The serine/threonine kinase Akt has been implicated in the control of cell survival and metabolism. Here we report the disruption of the most ubiquitously expressed member of the akt family of genes, akt1, in the mouse.

To elucidate the functions of the serine/threonine kinase Akt/PKB in vivo, we generated mice lacking both akt1 and akt2 genes. Akt1/Akt2 double-knockout (DKO) mice exhibit severe growth deficiency and die shortly after birth. These mice display impaired skin development because of a proliferation defect, severe skeletal muscle atrophy because of a marked decrease in individual muscle cell size, and impaired bone development. These defects are strikingly similar to the phenotypes of IGF-1 receptor-deficient mice and suggest that Akt may serve as the most critical downstream effector of the IGF-1 receptor during development. In addition, Akt1/Akt2 DKO mice display impeded adipogenesis. Specifically, Akt1 and Akt2 are required for the induced expression of PPAR␥, the master regulator of adipogenesis, establishing a new essential role for Akt in adipocyte differentiation. Overall, the combined deletion of Akt1 and Akt2 establishes in vivo roles for Akt in cell proliferation, growth, and differentiation. These functions of Akt were uncovered despite the observed lower level of Akt activity mediated by Akt3 in Akt1/Akt2 DKO cells, suggesting that a critical threshold level of Akt activity is required to maintain normal cell proliferation, growth, and differentiation.

Summary Akt-deficiency causes resistance to replicative senescence, oxidative stress- or oncogenic Ras-induced premature senescence, and to reactive oxygen species (ROS)-mediated apoptosis. Akt activation induces premature senescence and sensitizes cells to ROS-mediated apoptosis by increasing intracellular ROS through increased oxygen consumption and by inhibiting the expression of ROS-scavengers downstream of FoxO, particularly sestrin3 expression. This uncovers an Achilles’ heel of Akt, since in contrast to its ability to inhibit apoptosis induced by multiple apoptotic stimuli; Akt could not inhibit ROS-mediated apoptosis. Furthermore, treatment with rapamycin that led to further Akt activation and resistance to etoposide, hypersensitized cancer cells to ROS-mediated apoptosis. Given that rapamycin alone is mainly cytostatic, this constitutes a strategy for cancer therapy that selectively eradicates cancer cells via Akt activation.

Cancer patients can harbor significant numbers of CD8 and CD4 T cells with specificities to tumor antigens (Ags). Yet, in most cases, such T cells fail to eradicate the tumor in vivo. Here, we investigated the interference of Ag-specific CD4 ؉ CD25 ؉ regulatory T cells (Treg) with the tumor-specific CD8 T cell immune response in vivo, by monitoring the homing, expansion, and effector function of both subsets in draining and nondraining lymph nodes. The results show that CD8 cells expand to the same extent and produce similar levels of IFN-␥ in the presence or absence of Ag-specific Treg. Nevertheless, these Treg abrogate CD8 T cell-mediated tumor rejection by specifically suppressing the cytotoxicity of expanded CD8 cells. The molecular mechanism of suppression involves TGF-␤ because expression of a dominant-negative TGF-␤ receptor by tumor-specific CD8 cells renders them resistant to suppression and is associated with tumor rejection and unimpaired cytotoxicity. (5-7). Treg also can control the magnitude of recall CD8 T cell responses in different settings that include viral (8, 9) and bacterial (10) infections as well as allograft transplantation in vivo (11,12). A role in Treg function has been attributed to IL-2, which stimulates Treg and in turn may inhibit division of memory CD8 T cells (13,14). However, these studies did not trace Ag-specific Treg in draining lymph nodes (LN), and it is not clear to what extent the different outcomes reflect accumulation of Treg by specific homing and local expansion. In fact, most studies were conducted with polyclonal Treg of unknown specificity where such questions cannot be addressed, and, therefore, the mode of inhibition cannot be correlated with specific Treg accumulation. Analysis of tumor-bearing patients suggests that suppression of CD8 T cell cytotoxicity by Treg may be causally related to tumor progression, because tumor-specific CD8 cells and tumor-specific CD4 Treg frequently accumulate in tumors from melanoma patients, and tumor-specific CD8 cells fail to exert cytotoxic T lymphocyte effector function (15)(16)(17).This study investigates how Treg suppress primary CD8 T cell immune responses directed against tumor cells expressing influenza hemagglutinin (HA) as a surrogate tumor-specific Ag. Naïve CD8 and regulatory CD4 T cells with transgenic receptors specific for distinct peptides of HA were used to allow us to follow the fate of these cells in tumor draining LN. The results show that Treg interfere with CD8 T cell-mediated tumor rejection relatively early during the immune response, and the mechanism by which Treg suppress naïve CD8 cells differs from the ones that have been reported for memory CD8 cells (10)(11)(12)14). Treg influenced neither the kinetics of proliferation nor the commitment of recently activated CD8 cells to produce inflammatory cytokines. Nevertheless, CD8 cells failed to undergo normal functional maturation in the presence of Treg as evidenced by the fact that their cytotoxic potential to destroy specific targets in vivo was abolis...