Anatomy and Neuroscience - Research Publications

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Author: Blewitt, Marnie × Start date: 2020 × End date: 2020 ×

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    RNF41 regulates the damage recognition receptor Clec9A and antigen cross-presentation in mouse dendritic cells
    Tullett, KM ; Tan, PS ; Park, H-Y ; Schittenhelm, RB ; Michael, N ; Li, R ; Policheni, AN ; Gruber, E ; Huang, C ; Fulcher, AJ ; Danne, JC ; Czabotar, PE ; Wakim, LM ; Mintern, JD ; Ramm, G ; Radford, KJ ; Caminschi, I ; O'Keeffe, M ; Villadangos, JA ; Wright, MD ; Blewitt, ME ; Heath, WR ; Shortman, K ; Purcell, AW ; Nicola, NA ; Zhang, J-G ; Lahoud, MH (ELIFE SCIENCES PUBLICATIONS LTD, 2020-12-02)
    The dendritic cell receptor Clec9A facilitates processing of dead cell-derived antigens for cross-presentation and the induction of effective CD8+ T cell immune responses. Here, we show that this process is regulated by E3 ubiquitin ligase RNF41 and define a new ubiquitin-mediated mechanism for regulation of Clec9A, reflecting the unique properties of Clec9A as a receptor specialized for delivery of antigens for cross-presentation. We reveal RNF41 is a negative regulator of Clec9A and the cross-presentation of dead cell-derived antigens by mouse dendritic cells. Intriguingly, RNF41 regulates the downstream fate of Clec9A by directly binding and ubiquitinating the extracellular domains of Clec9A. At steady-state, RNF41 ubiquitination of Clec9A facilitates interactions with ER-associated proteins and degradation machinery to control Clec9A levels. However, Clec9A interactions are altered following dead cell uptake to favor antigen presentation. These findings provide important insights into antigen cross-presentation and have implications for development of approaches to modulate immune responses.
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    HBO1 is required for the maintenance of leukaemia stem cells
    MacPherson, L ; Anokye, J ; Yeung, MM ; Lam, EYN ; Chan, Y-C ; Weng, C-F ; Yeh, P ; Knezevic, K ; Butler, MS ; Hoegl, A ; Chan, K-L ; Burr, ML ; Gearing, LJ ; Willson, T ; Liu, J ; Choi, J ; Yang, Y ; Bilardi, RA ; Falk, H ; Nghi, N ; Stupple, PA ; Peat, TS ; Zhang, M ; de Silva, M ; Carrasco-Pozo, C ; Avery, VM ; Khoo, PS ; Dolezal, O ; Dennis, ML ; Nuttall, S ; Surjadi, R ; Newman, J ; Ren, B ; Leaver, DJ ; Sun, Y ; Baell, JB ; Dovey, O ; Vassiliou, GS ; Grebien, F ; Dawson, S-J ; Street, IP ; Monahan, BJ ; Burns, CJ ; Choudhary, C ; Blewitt, ME ; Voss, AK ; Thomas, T ; Dawson, MA (NATURE PORTFOLIO, 2020-01-09)
    Acute myeloid leukaemia (AML) is a heterogeneous disease characterized by transcriptional dysregulation that results in a block in differentiation and increased malignant self-renewal. Various epigenetic therapies aimed at reversing these hallmarks of AML have progressed into clinical trials, but most show only modest efficacy owing to an inability to effectively eradicate leukaemia stem cells (LSCs)1. Here, to specifically identify novel dependencies in LSCs, we screened a bespoke library of small hairpin RNAs that target chromatin regulators in a unique ex vivo mouse model of LSCs. We identify the MYST acetyltransferase HBO1 (also known as KAT7 or MYST2) and several known members of the HBO1 protein complex as critical regulators of LSC maintenance. Using CRISPR domain screening and quantitative mass spectrometry, we identified the histone acetyltransferase domain of HBO1 as being essential in the acetylation of histone H3 at K14. H3 acetylated at K14 (H3K14ac) facilitates the processivity of RNA polymerase II to maintain the high expression of key genes (including Hoxa9 and Hoxa10) that help to sustain the functional properties of LSCs. To leverage this dependency therapeutically, we developed a highly potent small-molecule inhibitor of HBO1 and demonstrate its mode of activity as a competitive analogue of acetyl-CoA. Inhibition of HBO1 phenocopied our genetic data and showed efficacy in a broad range of human cell lines and primary AML cells from patients. These biological, structural and chemical insights into a therapeutic target in AML will enable the clinical translation of these findings.
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    Unique properties of a subset of human pluripotent stem cells with high capacity for self-renewal
    Lau, KX ; Mason, EA ; Kie, J ; De Souza, DP ; Kloehn, J ; Tull, D ; McConville, MJ ; Keniry, A ; Beck, T ; Blewitt, ME ; Ritchie, ME ; Naik, SH ; Zalcenstein, D ; Korn, O ; Su, S ; Romero, IG ; Spruce, C ; Baker, CL ; McGarr, TC ; Wells, CA ; Pera, MF (Nature Research, 2020-05-15)
    Archetypal human pluripotent stem cells (hPSC) are widely considered to be equivalent in developmental status to mouse epiblast stem cells, which correspond to pluripotent cells at a late post-implantation stage of embryogenesis. Heterogeneity within hPSC cultures complicates this interspecies comparison. Here we show that a subpopulation of archetypal hPSC enriched for high self-renewal capacity (ESR) has distinct properties relative to the bulk of the population, including a cell cycle with a very low G1 fraction and a metabolomic profile that reflects a combination of oxidative phosphorylation and glycolysis. ESR cells are pluripotent and capable of differentiation into primordial germ cell-like cells. Global DNA methylation levels in the ESR subpopulation are lower than those in mouse epiblast stem cells. Chromatin accessibility analysis revealed a unique set of open chromatin sites in ESR cells. RNA-seq at the subpopulation and single cell levels shows that, unlike mouse epiblast stem cells, the ESR subset of hPSC displays no lineage priming, and that it can be clearly distinguished from gastrulating and extraembryonic cell populations in the primate embryo. ESR hPSC correspond to an earlier stage of post-implantation development than mouse epiblast stem cells.