The differentiation of B cells into immunoglobulin-secreting plasma cells is controlled by two transcription factors, Blimp-1 and XBP1. By gene expression profiling, we defined a set of genes whose induction during mouse plasmacytic differentiation is dependent on Blimp-1 and/or XBP1. Blimp-1-deficient B cells failed to upregulate most plasma cell-specific genes, including xbp1. Differentiating xbp1-deficient B cells induced Blimp-1 normally but failed to upregulate genes encoding many secretory pathway components. Conversely, ectopic expression of XBP1 induced a wide spectrum of secretory pathway genes and physically expanded the endoplasmic reticulum. In addition, XBP1 increased cell size, lysosome content, mitochondrial mass and function, ribosome numbers, and total protein synthesis. Thus, XBP1 coordinates diverse changes in cellular structure and function resulting in the characteristic phenotype of professional secretory cells.

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogen and nutrient signals to control cell proliferation and cell size. Hence, mTORC1 is implicated in a large number of human diseases--including diabetes, obesity, heart disease, and cancer--that are characterized by aberrant cell growth and proliferation. Although eukaryotic translation initiation factor 4E-binding proteins (4E-BPs) are critical mediators of mTORC1 function, their precise contribution to mTORC1 signaling and the mechanisms by which they mediate mTORC1 function have remained unclear. We inhibited the mTORC1 pathway in cells lacking 4E-BPs and analyzed the effects on cell size, cell proliferation, and cell cycle progression. Although the 4E-BPs had no effect on cell size, they inhibited cell proliferation by selectively inhibiting the translation of messenger RNAs that encode proliferation-promoting proteins and proteins involved in cell cycle progression. Thus, control of cell size and cell cycle progression appear to be independent in mammalian cells, whereas in lower eukaryotes, 4E-BPs influence both cell growth and proliferation.The mammalian target of rapamycin complex 1 (mTORC1) controls growth (increase in cell mass) and proliferation (increase in cell number) by modulating mRNA translation through phosphorylation of the eukaryotic translation initiation factor 4E (eIF4E)-binding proteins (4E-BP1, 2, and 3) and the ribosomal protein S6 kinases (S6K1 and 2) (1,2). 4E-BPs regulate the translation of a subset of mRNAs by competing with eIF4G for binding to eIF4E, thus preventing the assembly of the eIF4F complex, whereas the S6Ks control the phosphorylation status of a number of translational components (1-3). Rapamycin has been an important tool in understanding mTORC1 signaling; however, it inefficiently and transiently inhibits 4E-BP phosphorylation (4)( fig. S1A). Moreover, we found that rapamycin inhibited proliferation and G1/S cell cycle progression of WT and 4E-BP double knock-out (DKO) mouse embryonic fibroblasts (MEFs) to the same extent, which suggests that its effects are not mediated by 4E-BPs ( fig. S1, B to D). To directly address the role of 4E-BPs in mTORC1 signaling, we depleted raptor, a component of mTORC1 required for substrate binding (5), in these MEFs. 4E-BP DKO MEFs lack all three 4E-BPs as they do not express 4E-BP3 ( fig. S2A) 1A). Wild-type MEFs in which raptor was depleted proliferated more slowly than control cells, whereas raptor-depleted 4E-BP DKO MEFs proliferated at a rate indistinguishable from that of control cells (Fig. 1B). Similarly, in human embryonic kidney (HEK) 293T cells, raptor silencing had a pronounced effect on mTORC1 signaling and proliferation (Figs. 1C and D). The effect of raptor silencing on proliferation, but not mTOR signaling, was attenuated by codepletion of 4E-BPs (Fig. 1D). Thus, mTORC1-dependent proliferation requires 4E-BPs.To further assess the role of 4E-BPs in mTORC1-mediated cell proliferation, we depleted mTOR or rictor (an mTORC2 specific component), i...

Translational regulation plays a critical role in the control of cell growth and proliferation. A key player in translational control is eIF4E, the mRNA 5′ cap-binding protein. Aberrant expression of eIF4E promotes tumorigenesis and has been implicated in cancer development and progression. The activity of eIF4E is dysregulated in cancer. Regulation of eIF4E is partly achieved through phosphorylation. However, the physiological significance of eIF4E phosphorylation in mammals is not clear. Here, we show that knock-in mice expressing a nonphosphorylatable form of eIF4E are resistant to tumorigenesis in a prostate cancer model. By using a genome-wide analysis of translated mRNAs, we show that the phosphorylation of eIF4E is required for translational up-regulation of several proteins implicated in tumorigenesis. Accordingly, increased phospho-eIF4E levels correlate with disease progression in patients with prostate cancer. Our findings establish eIF4E phosphorylation as a critical event in tumorigenesis. These findings raise the possibility that chemical compounds that prevent the phosphorylation of eIF4E could act as anticancer drugs. PTEN | translational controlA berrations in the control of mRNA translation initiation have been documented in many tumor types (1-4). Translation initiation is controlled in part by eIF4E, the mRNA 5′ cap-binding protein. eIF4E is a proto-oncogene, inasmuch as its overexpression in immortalized rodent fibroblasts or human epithelial cells causes transformation (5, 6), and in mouse models its overexpression engenders tumor formation (7,8). eIF4E is phosphorylated by the MNK1/2 serine/threonine kinases, which are activated in response to mitogenic and stress signaling downstream of ERK1/2 and p38 MAP kinase, respectively (9, 10). eIF4E phosphorylation at serine 209 by MNK1/2 promotes its transformation activity (11,12). To study the role of eIF4E phosphorylation in tumorigenesis in the whole organism, we generated a knock-in (KI) mouse in which eIF4E serine 209 was mutated to alanine. Here, we show that mouse embryonic fibroblasts (MEFs) isolated from eIF4E S209A/S209A embryos display a marked resistance to oncogene-induced transformation. Furthermore, the mutant mice are viable, but are resistant to development of Pten loss-induced prostate cancer, and this resistance is associated with a decrease in MMP3, CCL2, VEGFC, and BIRC2 proteins. Moreover, eIF4E is highly phosphorylated in hormone-refractory prostate cancer, which correlates with poor clinical outcome. These results demonstrate the importance of eIF4E phosphorylation in tumorigenesis and validate the eIF4E phosphorylation pathway as a potential therapeutic target for cancer. ResultsSer209 Is the Only Phosphorylation Site in eIF4E. To address the role of eIF4E phosphorylation in tumorigenesis, a knock-in (KI) mouse in which serine 209 was replaced by an alanine residue was generated. The strategy and targeting vector construction for the generation, selection, and genotyping of the S209A mice is shown in Fig. S1. The eIF4E S...

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