scholarly journals mRNA-Selective Translation Induced by FSH in Primary Sertoli Cells

2012 ◽  
Vol 26 (4) ◽  
pp. 669-680 ◽  
Author(s):  
Astrid Musnier ◽  
Kelly León ◽  
Julia Morales ◽  
Eric Reiter ◽  
Thomas Boulo ◽  
...  
Keyword(s):  
Sertoli Cells ◽  
Mammalian Target Of Rapamycin ◽  
Initiation Factor ◽  
Target Of Rapamycin ◽  
Target Cells ◽  
Eukaryotic Initiation Factor ◽  
S6 Kinase ◽  
P70 S6 Kinase ◽  
Eukaryotic Initiation Factor 4E ◽  
Selective Translation

Abstract FSH is a key hormonal regulator of Sertoli cell secretory activity, required to optimize sperm production. To fulfil its biological function, FSH binds a G protein-coupled receptor, the FSH-R. The FSH-R-transduced signaling network ultimately leads to the transcription or down-regulation of numerous genes. In addition, recent evidence has suggested that FSH might also regulate protein translation. However, this point has never been demonstrated conclusively yet. Here we have addressed this issue in primary rat Sertoli cells endogenously expressing physiological levels of FSH-R. We observed that, within 90 min of stimulation, FSH not only enhanced overall protein synthesis in a mammalian target of rapamycin-dependent manner but also increased the recruitment of mRNA to polysomes. m7GTP pull-down experiments revealed the functional recruitment of mammalian target of rapamycin and p70 S6 kinase to the 5′cap, further supported by the enhanced phosphorylation of one of p70 S6 kinase targets, the eukaryotic initiation factor 4B. Importantly, the scaffolding eukaryotic initiation factor 4G was also recruited, whereas eukaryotic initiation factor 4E-binding protein, the eukaryotic initiation factor 4E generic inhibitor, appeared to play a minor role in translational regulations induced by FSH, in contrast to what is generally observed in response to anabolic factors. This particular regulation of the translational machinery by FSH stimulation might support mRNA-selective translation, as shown here by quantitative RT-PCR amplification of the c-fos and vascular endothelial growth factor mRNA but not of all FSH target mRNA, in polysomal fractions. These findings add a new level of complexity to FSH biological roles in its natural target cells, which has been underappreciated so far.

scholarly journals Ku-0063794 is a specific inhibitor of the mammalian target of rapamycin (mTOR)

2009 ◽  
Vol 421 (1) ◽  
pp. 29-42 ◽  
Author(s):  
Juan M. García-Martínez ◽  
Jennifer Moran ◽  
Rosemary G. Clarke ◽  
Alex Gray ◽  
Sabina C. Cosulich ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Cell Growth ◽  
Mammalian Target Of Rapamycin ◽  
Specific Inhibitor ◽  
Initiation Factor ◽  
Target Of Rapamycin ◽  
S6 Kinase ◽  
Class 1 ◽  
Hydrophobic Motif ◽  
Ribosomal S6 Kinase

mTOR (mammalian target of rapamycin) stimulates cell growth by phosphorylating and promoting activation of AGC (protein kinase A/protein kinase G/protein kinase C) family kinases such as Akt (protein kinase B), S6K (p70 ribosomal S6 kinase) and SGK (serum and glucocorticoid protein kinase). mTORC1 (mTOR complex-1) phosphorylates the hydrophobic motif of S6K, whereas mTORC2 phosphorylates the hydrophobic motif of Akt and SGK. In the present paper we describe the small molecule Ku-0063794, which inhibits both mTORC1 and mTORC2 with an IC50 of ∼10 nM, but does not suppress the activity of 76 other protein kinases or seven lipid kinases, including Class 1 PI3Ks (phosphoinositide 3-kinases) at 1000-fold higher concentrations. Ku-0063794 is cell permeant, suppresses activation and hydrophobic motif phosphorylation of Akt, S6K and SGK, but not RSK (ribosomal S6 kinase), an AGC kinase not regulated by mTOR. Ku-0063794 also inhibited phosphorylation of the T-loop Thr308 residue of Akt phosphorylated by PDK1 (3-phosphoinositide-dependent protein kinase-1). We interpret this as implying phosphorylation of Ser473 promotes phosphorylation of Thr308 and/or induces a conformational change that protects Thr308 from dephosphorylation. In contrast, Ku-0063794 does not affect Thr308 phosphorylation in fibroblasts lacking essential mTORC2 subunits, suggesting that signalling processes have adapted to enable Thr308 phosphorylation to occur in the absence of Ser473 phosphorylation. We found that Ku-0063794 induced a much greater dephosphorylation of the mTORC1 substrate 4E-BP1 (eukaryotic initiation factor 4E-binding protein 1) than rapamycin, even in mTORC2-deficient cells, suggesting a form of mTOR distinct from mTORC1, or mTORC2 phosphorylates 4E-BP1. Ku-0063794 also suppressed cell growth and induced a G1-cell-cycle arrest. Our results indicate that Ku-0063794 will be useful in delineating the physiological roles of mTOR and may have utility in treatment of cancers in which this pathway is inappropriately activated.

scholarly journals Immunopurified Mammalian Target of Rapamycin Phosphorylates and Activates p70 S6 Kinase αin Vitro

1999 ◽  
Vol 274 (48) ◽  
pp. 34493-34498 ◽  
Author(s):  
Shuji Isotani ◽  
Kenta Hara ◽  
Chiharu Tokunaga ◽  
Hitomi Inoue ◽  
Joseph Avruch ◽  
...  
Keyword(s):  
Mammalian Target Of Rapamycin ◽  
Target Of Rapamycin ◽  
S6 Kinase ◽  
P70 S6 Kinase

scholarly journals Serotonin Increases Phosphorylation of Synaptic 4EBP through TOR, but Eukaryotic Initiation Factor 4E Levels Do Not Limit Somatic Cap-Dependent Translation in Aplysia Neurons

2006 ◽  
Vol 26 (22) ◽  
pp. 8586-8598 ◽  
Author(s):  
Matthew Carroll ◽  
John Dyer ◽  
Wayne S. Sossin
Keyword(s):  
Binding Protein ◽  
Aplysia Californica ◽  
Memory Formation ◽  
Initiation Factor ◽  
Target Of Rapamycin ◽  
Phosphorylation Sites ◽  
Eukaryotic Initiation Factor ◽  
Cell Soma ◽  
Aplysia Neurons ◽  
Eukaryotic Initiation Factor 4E

ABSTRACT The target of rapamycin (TOR) plays an important role in memory formation in Aplysia californica. Here, we characterize one of the downstream targets of TOR, the eukaryotic initiation factor 4E (eIF4E) binding protein (4EBP) from Aplysia. Aplysia 4EBP contains the four critical phosphorylation sites regulated by TOR as well as an N-terminal RAIP motif and a C-terminal TOS site. Aplysia 4EBP was hypophosphorylated in synaptosomes, and serotonin addition caused a rapamycin-sensitive increase in 4EBP phosphorylation both in synaptosomes and in isolated neurites. Aplysia 4EBP was regulated in a fashion similar to that of mammalian 4EBPs, binding to eIF4E when dephosphorylated and releasing eIF4E after phosphorylation. Overexpression of 4EBP in the soma of Aplysia neurons caused a specific decrease in cap-dependent translation that was rescued by concomitant overexpression of eIF4E. However, eIF4E overexpression by itself did not increase cap-dependent translation, suggesting that increasing levels of free eIF4E by phosphorylating 4EBP is not important in regulating cap-dependent translation in the cell soma. Total levels of eIF4E were also regulated by 4EBP, suggesting that 4EBP can also act as an eIF4E chaperone. These studies demonstrate the conserved nature of 4EBP regulation and its role in cap-dependent translation and suggest differential roles of 4EBP phosphorylation in the soma and synapse.

A dose escalation, single arm, phase Ib/II combination study of BEZ235 with everolimus to determine the safety, pharmacodynamics, and pharmacokinetics in subjects with advanced solid malignancies including glioblastoma multiforme.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. TPS3116-TPS3116
Author(s):  
Mohamad Adham Salkeni ◽  
Muhammad Shaalan Beg ◽  
Olugbenga Olanrele Olowokure ◽  
Hassana Fathallah ◽  
Hala Thomas ◽  
...  
Keyword(s):  
Glioblastoma Multiforme ◽  
Solid Tumors ◽  
Mammalian Target Of Rapamycin ◽  
Competitive Inhibitor ◽  
Initiation Factor ◽  
Phase 2 ◽  
S6 Kinase ◽  
Eukaryotic Initiation Factor 4E ◽  
Solid Malignancies ◽  
Phase 1B

TPS3116 Background: Downregulation of a number of signaling pathways, including the mammalian target of rapamycin (mTOR) pathway, has been demonstrated to be efficacious in a large range of solid tumors such as breast, colon, endometrial, glial and hepatocellular carcinoma (HCC). However, we find that rapamycins lead to suppression of a negative feedback loop from S6 Kinase 1 (S6K1) to Protein Kinase B (PKB), leading to hyperactivation of PKB. In pre-clinical studies using a mouse model of carcinogen-induced HCC, we have demonstrated that combining BEZ235 (a potent and highly selective reversible ATP site competitive inhibitor of PI3K and mTOR) with everolimus (an allosteric inhibitor of mTOR) synergizes to inhibit tumor growth. BEZ235, an orally administered agent, has demonstrated preliminary antitumor activity in a first-in-human phase I study. The current study will evaluate this combination in patients with a variety of solid malignancies that includes glioblastoma multiforme (GBM). Methods: This study is divided into a phase 1b portion designed to determine safety of increasing doses of the combination, with extensive pharmacokinetics, pharmacodynamics and pharmacogenomics analysis; and a phase 2 portion that includes both solid tumors and GBM based on predominance of the mTOR and PI3K deregulation in these tumors, to determine preliminary anti-tumor activity and the recommended dose for phase 2 studies. We will also integrate biomarker assessment for gene expression products of the mTOR downstream pathway such as eukaryotic initiation factor 4E binding protein (4EBP1) and S6 kinase (S6K). The phase 1b portion has started accruing.

Activation of mammalian target of rapamycin and the p70 S6 kinase by arsenic trioxide in BCR-ABL–expressing cells

2006 ◽  
Vol 5 (11) ◽  
pp. 2815-2823 ◽  
Author(s):  
Patrick Yoon ◽  
Nick Giafis ◽  
Jessica Smith ◽  
Heather Mears ◽  
Efstratios Katsoulidis ◽  
...  
Keyword(s):  
Arsenic Trioxide ◽  
Mammalian Target Of Rapamycin ◽  
Target Of Rapamycin ◽  
S6 Kinase ◽  
P70 S6 Kinase
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