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Recent evidence suggests that resistance training with light or heavy loads to failure results in similar adaptations. Herein, we compared how both training modalities affect the molecular, neuromuscular, and recovery responses following exercise. Resistance‐trained males (mean ± SE: 22 ± 2 years, 84.8 ± 9.0 kg, 1.79 ± 0.06 m; n = 15) performed a crossover design of four sets of leg extensor exercise at 30% (light RE) or 80% (heavy RE) one repetition maximum (1RM) to repetition failure, and heavy RE or light RE 1 week later. Surface electromyography (EMG) was monitored during exercise, and vastus lateralis muscle biopsies were collected at baseline (PRE), 15 min (15mPOST), and 90 min following RE (90mPOST) for examination of molecular targets and fiber typing. Isokinetic dynamometry was also performed before (PRE), immediately after (POST), and 48 h after (48hPOST) exercise. Dependent variables were analyzed using repeated measures ANOVAs and significance was set at P ≤ 0.05. Repetitions completed were greater during light RE (P < 0.01), while EMG amplitude was greater during heavy RE (P ≤ 0.01). POST isokinetic torque was reduced following light versus heavy RE (P < 0.05). Postexercise expression of mRNAs and phosphoproteins associated with muscle hypertrophy were similar between load conditions. Additionally, p70s6k (Thr389) phosphorylation and fast‐twitch fiber proportion exhibited a strong relationship after both light and heavy RE (r > 0.5). While similar mRNA and phosphoprotein responses to both modalities occurred, we posit that heavy RE is a more time‐efficient training method given the differences in total repetitions completed, lower EMG amplitude during light RE, and impaired recovery response after light RE. Similar skeletal muscle mRNA and phosphoprotein responses to light versus heavy resistance exercise (RE) occurred. However, we posit that heavy RE is a more efficient training method given that significantly less total repetitions completed were completed to elicit similar physiological responses, lower skeletal muscle electromyography amplitude occurred during light RE, and an impaired functional recovery response occurred after light RE.

A classic metabolic concept posits that insulin promotes energy storage and adipose expansion, while catecholamines stimulate release of adipose energy stores by hydrolysis of triglycerides through β-adrenergic receptor (βARs) and protein kinase A (PKA) signaling. Here, we have shown that a key hub in the insulin signaling pathway, activation of p70 ribosomal S6 kinase (S6K1) through mTORC1, is also triggered by PKA activation in both mouse and human adipocytes. Mice with mTORC1 impairment, either through adipocyte-specific deletion of Raptor or pharmacologic rapamycin treatment, were refractory to the well-known βAR-dependent increase of uncoupling protein UCP1 expression and expansion of beige/brite adipocytes (so-called browning) in white adipose tissue (WAT). Mechanistically, PKA directly phosphorylated mTOR and RAPTOR on unique serine residues, an effect that was independent of insulin/AKT signaling. Abrogation of the PKA site within RAPTOR disrupted βAR/mTORC1 activation of S6K1 without affecting mTORC1 activation by insulin. Conversely, a phosphomimetic RAPTOR augmented S6K1 activity. Together, these studies reveal a signaling pathway from βARs and PKA through mTORC1 that is required for adipose browning by catecholamines and provides potential therapeutic strategies to enhance energy expenditure and combat metabolic disease.

During pregnancy, the appropriate allocation of nutrients between the mother and the fetus is dominated by maternal–fetal interactions, which is primarily governed by the placenta. The syncytiotrophoblast (STB) lining at the outer surface of the placental villi is directly bathed in maternal blood and controls feto–maternal exchange. The STB is the largest multinucleated cell type in the human body, and is formed through syncytialization of the mononucleated cytotrophoblast. However, the physiological advantage of forming such an extensively multinucleated cellular structure remains poorly understood. Here, we discover that the STB uniquely adapts to nutrient stress by inducing the macropinocytosis machinery through repression of mammalian target of rapamycin (mTOR) signaling. In primary human trophoblasts and in trophoblast cell lines, differentiation toward a syncytium triggers macropinocytosis, which is greatly enhanced during amino acid shortage, induced by inhibiting mTOR signaling. Moreover, inhibiting mTOR in pregnant mice markedly stimulates macropinocytosis in the syncytium. Blocking macropinocytosis worsens the phenotypes of fetal growth restriction caused by mTOR-inhibition. Consistently, placentas derived from fetal growth restriction patients display: 1) Repressed mTOR signaling, 2) increased syncytialization, and 3) enhanced macropinocytosis. Together, our findings suggest that the unique ability of STB to undergo macropinocytosis serves as an essential adaptation to the cellular nutrient status, and support fetal survival and growth under nutrient deprivation.

Growth signals, such as extracellular nutrients and growth factors, have substantial effects on genome integrity; however, the direct underlying link remains unclear. Here, we show that the mechanistic target of rapamycin (mTOR)–ribosomal S6 kinase (S6K) pathway, a central regulator of growth signalling, phosphorylates RNF168 at Ser60 to inhibit its E3 ligase activity, accelerate its proteolysis and impair its function in the DNA damage response, leading to accumulated unrepaired DNA and genome instability. Moreover, loss of the tumour suppressor liver kinase B1 ( LKB1 ; also known as STK11 ) hyperactivates mTOR complex 1 (mTORC1)–S6K signalling and decreases RNF168 expression, resulting in defects in the DNA damage response. Expression of a phospho-deficient RNF168-S60A mutant rescues the DNA damage repair defects and suppresses tumorigenesis caused by Lkb1 loss. These results reveal an important function of mTORC1–S6K signalling in the DNA damage response and suggest a general mechanism that connects cell growth signalling to genome stability control. Xie and colleagues find that activated mTORC1 growth signalling impairs DNA repair through S6K-mediated phosphorylation and inhibition of the RNF168 ligase.

Angiogenesis is a crucial step in the growth and metastasis of cancers, since it enables the growing tumor to receive oxygen and nutrients. Cancer prevention using natural products has become an integral part of cancer control. We studied the antiangiogenic activity of quercetin using ex vivo, in vivo and in vitro models. Rat aortic ring assay showed that quercetin at non-toxic concentrations significantly inhibited microvessel sprouting and exhibited a significant inhibition in the proliferation, migration, invasion and tube formation of endothelial cells, which are key events in the process of angiogenesis. Most importantly, quercetin treatment inhibited ex vivo angiogenesis as revealed by chicken egg chorioallantoic membrane assay (CAM) and matrigel plug assay. Western blot analysis showed that quercetin suppressed VEGF induced phosphorylation of VEGF receptor 2 and their downstream protein kinases AKT, mTOR, and ribosomal protein S6 kinase in HUVECs. Quercetin (20 mg/kg/d) significantly reduced the volume and the weight of solid tumors in prostate xenograft mouse model, indicating that quercetin inhibited tumorigenesis by targeting angiogenesis. Furthermore, quercetin reduced the cell viability and induced apoptosis in prostate cancer cells, which were correlated with the downregulation of AKT, mTOR and P70S6K expressions. Collectively the findings in the present study suggest that quercetin inhibits tumor growth and angiogenesis by targeting VEGF-R2 regulated AKT/mTOR/P70S6K signaling pathway, and could be used as a potential drug candidate for cancer therapy.

Emerging evidence suggests that the mammalian target of rapamcyin (mTOR) pathway is associated with radio-resistance in cancer treatment. We hypothesised that phosphorylated ribosomal S6 kinase 1 (p-S6K1), a major downstream regulator of the mTOR pathway, may play a role in predicting radio-resistance. Therefore, we evaluated the association of p-S6K1 expression with radio-resistance in breast cancer cell lines and patients. During median follow-up of 33 (range, 0.1–111) months for 1770 primary breast cancer patients who underwent surgery, patients expressing p-S6K1 showed worse 10-year loco-regional recurrence-free survival (LRFS) compared to that of p-S6K1-negative patients after radiotherapy (93.4% vs. 97.7%, p  = 0.015). Multivariate analysis revealed p-S6K1 expression as a predictor of radio-resistance (hazard ratio 7.9, 95% confidence interval 1.1–58.5, p  = 0.04). In vitro , CD44 high /CD24 low MCF7 cells with a radioresistant phenotype expressed higher levels of p-S6K1 than control MCF7 cells. Furthermore, the combination of radiation with treatment of everolimus, an mTOR-S6K1 pathway inhibitor, sensitised CD44 high /CD24 low MCF7 cells to a greater extent than MCF7 cells. This study provides in vivo and in vitro evidence for p-S6K1 expression status as an important marker for predicting the resistance to radiotherapy and as a possible target for radio-sensitization in breast cancer patients.

Deregulated AKT kinase activity due to PTEN deficiency in cancer cells contributes to oncogenesis by incompletely understood mechanisms. Here, we show that PTEN deletion in HCT116 and DLD1 colon carcinoma cells leads to suppression of CHK1 and CHK2 activation in response to irradiation, impaired G2 checkpoint proficiency and radiosensitization. These defects are associated with reduced expression of MRE11, RAD50 and NBS1, components of the apical MRE11/RAD50/NBS1 (MRN) DNA damage response complex. Consistent with reduced MRN complex function, PTEN-deficient cells fail to resect DNA double-strand breaks efficiently after irradiation and show greatly diminished proficiency for DNA repair via the error-free homologous recombination (HR) repair pathway. MRE11 is highly unstable in PTEN-deficient cells but stability can be significantly restored by inhibiting mTORC1 or p70S6 kinase (p70S6K), downstream kinases whose activities are stimulated by AKT, or by mutating a residue in MRE11 that we show is phosphorylated by p70S6K in vitro . In primary human fibroblasts, activated AKT suppresses MRN complex expression to escalate RAS-induced DNA damage and thereby reinforce oncogene-induced senescence. Taken together, our data demonstrate that deregulation of the PI3K-AKT/ mTORC1/ p70S6K pathways, an event frequently observed in cancer, exert profound effects on genome stability via MRE11 with potential implications for tumour initiation and therapy.

Aims/hypothesis Mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of nutritional status at the cellular and organismic level. While mTORC1 mediates beta cell growth and expansion, its hyperactivation has been observed in pancreatic islets from animal models of type 2 diabetes and leads to beta cell loss. We sought to determine whether such mTORC1 activation occurs in humans with type 2 diabetes or in metabolically stressed human islets and whether mTORC1 blockade can restore beta cell function of diabetic islets. Methods Human islets isolated from non-diabetic controls and individuals with type 2 diabetes, as well as human islets and INS-1E cells exposed to increased glucose (22.2 mmol/l), were examined for mTORC1/2 activity by western blotting analysis of phosphorylation of mTORC1 downstream targets ribosomal protein S6 kinase 1 (S6K1), S6 and eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1) and mTORC2 downstream targets Akt and N-myc downstream regulated 1 (NDRG1). mTORC1/2 complexes’ integrity was assessed by immunoprecipitation and subsequent western blot analysis. Cell-type specific expression of activated mTORC1 in human islets was examined by immunostaining of pS6 (Ser 235/236) in human islet sections. Beta cell function was measured by glucose-stimulated insulin secretion (GSIS). Results While mTORC2 signalling was diminished, mTORC1 activity was markedly increased in islets from patients with type 2 diabetes and in islets and beta cells exposed to increased glucose concentrations. Under high-glucose conditions in metabolically stressed human islets, we identified a reciprocal regulation of different mTOR complexes, with functional upregulation of mTORC1 and downregulation of mTORC2. pS6 immunostaining showed beta cell-specific upregulation of mTORC1 in islets isolated from patients with type 2 diabetes. Inhibition of mTORC1–S6K1 signalling improved GSIS and restored mTORC2 activity in islets from patients with type 2 diabetes as well as in islets isolated from diabetic db / db mice and mice fed a high-fat/high-sucrose diet. Conclusions/interpretation Our data show the aberrant mTORC1 activity in islets from patients with type 2 diabetes, in human islets cultured under diabetes-associated increased glucose conditions and in diabetic mouse islets. This suggests that elevated mTORC1 activation is a striking pathogenic hallmark of islets in type 2 diabetes, contributing to impaired beta cell function and survival in the presence of metabolic stress.

One characteristic of cancer cells is the abnormally high rate of cell metabolism to sustain their enhanced proliferation. However, the behind mechanism of this phenomenon is still elusive. Here we find that enhanced precursor 45s ribosomal RNA (pre-45s rRNA) is one of the core mechanisms in promoting the pathogenesis of colorectal cancer (CRC). Pre-45s rRNA expression is significantly higher in primary CRC tumor tissues samples and cancer cell lines compared with the non-tumorous colon tissues, and is associated with tumor sizes. Knockdown of pre-45s rRNA inhibits G1/S cell-cycle transition by stabilizing p53 through inducing murine double minute 2 (MDM2) and ribosomal protein L11 (RpL11) interaction. In addition, we revealed that high rate of cancer cell metabolism triggers the passive release of calcium ion from endoplasmic reticulum to the cytoplasm. The elevated calcium ion in the cytoplasm activates the signaling cascade of calcium/calmodulin-dependent protein kinase II, ribosomal S6 kinase (S6K) and ribosomal S6K (CaMKII-S6K-UBF). The activated UBF promotes the transcription of rDNA , which therefore increases pre-45s rRNA . Disruption of CaMKII-S6K-UBF axis by either RNAi or pharmaceutical approaches leads to reduction of pre-45s rRNA expression, which subsequently suppresses cell proliferation in colon cancer cells by causing cell-cycle arrest. Knockdown of APC activates CaMKII-S6K-UBF cascade and thus enhances pre-45s rRNA expression. Moreover, the high expression level of pre-45s rRNA is associated with poor survival of CRC patients in two independent cohorts. Our study identifies a novel mechanism in CRC pathogenesis mediated by pre-45s rRNA and a prognostic factor of pre-45s rRNA in CRC patients.

External loads applied to skeletal muscle cause increases in the protein translation rate, which leads to muscle hypertrophy. Although some studies have demonstrated that increases in the capacity and efficiency of translation are involved in this process, it remains unclear how these two factors are related to the magnitude of muscle hypertrophy. The present study aimed to clarify the roles played by the capacity and efficiency of translation in muscle hypertrophy. We used an improved synergist ablation in which the magnitude of compensatory hypertrophy could be controlled by partial removal of synergist muscles. Male rats were assigned to four groups in which the plantaris muscle was unilaterally subjected to weak (WK), moderate (MO), middle (MI), and strong (ST) overloading by four types of synergist ablation. Fourteen days after surgery, the weight of the plantaris muscle per body weight increased by 8%, 22%, 32% and 45%, in the WK, MO, MI and ST groups, respectively. Five days after surgery, 18+28S rRNA content (an indicator of translational capacity) increased with increasing overload, with increases of 1.8-fold (MO), 2.2-fold (MI), and 2.5-fold (ST), respectively, relative to non-overloaded muscle (NL) in the WK group. rRNA content showed a strong correlation with relative muscle weight measured 14 days after surgery (r = 0.98). The phosphorylated form of p70S6K (a positive regulator of translational efficiency) showed a marked increase in the MO group, but no further increase was observed with further increase in overload (increases of 22.6-fold (MO), 17.4-fold (MI), and 18.2-fold (ST), respectively, relative to NL in the WK group). These results indicate that increases in ribosome biogenesis at the early phase of overloading are strongly dependent on the amount of overloading, and may play an important role in increasing the translational capacity for further gain of muscular size.