Despite two decades of studies documenting thein vitroblood-forming potential of murine embryonic stem cells (ESCs), achieving stable long-term blood engraftment of ESC-derived hematopoietic stem cells in irradiated mice has proven difficult. We have exploited theCdx-Hoxpathway, a genetic program important for blood development, to enhance the differentiation of ESCs along the hematopoietic lineage. Using an embryonic stem cell line engineered with tetracycline-inducibleCdx4, we demonstrate that ectopicCdx4expression promotes hematopoietic mesoderm specification, increases hematopoietic progenitor formation, and, together withHoxB4, enhances multilineage hematopoietic engraftment of lethally irradiated adult mice. Clonal analysis of retroviral integration sites confirms a common stem cell origin of lymphoid and myeloid populations in engrafted primary and secondary mice. These data document the cardinal stem cell features of self-renewal and multilineage differentiation of ESC-derived hematopoietic stem cells.

The SWI/SNF complex plays an important role in mouse embryonic stem cells (mESCs), but it remains to be determined whether this complex is required for the pluripotency of human ESCs (hESCs). Using RNAi, we demonstrated that depletion of BRG1, the catalytic subunit of the SWI/SNF complex, led to impaired self-renewing ability and dysregulated lineage specification of hESCs. A unique composition of the BRG1-SWI/SNF complex in hESCs was further defined by the presence of BRG1, BAF250A, BAF170, BAF155, BAF53A, and BAF47. Genome-wide expression analyses revealed that BRG1 participated in a broad range of biological processes in hESCs through pathways different from those in mESCs. In addition, chromatin immunoprecipitation sequencing (ChIP-seq) demonstrated that BRG1 played a repressive role in transcriptional regulation by modulating the acetylation levels of H3K27 at the enhancers of lineage-specific genes. Our data thus provide valuable insights into molecular mechanisms by which transcriptional repression affects the self-renewal and differentiation of hESCs.

Metabolic fitness of T cells is essential for their vitality, which is largely dependent on the behavior of the mitochondria. The nature of mitochondrial behavior in tumor-infiltrating T cells remains poorly understood. In this study, we show that mitofusin-2 (MFN2) expression is positively correlated with the prognosis of multiple cancers. Genetic ablation of Mfn2 in CD8 ⁺ T cells dampens mitochondrial metabolism and function and promotes tumor progression. In tumor-infiltrating CD8 ⁺ T cells, MFN2 enhances mitochondria–endoplasmic reticulum (ER) contact by interacting with ER-embedded Ca ²⁺ -ATPase SERCA2, facilitating the mitochondrial Ca ²⁺ influx required for efficient mitochondrial metabolism. MFN2 stimulates the ER Ca ²⁺ retrieval activity of SERCA2, thereby preventing excessive mitochondrial Ca ²⁺ accumulation and apoptosis. Elevating mitochondria-ER contact by increasing MFN2 in CD8 ⁺ T cells improves the efficacy of cancer immunotherapy. Thus, we reveal a tethering-and-buffering mechanism of organelle cross-talk that regulates the metabolic fitness of tumor-infiltrating CD8 ⁺ T cells and highlights the therapeutic potential of enhancing MFN2 expression to optimize T cell function.

Cdx genes ( Cdx1 , Cdx2 , and Cdx4 ) encode a family of caudal -related transcription factors that mediate anterior–posterior patterning during embryogenesis through Hox gene regulation. Homologues in the zebrafish have been shown to play key roles in blood formation. To define the role of Cdx genes during embryonic hematopoiesis in a mammalian system, we examined the hematopoietic potential of Cdx -deficient mouse embryonic stem cells (ESCs) in vitro and in vivo . Individual Cdx -deficient ESCs exhibited impaired embryonic hematopoietic progenitor formation and altered Hox gene expression, most notably for Cdx2 deficiency. A more severe hematopoietic defect was observed with compound Cdx deficiency than loss of function of any single Cdx gene. Reduced hematopoietic progenitor formation of ESCs deficient in multiple Cdx genes could be rescued by ectopic expression of Cdx4 , concomitant with partially restored Hox gene expression. These results reveal an essential and partially redundant role for multiple Cdx genes during embryonic hematopoiesis in the mouse.