Biology subjects

Kent, D. G.

Publications and source records attributed to Kent, D. G..

6 recordsLinked to original sources

bioRxiv2026

A scalable in vitro cell competition model identifies preleukemic super competitor clones

Preleukemic and leukemic conditions are frequently characterized by the acquisition of recurrent genetic mutations in hematopoietic stem cells (HSCs). One of the most frequently mutated genes in both clonal hematopoiesis and myeloid malignancies is the ten-eleven translocation 2 (TET2) gene, which encodes an epigenetic enzyme with a critical role in the regulation of gene transcription. While substantial work has shown that TET2 loss-of-function mutations lead to a clonal advantage, the underpinning cellular mechanism remains unclear, especially with respect to the early interactions with other cells. To study these early cell interactions of mutant and non-mutant cells, we developed an in vitro clonal competition platform where single-cell derived cultures are split into subclonal fractions which can be simultaneously cross-competed against multiple different clones of an opposing genotype. We identify a novel preleukemic cell phenotype that parallels the developmental biology phenomenon of super-competition, whereby ~10% of TET2-mutant clones actively and selectively suppress wild type (WT) cell expansion and, in some cases, completely eliminate WT clones. Molecular profiling and functional assays point to RAC1-GEF signalling inhibition as a specific WT cell vulnerability that is promoted, and tolerated, by neighbouring TET2-mutated clones. In addition to pinpointing selective drivers of non-mutant cell suppression, this clonal competition platform offers a broadly applicable new approach to study cell competition dynamics at an unprecedented scale.

cancer biology↗

bioRxiv2026

Sex-specific lymphoid lineage differentiation is modulated by CIZ1

The epigenetic stability factor CIZ1 helps maintain X-chromosome inactivation, and its absence results in murine female-specific splenomegaly. By exploring the aetiology of this pathology, we reveal unexpected evidence for dosage compensation via modification of X-linked gene expression in males. Genes known to escape repression on the inactive X-chromosome in females, including the B-cell maturation factor DDX3X, require CIZ1 in males to maintain parity between the sexes. Furthermore, absence of this single regulator triggers stark sex-specific changes to large autosomal domains in B cells, causing naive female B cells to shift prematurely towards germinal centre transcriptional signatures, including immunoglobulin and pro-proliferation genes even without immune challenge. Conversely, males acquire natural killer-like gene expression through elevation of Killer cell Lectin-like Receptors. Together, the data indicate that CIZ1 limits sexually dimorphic gene expression on autosomes, and promotes dosage compensation by modulation of the male X-chromosome, introducing a new paradigm for sex biased disorders of the immune system.

cell biology↗

bioRxiv2025

Non-canonical inflammasome drives intrinsic anti-microbial responses in human Natural Killer (NK) cells.

Inflammasome pathways are critical for detecting pathogens and initiating inflammatory responses, yet how individual cell types configure these signalling platforms remains poorly understood. Here, we demonstrate that human Natural Killer cells possess distinct inflammasome machinery, lacking canonical components and innate sensors (NLRP3, NLRC4, TLR4) but harbour a functional non-canonical inflammasome (NCI). Using Salmonella enterica Typhimurium infection of primary human NK cells, we show that cytosolic bacterial access triggers caspase-4-dependent pyroptosis and IL-18 secretion, without canonical inflammasome activation or IL-1{beta} production. Strikingly, murine NK cells express inflammasome transcripts but lack functional caspase-11 protein and do not undergo pyroptosis upon infection, revealing species-specific post-transcriptional regulation. We further identify IL-12, but not IFN{gamma}, as a priming signal for NCI components in human NK cells, a functional distinction from macrophages and epithelial cells. Together, these findings reveal that human NK cells have evolved a specialised inflammasome with unique regulatory inputs, establishing these cells as autonomous sensors of intracellular Gram-negative infection.

immunology↗

bioRxiv2025

Selection of pre-leukemic hematopoietic stem cells driven by distinct extracellular matrix molecules

Despite rapid advances in mapping genetic drivers and gene expression changes in hematopoietic stem cells (HSCs), there is a relative paucity of studies at the protein level. Here, we perform a deep, multi-omic characterization (epigenome, transcriptome and proteome) of HSCs carrying a loss-of-function mutation in Tet2, a key driver of increased self-renewal in blood cancers. Using state-of-the-art, multiplexed, low-input mass spectrometry (MS)-based proteomics, we profile wildtype (WT) and TET2-deficient (Tet2-/-) HSCs and show that the proteome captures previously unrecognized molecular processes which define the pre-leukemic HSC molecular landscape. Specifically, we obtain more accurate stratification of WT and Tet2-/- HSCs than transcriptomic approaches and identify extracellular matrix (ECM) molecules as novel points of dysregulation upon TET2 loss. HSC expansion assays using ECM-functionalized hydrogels confirm a selective effect on the expansion of Tet2-mutant HSCs. Taken together, our study represents a comprehensive molecular characterization of Tet2-mutant HSCs and identifies a previously unanticipated role of ECM molecules in regulating self-renewal of disease-driving HSCs.

cell biology↗

bioRxiv2021

Prospective isolation of mouse and human hematopoietic stem cells using Plexin domain containing 2

Numerous strategies exist to isolate hematopoietic stem cells (HSCs) using complex combinations of markers and flow cytometry. However, robust identification of HSCs using imaging techniques is substantially more challenging which has prompted the recent development of HSC reporter mice. To date, none of the molecules used in these reporters have been useful for human HSC identification. Here we report that PLXDC2 is a useful marker for both mouse and human HSCs. Using a green fluorescent protein (GFP) knock-in at the Plxdc2 locus in mice (hereafter denoted as Plxdc2-GFP), we showed that Plxdc2-GFP is highly expressed in HSCs with 1 in 2.8 Plxdc2-GFP+CD150+ cells giving long-term multi-lineage reconstitution in transplantation. Moreover, we developed a novel human PLXDC2 antibody and showed that human PLXDC2+ HSCs have stronger long-term multilineage reconstitution ability compared with PLXDC2- HSCs in a xenograft model. Thus, our study identifies PLXDC2 as a highly relevant molecule in HSC identification, potentially allowing greater purity and live in vivo tracking of these cells. SummaryTo date, few molecules are available for isolation of HSCs across species. The present study shows that PLXDC2 is a highly useful molecule for isolation of HSCs, which works across mouse and human.

cell biology↗

bioRxiv2021

A highly efficient reporter system for identifying and characterizing in vitro expanded hematopoietic stem cells

Hematopoietic stem cells (HSCs) cultured outside the body are the fundamental component of a wide range of cellular and gene therapies. Recent efforts have achieved more than 200-fold expansion of functional HSCs, but their molecular characterization has not been possible due to the substantial majority of cells being non-HSCs and single cell-initiated cultures displaying substantial clone-to-clone variability. Using the Fgd5 reporter mouse in combination with the EPCR surface marker, we report exclusive identification of HSCs from non-HSCs in expansion cultures. Linking single clone functional transplantation data with single clone gene expression profiling, we show that the molecular profile of expanded HSCs is similar to actively cycling fetal liver HSCs and shares a gene expression signature with functional HSCs from all sources, including Prdm16, Fstl1 and Palld. This new tool can now be applied to a wide-range of functional screening and molecular experiments previously not possible due to limited HSC numbers.

cell biology↗