Unknown,Transcriptomics,Genomics,Proteomics

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GSK-3Î² inhibition promotes engraftment of ex vivo expanded hematopoietic stem cells

ABSTRACT: Glycogen synthase kinase-3Î² (GSK-3Î²) has been recently identified as an important regulator of stem cell function. In vitro studies show that GSK-3Î² inhibition delays proliferation of human haematopoietic progenitor cells while increasing numbers of late dividing multipotent progenitors. Gene expression analysis revealed that GSK-3Î² inhibition modulates the expression of a subset of genes that are transcriptional targets for cytokines. GSK-3Î² inhibition antagonised down-regulation of genes encoding cyclin dependent kinase inhibitor p57 and a member of the growth arrest and DNA damage 45 family, GADD45B as well as up-regulation of cyclin D1 by cytokines, providing a possible mechanism for the BIO-induced delay in cell cycle progression. Surprisingly, inhibition of GSK-3Î² earlier shown to prevent Î²-catenin degradation and promote the nuclear accumulation of Î²-catenin was not sufficient to activate its transcriptional targets in haematopoietic stem cells. GSK-3Î² inhibition up-regulated the expression of a several positive regulators of stem cell function suppressed during cytokine-induced proliferation. The data supports a clinical role for GSK-3Î² inhibition to improve engraftment efficiency of ex vivo expanded stem cells. Total RNA was isolated from three groups following expansion of CD34+ cells in cytokikes and then treatment with BIO, as described below.

ORGANISM(S): Homo sapiens

SUBMITTER: Kap-hyoun Ko

PROVIDER: E-GEOD-21073 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:GSK-3b plays a critical role in regulating the Wnt/b-catenin signaling pathway, and manipulating GSK-3b in dendritic cells (DCs) has been shown to improve anti-tumor efficacy of DC vaccines. Since inhibition of GSK-3b leads to activation of b-catenin, we hypothesize that blocking GSK-3b in DCs negatively regulates DC-mediated CD8 T cell immunity and anti-tumor immunity. Using CD11c-GSK-3b-/- conditional knockout mice in which GSK-3b is genetically deleted in CD11c-expressing DCs, we surprisingly found that deletion of GSK-3b in DCs resulted in increased anti-tumor immunity, which contradicted our initial expectation of reduced anti-tumor immunity due to presumed upregulation of b-catenin in DCs. Indeed, we found by both western blot and flow cytometry that deletion of GSK-3b in DCs did not lead to augmented expression of b-catenin protein, suggesting that GSK-3b exerts its function independent of b-catenin. Supporting this notion, our single cell RNA sequencing (scRNA-seq) analysis revealed that GSK-3b-deficient DCs exhibited distinct gene expression patterns with minimally overlapping differentially expressed genes (DEGs) compared to DCs with activated b-catenin. This suggests that deletion of GSK-3b in DCs is unlikely to lead to upregulation of b-catenin at the transcriptional level. Consistent with enhanced anti-tumor immunity, we also found that CD11c-GSK-3b-/- mice exhibited significantly augmented cross-priming of antigen-specific CD8 T cells following DC-targeted vaccines. We further found that deletion of GSK-3b in DCs completely abrogated memory CD8 T cell responses, suggesting that GSK-3b in DCs also plays a negative role in regulating the differentiation and/or maintenance of memory CD8 T cells. scRNA-seq analysis further revealed that although deletion of GSK-3b in DCs positively regulated transcriptional programs for effector differentiation and function of primed antigen-specific CD8 T cells in CD11c-GSK-3b-/- mice during the priming phase, it resulted in significantly reduced antigen-specific memory CD8 T cells, consistent with diminished memory responses. Taken together, our data demonstrate that GSK-3b in DCs has opposite functions in regulating cross-priming and memory CD8 T cell responses, and GSK-3b exerts its functions independent of its regulation of b-catenin.

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GSK-3Î² inhibition promotes engraftment of ex vivo expanded hematopoietic stem cells

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