Regulation of embryonic diapause, dormancy that interrupts the tight connection between developmental stage and time, is still poorly understood. Here, we characterize the transcriptional and metabolite profiles of mouse diapause embryos and identify unique gene expression and metabolic signatures with activated lipolysis, glycolysis, and metabolic pathways regulated by AMPK. Lipolysis is increased due to mTORC2 repression, increasing fatty acids to support cell survival. We further show that starvation in pre-implantation ICM-derived mouse ESCs induces a reversible dormant state, transcriptionally mimicking the in vivo diapause stage. During starvation, Lkb1, an upstream kinase of AMPK, represses mTOR, which induces a reversible glycolytic and epigenetically H4K16Ac-negative, diapause-like state. Diapause furthermore activates expression of glutamine transporters SLC38A1/2. We show by genetic and small molecule inhibitors that glutamine transporters are essential for the H4K16Ac-negative, diapause state. These data suggest that mTORC1/2 inhibition, regulated by amino acid levels, is causal for diapause metabolism and epigenetic state. In BriefHussein et al. report that, during starvation, mTOR is repressed through LKB1-AMPK, inducing a reversible metabolically active but epigenetically silenced embryonic diapause-like state that upregulates expression of the glutamine transporters SLC38A1/2. These transporters are required for the H4K16ac-negative, diapause state.

A major obstacle to stem-cell gene therapy rests in the inability to deliver a gene into a therapeutically relevant fraction of stem cells. One way to circumvent this obstacle is to use selection. Vectors containing two linked genes serve as the basis for selection, with one gene encoding a selectable product and the other, a therapeutic protein. Applying selection in vivo has the potential to bring a minor population of genetically corrected cells into the therapeutic range. But strategies for achieving in vivo selection have traditionally relied on genes that confer resistance to cytotoxic drugs and are encumbered by toxicity. Here we describe a new system for in vivo selection that uses a 'cell-growth switch', allowing a minor population of genetically corrected cells into the therapeutic range. But strategies for achieving in vivo selection have traditionally relied on genes that confer resistance to cytotoxic drugs and are encumbered by toxicity. Here we describe a new system for in vivo selection that uses a 'cell-growth switch', allowing a minor population of genetically modified cells to be inducibly amplified, thereby averting the risks associated with cytotoxic drugs. This system provides a general platform for conditionally expanding genetically modified cell populations in vivo, and may have widespread applications in gene and cell therapy.

No abstract

Receptor dimerization is the key signaling event for many cytokines, including erythropoietin. A system has been recently developed that permits intracellular protein dimerization to be reversibly activated in response to a lipid-soluble dimeric form of the drug FK506, called FK1012. FK1012 is used as a pharmacological mediator of dimerization to bring together FK506 binding domains, taken from the endogenous protein FKBP12. In experiments reported herein, FK1012-induced dimerization of a fusion protein containing the intracellular portion of the erythropoietin receptor allowed cells normally dependent on interleukin 3 to proliferate in its absence. FK506 competitively reversed the proliferative effect of FK1012 but had no inf luence on the proliferative effect of interleukin 3. Signaling pathways activated by FK1012 mimicked those activated by erythropoietin, because both JAK2 and STAT5 were phosphorylated in response to FK1012. This approach may provide a means to specifically and reversibly stimulate the proliferation of genetically modified cell populations in vitro or in vivo.