukemic stem cells (LSCs), likely reflecting distinct cellular origins. Furthermore, distinct LSC-driven hierarchies can co-exist within individual patients, providing insights into AML evolution. Together, precise mapping of normal and malignant cell states provides a framework for advancing the study and disease classification of hematologic malignancies thereby informing therapy development.

cancer biology↗

bioRxiv2023

Massively parallel base editing screens to map variant effects on anti-tumor hallmarks of primary human T cells

Base editing enables generation of single nucleotide variants, but large-scale screening in primary human T cells is limited due to low editing efficiency, among other challenges1. Here, we developed a high-throughput approach for high-efficiency and massively parallel adenine and cytosine base-editor screening in primary human T cells. We performed multiple large-scale screens editing 102 genes with central functions in T cells and full-length tiling mutagenesis of selected genes, and read out variant effects on hallmarks of T cell anti-tumor immunity, including activation, proliferation, and cytokine production. We discovered a broad landscape of gain- and loss-of-function mutations, including in PIK3CD and its regulatory subunit encoded by PIK3R1, LCK, AKT1, CTLA-4 and JAK1. We identified variants that affected several (e.g., PIK3CD C416R) or only selected (e.g. LCK Y505C) hallmarks of T cell activity, and functionally validated several hits by probing downstream signaling nodes and testing their impact on T cell polyfunctionality and proliferation. Using primary human T cells in which we engineered a T cell receptor (TCR) specific to a commonly presented tumor testis antigen as a model for cellular immunotherapy, we demonstrate that base edits identified in our screens can tune specific or broad T cell functions and ultimately improve tumor elimination while exerting minimal off-target activity. In summary, we present the first large-scale base editing screen in primary human T cells and provide a framework for scalable and targeted base editing at high efficiency. Coupled with multi-modal phenotypic mapping, we accurately nominate variants that produce a desirable T cell state and leverage these synthetic proteins to improve models of cellular cancer immunotherapies.

immunology↗

bioRxiv2023

ProtPipe: A Multifunctional Data Analysis Pipeline for Proteomics and Peptidomics

Mass spectrometry (MS) is a technique widely employed for the identification and characterization of proteins, personalized medicine, systems biology and biomedical applications. By combining MS with different proteomics approaches such as immunopurification MS, immunopeptidomics, and total protein proteomics, researchers can gain insights into protein-protein interactions, immune responses, cellular processes, and disease mechanisms. The application of MS-based proteomics in these areas continues to advance our understanding of protein function, cellular signaling, and complex biological systems. Data analysis for mass spectrometry is a critical process that includes identifying and quantifying proteins and peptides and exploring biological functions for these proteins in downstream analysis. To address the complexities associated with MS data analysis, we developed ProtPipe to streamline and automate the processing and analysis of high-throughput proteomics and peptidomics datasets. The pipeline facilitates data quality control, sample filtering, and normalization, ensuring robust and reliable downstream analysis. ProtPipe provides downstream analysis including identifying differential abundance proteins and peptides, pathway enrichment analysis, protein-protein interaction analysis, and MHC1-peptide binding affinity. ProtPipe generates annotated tables and diagnostic visualizations from statistical postprocessing and computation of fold-changes across pairwise conditions, predefined in an experimental design. ProtPipe is well-documented open-source software and is available at https://github.com/NIH-CARD/ProtPipe, accompanied by a web interface.

bioinformatics↗

bioRxiv2023

Single cell dissection of developmental origins and transcriptional heterogeneity in B-cell acute lymphoblastic leukemia

Sequencing of bulk tumor populations has improved genetic classification and risk assessment of B-ALL, but does not directly examine intratumor heterogeneity or infer leukemia cellular origins. We profiled 89 B-ALL samples by single-cell RNA-seq (scRNA-seq) and compared them to a reference map of normal human B-cell development established using both functional and molecular assays. Intra-sample heterogeneity was driven by cell cycle, metabolism, differentiation, and inflammation transcriptional programs. By inference of B lineage developmental state composition, nearly all samples possessed a high abundance of pro-B cells, with variation between samples mainly driven by sub-populations. However, ZNF384-r and DUX4- r B-ALL showed composition enrichment of hematopoietic stem cells, BCR::ABL1 and KMT2A-r ALL of Early Lymphoid progenitors, MEF2D-r and TCF3::PBX1 of Pre-B cells. Enrichment of Early Lymphoid progenitors correlated with high-risk clinical features. Understanding variation in transcriptional programs and developmental states of B-ALL by scRNA-seq refines existing clinical and genomic classifications and improves prediction of treatment outcome.

cancer biology↗

bioRxiv2023

Targeting F-actin stress fibers to suppress the dedifferentiated phenotype in chondrocytes

Actin is a central mediator of the chondrocyte phenotype. Monolayer expansion of articular chondrocytes on tissue culture polystyrene, for cell-based repair therapies, leads to chondrocyte dedifferentiation. During dedifferentiation, chondrocytes spread and filamentous (F-)actin reorganizes from a cortical to a stress fiber arrangement causing a reduction in cartilage matrix expression and an increase in fibroblastic matrix and contractile molecule expression. While the downstream mechanisms regulating chondrocyte molecular expression by alterations in F-actin organization have become elucidated, the critical upstream regulators of F-actin networks in chondrocytes are not completely known. Tropomyosin (TPM) and the RhoGTPases are known regulators of F-actin networks. The purpose of this study is to elucidate the regulation of passaged chondrocyte F-actin stress fiber networks and cell phenotype by the specific TPM, TPM3.1, and the RhoGTPase, CDC42. Our results demonstrated that TPM3.1 associates with cortical F-actin and stress fiber F-actin in primary and passaged chondrocytes, respectively. In passaged cells, we found that TPM3.1 inhibition causes F-actin reorganization from stress fibers back to cortical F-actin and also causes an increase in G/F-actin. CDC42 inhibition also causes formation of cortical F-actin. However, CDC42 inhibition, but not TPM3.1 inhibition, leads to the re-association of TPM3.1 with cortical F-actin. Both TPM3.1 and CDC42 inhibition reduces nuclear localization of myocardin related transcription factor, which is known to suppress dedifferentiated molecule expression. We confirmed that TPM3.1 or CDC42 inhibition partially redifferentiates passaged cells by reducing fibroblast matrix and contractile expression, and increasing chondrogenic SOX9 expression. A further understanding on the regulation of F-actin in passaged cells may lead into new insights to stimulate cartilage matrix expression in cells for regenerative therapies.

cell biology↗

bioRxiv2023

Peripheral gating of pain by glial endozepine

We report that diazepam binding inhibitor (DBI) is a glial messenger mediating satellite glia-sensory neuron crosstalk in the dorsal root ganglion (DRG). DBI is highly and specifically expressed in satellite glia cells (SGCs) of mice, rat and human, but not in sensory neurons or other DRG-resident cells. Knockdown of DBI results in a robust mechanical hypersensitivity without significant effects on other sensory modalities. In vivo overexpression of DBI in SGCs reduces sensitivity to mechanical stimulation and alleviates mechanical allodynia in neuropathic and inflammatory pain models. We further show that DBI acts as a partial agonist and positive allosteric modulator at the neuronal GABAA receptors, particularly strongly effecting those with a high-affinity benzodiazepine binding site. Such receptors are selectively expressed by a subpopulation of mechanosensitive DRG neurons and these are also more enwrapped with DBI-expressing glia, as compared to other DRG neurons, suggesting a mechanism for specific effect of DBI on mechanosensation. These findings identified a new, peripheral neuron-glia communication mechanism modulating pain signalling, which can be targeted therapeutically.

neuroscience↗

bioRxiv2023

Antibiotic resistance alters the ability of Pseudomonas aeruginosa to invade the respiratory microbiome

The emergence and spread of antibiotic resistance in bacterial pathogens is a global health threat. One important unanswered question is how antibiotic resistance influences the ability of a pathogen to invade the host-associated microbiome. Here we investigate how antibiotic resistance impacts the ability of the opportunistic bacterial pathogen Pseudomonas aeruginosa to invade the respiratory microbiome, by measuring the ability of P. aeruginosa spontaneous antibiotic resistant mutants to invade pre-established cultures of commensal respiratory microbes. We find that commensal respiratory microbes tend to inhibit the growth of P. aeruginosa, and antibiotic resistance is a double-edged sword that can either help or hinder the ability of P. aeruginosa to overcome this inhibition. The directionality of this help or hinderance depends on both P. aeruginosa genotype and respiratory microbe identity. Antibiotic resistance facilitates the invasion of P. aeruginosa into Staphylococcus lugdunensis, yet impairs invasion into Rothia mucilaginosa and Staphylococcus epidermidis. Streptococcus species provide the strongest inhibition to P. aeruginosa invasion, and this is maintained regardless of antibiotic resistance genotype. Our study demonstrates how antibiotic resistance can alter the ability of a bacterial pathogen to invade the respiratory microbiome and suggests that attempts to manipulate the microbiome should focus on promoting the growth of commensals that can provide robust inhibition of both wildtype and antibiotic resistant pathogen strains.

microbiology↗

bioRxiv2023

Three-dimensional electro-neural interfaces electroplated on subretinal prostheses

ObjectiveHigh-resolution retinal prosthetics offer partial restoration of sight to patients blinded by retinal degenerative diseases through electrical stimulation of the remaining neurons. Decreasing the pixel size enables an increase in prosthetic visual acuity, as demonstrated in animal models of retinal degeneration. However, scaling down the size of planar pixels is limited by the reduced penetration depth of the electric field in tissue. We investigate 3-dimensional structures on top of the photovoltaic arrays for enhanced penetration of electric field to permit higher-resolution implants. ApproachWe developed 3D COMSOL models of subretinal photovoltaic arrays that accurately quantify the device electrodynamics during stimulation and verified it experimentally through comparison with the standard (flat) photovoltaic arrays. The models were then applied to optimise the design of 3D electrode structures (pillars and honeycombs) to efficiently stimulate the inner retinal neurons. The return electrodes elevated on top of the honeycomb walls surrounding each pixel orient the electric field inside the cavities vertically, aligning it with bipolar cells for optimal stimulation. Alternatively, pillars elevate the active electrode into the inner nuclear layer, improving proximity to the target neurons. Modelling results informed a microfabrication process of electroplating the 3D electrode structures on top of the existing flat subretinal prosthesis. Main resultsSimulations demonstrate that despite the conductive sidewalls of the 3D electrodes being exposed to electrolyte, most of the charge flows via the high-capacitance sputtered Iridium Oxide film that caps the top of the 3D structures. The 24 {micro}m height of the electroplated honeycomb structures was optimised for integration with the inner nuclear layer cells in rat retina, while 35 {micro}m height of the pillars was optimized for penetrating the debris layer in human patients. Release from the wafer and implantation of the 3D arrays demonstrated that they are mechanically robust to withstand the associated forces. Histology demonstrated successful integration of the 3D structures with the rat retina in-vivo. SignificanceElectroplated 3D honeycomb structures produce a vertically oriented electric field that offers low stimulation threshold, high spatial resolution and high contrast for the retinal implants with pixel sizes down to 20{micro}m in width. Pillar electrodes offer an alternative configuration for extending the stimulation past the debris layers. Electroplating of the 3D structures is compatible with the fabrication process of the flat photovoltaic arrays, thereby enabling much more efficient stimulation than in their original flat configuration.

neuroscience↗

bioRxiv2023

Selective advantage of mutant stem cells in clonal hematopoiesis occurs by attenuating the deleterious effects of inflammation and aging

Clonal hematopoiesis (CH) arises when hematopoietic stem cells (HSC) acquire mutations in genes, including DNMT3A and TET2, conferring a competitive advantage through a mechanism that remains unclear. To gain insight into how CH mutations enable gradual clonal expansion, we used single-cell multi-omics with high-fidelity genotyping on CH bone marrow samples. Most of the selective advantage of mutant cells occurs within HSCs. DNMT3A and TET2-mutant clones expand further in early progenitors, while TET2 mutations accelerate myeloid maturation in a dose-dependent manner. Unexpectedly, both mutant and non-mutant HSCs from CH samples are enriched for inflammatory and aging transcriptomic signatures, compared to HSC from non-CH samples, revealing a non-cell autonomous mechanism. However, DNMT3A and TET2-mutant HSCs have an attenuated inflammatory response relative to wild-type HSCs within the same sample. Our data support a model whereby CH clones are gradually selected because they are more resistant to the deleterious impact of inflammation and aging.

cancer biology↗

bioRxiv2023

A hematopoietic stem cell subset that retains memory of prior inflammatory stress accumulates in aging and clonal hematopoiesis

Inflammation activates many blood cell types, driving aging and malignancy. Yet, hematopoietic stem cells (HSCs) survive a lifetime of infection to sustain life-long blood production. To understand HSC adaptation to inflammation, we developed xenograft inflammation-recovery models and performed single cell multiomics on isolated human HSC. Two transcriptionally and epigenetically distinct HSC subsets expressing canonical HSC programs were identified. Only one showed sustained transcriptional and epigenetic changes after recovery from inflammatory treatments. This HSC inflammatory memory (HSC-iM) program is enriched in memory T cells and HSCs from recovered COVID-19 patients. Importantly, HSC-iM accumulates with age and with clonal hematopoiesis. Overall, heritable molecular alterations in a subset of human HSCs, an adaptation to long-term inflammatory stress, may predispose to heightened age-related risk of blood cancer and infection. One-Sentence SummaryInflammation across a lifetime rewires human HSCs to produce a distinct HSC subset with both beneficial and deleterious fitness consequences.

cell biology↗