Regulation of PTEN translation by PI3K signaling maintains pathway homeostasis.

The PI3K pathway regulates cell metabolism, proliferation, and migration, and its dysregulation is common in cancer. We now show that both physiologic and oncogenic activation of PI3K signaling increase the expression of its negative regulator PTEN. This limits the duration of the signal and output of the pathway. Physiologic and pharmacologic inhibition of the pathway reduces PTEN and contributes to the rebound in pathway activity in tumors treated with PI3K inhibitors and limits their efficacy. Regulation of PTEN is due to mTOR/4E-BP1-dependent control of its translation and is lost when 4E-BP1 is deleted. Translational regulation of PTEN is therefore a major homeostatic regulator of physiologic PI3K signaling and plays a role in reducing the pathway activation by oncogenic PIK3CA mutants and the antitumor activity of PI3K pathway inhibitors. However, pathway output is hyperactivated in tumor cells with coexistent PI3K mutation and loss of PTEN function.

CDK12 phosphorylates 4E-BP1 to enable mTORC1-dependent translation and mitotic genome stability.

The RNA polymerase II (RNAPII) C-terminal domain kinase, CDK12, regulates genome stability, expression of DNA repair genes, and cancer cell resistance to chemotherapy and immunotherapy. In addition to its role in mRNA biosynthesis of DNA repair genes, we show here that CDK12 phosphorylates the mRNA 5' cap-binding repressor, 4E-BP1, to promote translation of mTORC1-dependent mRNAs. In particular, we found that phosphorylation of 4E-BP1 by mTORC1 (T37 and T46) facilitates subsequent CDK12 phosphorylation at two Ser-Pro sites (S65 and T70) that control the exchange of 4E-BP1 with eIF4G at the 5' cap of CHK1 and other target mRNAs. RNA immunoprecipitation coupled with deep sequencing (RIP-seq) revealed that CDK12 regulates release of 4E-BP1, and binding of eIF4G, to many mTORC1 target mRNAs, including those needed for MYC transformation. Genome-wide ribosome profiling (Ribo-seq) further identified specific CDK12 "translation-only" target mRNAs, including many mTORC1 target mRNAs as well as many subunits of mitotic and centromere/centrosome complexes. Accordingly, confocal imaging analyses revealed severe chromosome misalignment, bridging, and segregation defects in cells deprived of CDK12 or CCNK. We conclude that the nuclear RNAPII-CTD kinase CDK12 cooperates with mTORC1, and controls a specialized translation network that is essential for mitotic chromosome stability.

A Mad2-Mediated Translational Regulatory Mechanism Promoting S-Phase Cyclin Synthesis Controls Origin Firing and Survival to Replication Stress.

Cell survival to replication stress depends on the activation of the Mec1ATR-Rad53 checkpoint response that protects the integrity of stalled forks and controls the origin firing program. Here we found that Mad2, a member of the spindle assembly checkpoint (SAC), contributes to efficient origin firing and to cell survival in response to replication stress. We show that Rad53 and Mad2 promote S-phase cyclin expression through different mechanisms: while Rad53 influences Clb5,6 degradation, Mad2 promotes their protein synthesis. We found that Mad2 co-sediments with polysomes and modulates the association of the translation inhibitor Caf204E-BP with the translation machinery and the initiation factor eIF4E. This Mad2-dependent translational regulatory process does not depend on other SAC proteins. Altogether our observations indicate that Mad2 has an additional function outside of mitosis to control DNA synthesis and collaborates with the Mec1-Rad53 regulatory axis to allow cell survival in response to replication stress.

Selective Inhibition of mTORC1 Signaling Supports the Development and Maintenance of Pluripotency.

Insight into the molecular mechanisms governing the development and maintenance of pluripotency is important for understanding early development and the use of stem cells in regenerative medicine. We demonstrate the selective inhibition of mTORC1 signaling is important for developing the inner cell mass (ICM) and the self-renewal of human embryonic stem cells. S6K suppressed the expression and function of pluripotency-related transcription factors (PTFs) OCT4, SOX2, and KLF4 through phosphorylation and ubiquitin proteasome-mediated protein degradation, indicating that S6K inhibition is required for pluripotency. PTFs inhibited mTOR signaling. The phosphorylation of S6 was decreased in PTF-positive cells of the ICM in embryos. Activation of mTORC1 signaling blocked ICM formation and the selective inhibition of S6K by rapamycin increased the ICM size in mouse blastocysts. Thus, selective inhibition of mTORC1 signaling supports the development and maintenance of pluripotency.

mTOR Controls Mitochondrial Dynamics and Cell Survival via MTFP1.

The mechanisms that link environmental and intracellular stimuli to mitochondrial functions, including fission/fusion, ATP production, metabolite biogenesis, and apoptosis, are not well understood. Here, we demonstrate that the nutrient-sensing mechanistic/mammalian target of rapamycin complex 1 (mTORC1) stimulates translation of mitochondrial fission process 1 (MTFP1) to control mitochondrial fission and apoptosis. Expression of MTFP1 is coupled to pro-fission phosphorylation and mitochondrial recruitment of the fission GTPase dynamin-related protein 1 (DRP1). Potent active-site mTOR inhibitors engender mitochondrial hyperfusion due to the diminished translation of MTFP1, which is mediated by translation initiation factor 4E (eIF4E)-binding proteins (4E-BPs). Uncoupling MTFP1 levels from the mTORC1/4E-BP pathway upon mTOR inhibition blocks the hyperfusion response and leads to apoptosis by converting mTOR inhibitor action from cytostatic to cytotoxic. These data provide direct evidence for cell survival upon mTOR inhibition through mitochondrial hyperfusion employing MTFP1 as a critical effector of mTORC1 to govern cell fate decisions.

Yeast Tor complex 1 phosphorylates eIF4E-binding protein, Caf20.

Tor complex 1 (TORC1), a master regulator of cell growth, is an evolutionarily conserved protein kinase within eukaryotic organisms. To control cell growth, TORC1 governs translational processes by phosphorylating its substrate proteins in response to cellular nutritional cues. Mammalian TORC1 (mTORC1) assumes the responsibility of phosphorylating the eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1) to regulate its interaction with eIF4E. The budding yeast Saccharomyces cerevisiae possesses a pair of 4E-BP genes, CAF20 and EAP1. However, the extent to which the TORC1-4E-BP axis regulates translational initiation in yeast remains uncertain. In this study, we demonstrated the influence of TORC1 on the phosphorylation status of Caf20 in vivo, as well as the direct phosphorylation of Caf20 by TORC1 in vitro. Furthermore, we found the TORC1-dependent recruitment of Caf20 to the 80S ribosome. Consequently, our study proposes a plausible involvement of yeast's 4E-BP in the efficacy of translation initiation, an aspect under the control of TORC1.

PTEN and P-4E-BP1 might be associated with postoperative recurrence of rectal cancer patients undergoing concurrent radiochemotherapy.

BACKGROUND: Local recurrence after surgery and radiochemotherapy seriously affects the prognosis of locally advanced rectal cancer (LARC) patients. Studies on molecular markers related to the radiochemotherapy sensitivity of cancers have been widely carried out, which might provide valued information for clinicians to carry out individual treatment. AIM: To find potential biomarkers of tumors for predicting postoperative recurrence. METHODS: In this study, LARC patients undergoing surgery and concurrent radiochemotherapy were enrolled. We focused on clinicopathological factors and PTEN, SIRT1, p-4E-BP1, and pS6 protein expression assessed by immunohistochemistry in 73 rectal cancer patients with local recurrence and 76 patients without local recurrence. RESULTS: The expression of PTEN was higher, while the expression of p-4E-BP1 was lower in patients without local recurrence than in patients with local recurrence. Moreover, TNM stage, lymphatic vessel invasion (LVI), PTEN and p-4E-BP1 might be independent risk factors for local recurrence after LARC surgery combined with concurrent radiochemotherapy. CONCLUSIONS: This study suggests that PTEN and p-4E-BP1 might be potential biomarkers for prognostic prediction and therapeutic targets for LARC.

Inhibition of the rapamycin-insensitive mTORC1 /4E-BP1 axis attenuates TGF-ß1-induced fibrotic response in human Tenon's fibroblasts.

Subconjunctival fibrosis is the major cause of failure in both conventional and modern minimally invasive glaucoma surgeries (MIGSs) with subconjunctival filtration. The search for safe and effective anti-fibrotic agents is critical for improving long-term surgical outcomes. In this study, we investigated the effect of inhibiting the rapamycin-insensitive mTORC1/4E-BP1 axis on the transforming growth factor-beta 1(TGF-ß1)-induced fibrotic responses in human Tenon's fibroblasts (HTFs), as well as in a rat model of glaucoma filtration surgery (GFS). Primary cultured HTFs were treated with 3 ng/mL TGF-ß1 for 24 h, followed by treatment with 10 µM CZ415 for additional 24 h. Rapamycin (10 µM) was utilized as a control for mTORC1/4E-BP1 signaling insensitivity. The expression levels of fibrosis-associated molecules were measured using quantitative real-time PCR, Western blotting, and immunofluorescence analysis. Cell migration was assessed through the scratch wound assay. Additionally, a rat model of GFS was employed to evaluate the anti-fibrotic effect of CZ415 in vivo. Our findings indicated that both rapamycin and CZ415 treatment significantly reduced the TGF-ß1-induced cell proliferation, migration, and the expression of pro-fibrotic factors in HTFs. CZ415 also more effectively inhibited TGF-ß1-mediated collagen synthesis in HTFs compared to rapamycin. Activation of mTORC1/4E-BP signaling following TGF-ß1 exposure was highly suppressed by CZ415 but was only modestly inhibited by rapamycin. Furthermore, CZ415 was found to decrease subconjunctival collagen deposition in rats post GFS. Our results suggest that rapamycin-insensitive mTORC1/4E-BP1 signaling plays a critical role in TGF-ß1-driven collagen synthesis in HTFs. This study demonstrated that inhibition of the mTORC1/4E-BP1 axis offers superior anti-fibrotic efficacy compared to rapamycin and represents a promising target for improving the success rate of both traditional and modern GFSs.

The Structures of eIF4E-eIF4G Complexes Reveal an Extended Interface to Regulate Translation Initiation.

Eukaryotic initiation factor 4G (eIF4G) plays a central role in translation initiation through its interactions with the cap-binding protein eIF4E. This interaction is a major drug target for repressing translation and is naturally regulated by 4E-binding proteins (4E-BPs). 4E-BPs and eIF4G compete for binding to the eIF4E dorsal surface via a shared canonical 4E-binding motif, but also contain auxiliary eIF4E-binding sequences, which were assumed to contact non-overlapping eIF4E surfaces. However, it is unknown how metazoan eIF4G auxiliary sequences bind eIF4E. Here, we describe crystal structures of human and Drosophila melanogaster eIF4E-eIF4G complexes, which unexpectedly reveal that the eIF4G auxiliary sequences bind to the lateral surface of eIF4E, using a similar mode to that of 4E-BPs. Our studies provide a molecular model of the eIF4E-eIF4G complex, shed light on the competition mechanism of 4E-BPs, and enable the rational design of selective eIF4G inhibitors to dampen dysregulated translation in disease.

The rate of protein synthesis in hematopoietic stem cells is limited partly by 4E-BPs.

Adult stem cells must limit their rate of protein synthesis, but the underlying mechanisms remain largely unexplored. Differences in protein synthesis among hematopoietic stem cells (HSCs) and progenitor cells did not correlate with differences in proteasome activity, total RNA content, mRNA content, or cell division rate. However, adult HSCs had more hypophosphorylated eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP2 as compared with most other hematopoietic progenitors. Deficiency for 4E-BP1 and 4E-BP2 significantly increased global protein synthesis in HSCs, but not in other hematopoietic progenitors, and impaired their reconstituting activity, identifying a mechanism that promotes HSC maintenance by attenuating protein synthesis.

Protein Translation in the Pathogenesis of Parkinson's Disease.

In recent years, research into Parkinson's disease and similar neurodegenerative disorders has increasingly suggested that these conditions are synonymous with failures in proteostasis. However, the spotlight of this research has remained firmly focused on the tail end of proteostasis, primarily aggregation, misfolding, and degradation, with protein translation being comparatively overlooked. Now, there is an increasing body of evidence supporting a potential role for translation in the pathogenesis of PD, and its dysregulation is already established in other similar neurodegenerative conditions. In this paper, we consider how altered protein translation fits into the broader picture of PD pathogenesis, working hand in hand to compound the stress placed on neurons, until this becomes irrecoverable. We will also consider molecular players of interest, recent evidence that suggests that aggregates may directly influence translation in PD progression, and the implications for the role of protein translation in our development of clinically useful diagnostics and therapeutics.

Inhibiting eukaryotic initiation factor 5A (eIF5A) hypusination attenuated activation of the SIK2 (salt-inducible kinase 2)-p4E-BP1 pathway involved in ovarian cancer cell proliferation and migration.

BACKGROUND: Eukaryotic initiation factor 5A hypusine (eIF5AHyp) stimulates the translation of proline repeat motifs. Salt inducible kinase 2 (SIK2) containing a proline repeat motif is overexpressed in ovarian cancers, in which it promotes cell proliferation, migration, and invasion. METHODS AND RESULTS: Western blotting and dual luciferase analyses showed that depletion of eIF5AHyp by GC7 or eIF5A-targeting siRNA downregulated SIK2 level and decreased luciferase activity in cells transfected with a luciferase-based reporter construct containing consecutive proline residues, whereas the activity of the mutant control reporter construct (replacing P825L, P828H, and P831Q) did not change. According to the MTT assay, GC7, which has a potential antiproliferative effect, reduced the viability of several ovarian cancer cell lines by 20-35% at high concentrations (ES2 > CAOV-3 > OVCAR-3 > TOV-112D) but not at low concentrations. In a pull-down assay, we identified eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP1 (p4E-BP1) phosphorylated at Ser 65 as downstream binding partners of SIK2, and we validated that the level of p4E-BP1(Ser 65) was downregulated by SIK2-targeting siRNA. Conversely, in ES2 cells overexpressing SIK2, the p4E-BP1(Ser 65) level was increased but decreased in the presence of GC7 or eIF5A-targeting siRNA. Finally, the migration, clonogenicity, and viability of ES2 ovarian cancer cells were reduced by GC7 treatment as well as by siRNA for eIF5A gene silencing and siRNA for SIK2 and 4E-BP1 gene silencing. Conversely, those activities were increased in cells overexpressing SIK2 or 4E-BP1 and decreased again in the presence of GC7. CONCLUSION: The depletion of eIF5AHyp by GC7 or eIF5A-targeting siRNA attenuated activation of the SIK2-p4EBP1 pathway. In that way, eIF5AHyp depletion reduces the migration, clonogenicity, and viability of ES2 ovarian cancer cells.

Nuclear-targeted 4E-BP1 is dephosphorylated, induces nuclear translocation of eIF4E, and alters mRNA translation.

Mechanistic target of rapamycin complex 1 (mTORC1) phosphorylates and inhibits eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1). This leads to the release of eIF4E from 4E-BP1 and the initiation of eIF4E-dependent mRNA translation. In this study, we examined the expression of a 4E-BP1-based reporter (mTORC1 activity reporter; TORCAR) with various localization signal tags to clarify the relationship between the localization of 4E-BP1 and its phosphorylation. Phosphorylation of 4E-BP1 at threonine 37/46 and serine 65 was efficient at lysosomes and the plasma membrane, whereas it was significantly decreased in the nucleus. In addition, the localization of endogenous eIF4E shifted from the cytoplasm to the nucleus only when nuclear-localized TORCAR was expressed. Nuclear-localized TORCAR decreased cyclin D1 protein levels and altered cell cycle distribution. These data provide an experimental tool to manipulate the localization of endogenous eIF4E without affecting mTORC1 and highlight the important role of nuclear-cytoplasmic shuttling of eIF4E.

Glutamine stimulates the S6K/4E-BP branch of insulin signalling pathway to mitigate human poly(Q) disorders in Drosophila disease models.

OBJECTIVE AND METHODS: Since, the S6K/4E-BP sub-pathway can be stimulated by various amino acids; we extended our investigation to examine if oral feeding of amino acids delivers rescue against human poly(Q) toxicity in Drosophila. We utilised Drosophila models of two different poly(Q) disorders to test our hypothesis. Glutamine was fed to the test flies orally mixed in the food. Control and treated flies were then tested for different parameters, such as formation of poly(Q) aggregates and neurodegeneration, to evaluate glutamine's proficiency in mitigating poly(Q) neurotoxicity. RESULTS: Our study, for the first time, reports that glutamine feeding stimulates the growth promoting S6K/4E-BP branch of insulin signalling pathway and restricts pathogenesis of poly(Q) disorders in Drosophila disease models. We noted that glutamine treatment restricts the formation of neurotoxic poly(Q) aggregates and minimises neuronal deaths. Further, glutamine treatment re-establishes the chromatin architecture by improving the histone acetylation which is otherwise compromised in poly(Q) expressing neuronal cells. DISCUSSION: Since, the insulin signalling pathway as well as mechanism of action of glutamine are fairly conserved between human and Drosophila, our finding strongly suggests that glutamine holds immense potential to be developed as an intervention therapy against the incurable human poly(Q) disorders.

The isolated effects of local cold application on proteolytic and myogenic signaling.

Post-exercise cooling studies reveal inhibitory effects on markers of skeletal muscle growth. However, the isolated effect of local cold application has not been adequately addressed. It is unclear if the local cold or the combination of local cold and exercise is driving negatively altered skeletal muscle gene expression. The purpose was to determine the effects of a 4 h local cold application to the vastus lateralis on the myogenic and proteolytic response. Participants (n = 12, 27 ± 6 years, 179 ± 9 cm, 82.8 ± 13.0 kg, 18.4 ± 7.1 %BF) rested with a thermal wrap placed on each leg with either circulating cold fluid (10 °C, COLD) or no fluid circulation (room temperature, RT). Muscle samples were collected to quantify mRNA (RT-qPCR) and proteins (Western Blot) associated with myogenesis and proteolysis. Temperatures in COLD were lower than RT at the skin (13.2 ± 1.0 °C vs. 34.8 ± 0.9 °C; p < 0.001) and intramuscularly (20.5 ± 1.3 °C vs. 35.6 ± 0.8 °C, p < 0.001). Myogenic-related mRNA, MYO-G and MYO-D1, were lower in COLD (p = 0.001, p < 0.001, respectively) whereas myogenic-mRNA, MYF6, was greater in COLD (p = 0.002). No other myogenic associated genes were different between COLD and RT (MSTN, p = 0.643; MEF2a, p = 0.424; MYF5, p = 0.523; RPS3, p = 0.589; RPL3-L, p = 0.688). Proteolytic-related mRNA was higher in COLD (FOXO3a, p < 0.001; Atrogin-1, p = 0.049; MURF-1, p < 0.001). The phosphorylation:total protein ratio for the translational repressor of muscle mass, 4E-BP1Thr37/46, was lower in COLD (p = 0.043), with no differences in mTORser2448 (p = 0.509) or p70S6K1Thr389 (p = 0.579). Isolated local cooling over 4 h exhibits inhibited myogenic and higher proteolytic skeletal muscle molecular response.

The independent effects of local heat application on muscle growth program associated mRNA and protein phosphorylation.

The development and maintenance of skeletal muscle is crucial for the support of daily function. Recent evidence suggests that genes coded for proteins associated with the human muscle growth program (myogenic and proteolytic genes) are sensitive to local heat application. Therefore, the purpose of this investigation was to determine the effect of 4 h of local heat application to the vastus lateralis at rest on acute phosphorylation (mTORSer2448, p70-S6K1Thr389, and 4E-BP1Thr47/36) and gene expression changes for proteins associated with the muscle growth program. Intramuscular temperature of the HOT limb was 1.2 ± 0.2 °C higher than CON limb after 4 h of local heating. However, this local heat stimulus did not influence transcription of genes associated with myogenesis (MSTN, p = 0.321; MYF5, p = 0.445; MYF6, p = 0.895; MEF2a, p = 0.809; MYO-G, p = 0.766; MYO-D1, p = 0.118; RPS3, p = 0.321; and RPL-3L, p = 0.577), proteolysis (Atrogin-1, p = 0.573; FOXO3a, p = 0.452; MURF-1, p = 0.284), nor protein phosphorylation (mTORSer2448, p = 0.981; P70-S6K1Thr389, p = 0.583; 4E-BP1Thr37/46, p = 0.238) associated with the muscle growth program. These findings suggest little to no association between the local application of heat, at rest, and the activation of the observed muscle growth program-related markers.

Mextli proteins use both canonical bipartite and novel tripartite binding modes to form eIF4E complexes that display differential sensitivity to 4E-BP regulation.

The eIF4E-binding proteins (4E-BPs) are a diverse class of translation regulators that share a canonical eIF4E-binding motif (4E-BM) with eIF4G. Consequently, they compete with eIF4G for binding to eIF4E, thereby inhibiting translation initiation. Mextli (Mxt) is an unusual 4E-BP that promotes translation by also interacting with eIF3. Here we present the crystal structures of the eIF4E-binding regions of the Drosophila melanogaster (Dm) and Caenorhabditis elegans (Ce) Mxt proteins in complex with eIF4E in the cap-bound and cap-free states. The structures reveal unexpected evolutionary plasticity in the eIF4E-binding mode, with a classical bipartite interface for Ce Mxt and a novel tripartite interface for Dm Mxt. Both interfaces comprise a canonical helix and a noncanonical helix that engage the dorsal and lateral surfaces of eIF4E, respectively. Remarkably, Dm Mxt contains a C-terminal auxiliary helix that lies anti-parallel to the canonical helix on the eIF4E dorsal surface. In contrast to the eIF4G and Ce Mxt complexes, the Dm eIF4E-Mxt complexes are resistant to competition by bipartite 4E-BPs, suggesting that Dm Mxt can bind eIF4E when eIF4G binding is inhibited. Our results uncovered unexpected diversity in the binding modes of 4E-BPs, resulting in eIF4E complexes that display differential sensitivity to 4E-BP regulation.

Prothioconazole induces cell cycle arrest by up-regulation of EIF4EBP1 in extravillous trophoblast cells.

Prothioconazole (PTC) is a new broad-spectrum triazole antibacterial agent that is being widely used in agriculture. PTC has been linked to a number of reproductive outcomes including embryo implantation disorder; however, the exact mechanism underlying this relationship has yet to be determined. Proper trophoblast proliferation and migration is a prerequisite for successful embryo implantation. To elucidate the underlying molecular perturbations, we detect the effect of PTC on extravillous trophoblast cells proliferation and migration, and investigate its potential mechanisms. Exposure to different concentrations of PTC (0-500 µM) significantly inhibited the cell viability and migration ability (5 µM PTC exposure), and also caused the cell cycle arrest at the lowest dose (1 µM PTC exposure). Transcriptome analysis revealed that PTC exposure disturbed multiple biological processes including cell cycle and apoptosis, consistent with cell phenotype. Specifically, eukaryotic translation initiation factor 4E binding protein 1 (EIF4EBP1, 4E-BP1) was identified as up-regulated in PTC exposure group and knockdown of EIF4EBP1, and attenuated the G1 phase arrest induced by PTC exposure. In summary, our data demonstrated that 4E-BP1 participated in PTC-induced cell cycle arrest in extravillous trophoblast cells by regulating cyclin D1. These findings shed light on the potential adverse effect of PTC exposure on the embryo implantation.

The expression and significance of p4E-BP1/4E-BP1 in prostate cancer.

BACKGROUND: Although the phosphorylation of 4E-BP1 that has been detected in high-grade prostate cancer has been reported in previous studies, overexpression of p4E-BP1 and 4EBP1 and their clinical significance in prostate cancer still remain unknown. METHODS: One hundred six samples of prostate tissues were collected and analyzed by immunohistochemistry with p4E-BP1 or 4E-BP1 specific antibodies. Everolimus was used to block the phosphorylation of p4E-BP1, and then flow cytometry, clone formation, transwell, and wound healing assays were performed to detect the survival and invasive ability of the prostate cancer cells. RESULTS: We found that the expression of 4E-BP1 and p4E-BP1 was higher in prostate cancer tissues than in normal tissues. Interestingly, the expression of p4E-BP1 was significantly associated with Gleason score and lymph node metastasis, but had no obvious correlation with PSA and the presence of bone or visceral metastasis. However, no evident correlation was found between the positive expression of 4E-BP1 and these clinical characteristics. In in vitro experiments, we found similar results as the clinical presentation that 4E-BP1 and p4E-BP1 were low expressed in normal prostate epithelial cells, but in prostate cancer cells, as the malignancy increasing, 4E-BP1 and p4E-BP1 expression also gradually increased. Then, we used Everolimus to inhibit the phosphorylation of 4E-BP1 and found that Everolimus effectively reduced cloning formation, inhibited cell migration, and promoted apoptosis in a dose-dependent manner in PC3 cells. CONCLUSIONS: These findings suggest that p4E-BP1 is a potential biomarker and therapy target for prostate cancer, and patients with high expressions of p4E-BP1 may benefit from Everolimus treatment.

Inhibitory interneurons mediate autism-associated behaviors via 4E-BP2.

Translational control plays a key role in regulation of neuronal activity and behavior. Deletion of the translational repressor 4E-BP2 in mice alters excitatory and inhibitory synaptic functions, engendering autistic-like behaviors. The contribution of 4E-BP2-dependent translational control in excitatory and inhibitory neurons and astrocytic cells to these behaviors remains unknown. To investigate this, we generated cell-type-specific conditional 4E-BP2 knockout mice and tested them for the salient features of autism, including repetitive stereotyped behaviors (self-grooming and marble burying), sociability (3-chamber social and direct social interaction tests), and communication (ultrasonic vocalizations in pups). We found that deletion of 4E-BP2 in GABAergic inhibitory neurons, defined by Gad2, resulted in impairments in social interaction and vocal communication. In contrast, deletion of 4E-BP2 in forebrain glutamatergic excitatory neurons, defined by Camk2a, or in astrocytes, defined by Gfap, failed to cause autistic-like behavioral abnormalities. Taken together, we provide evidence for an inhibitory-cell-specific role of 4E-BP2 in engendering autism-related behaviors.