LARP1 facilitates translational recovery after amino acid refeeding by preserving long poly(A)-tailed TOP mRNAs

ABSTRACTOccasionally, cells must adapt to an inimical growth conditions like amino acid starvation (AAS) by downregulating protein synthesis. A class of transcripts containing 5’terminal oligopyrimidine (5’TOP) motif encodes translation-related proteins such as ribosomal proteins (RPs) and elongation factors, and therefore, their translation is severely repressed during AAS to conserve energy1. The RNA-binding protein LARP1 transduces amino acid signaling to TOP gene expression by controlling translation and stability of TOP mRNAs2-6. When released from AAS, translation machineries in turn have to be restored, however, the underlying mechanism of such re-adaptation is largely unknown. Here we show that LARP1 preserves TOP mRNAs in a long polyadenylated state during long-term AAS. We found that TOP mRNAs become highly polyadenylated when cells are in AAS or treated with the mTOR (mechanistic target of rapamycin) inhibitor Torin1. Importantly, depletion of LARP1 completely abrogated the polyadenylation of TOP mRNAs. Comprehensive analysis of poly(A) tail length using the Nanopore direct RNA sequencing revealed that TOP mRNAs are selectively polyadenylated under mTOR inhibition. Since a long poly(A) tail confers increased stability and polysome formation of TOP mRNAs, we predict that LARP1-dependent preservation of TOP mRNAs enables rapid translational resumption after the release from AAS.

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p27 regulates the autophagy-lysosomal pathway via the control of Ragulator and mTOR activity in amino acid deprived cells

10.1101/2020.01.07.896860 ◽

2020 ◽

Author(s):

Ada Nowosad ◽

Pauline Jeannot ◽

Caroline Callot ◽

Justine Creff ◽

Renaud T. Perchey ◽

...

Keyword(s):

Amino Acid ◽

Energy Homeostasis ◽

Underlying Mechanism ◽

Mtor Inhibition ◽

Mtorc1 Signaling ◽

And Function ◽

Mtorc1 Activation ◽

Catabolic Process ◽

Mtor Activity ◽

Dependent Mechanism

SummaryAutophagy is a catabolic process whereby cytoplasmic components are degraded within lysosomes, allowing cells to maintain energy homeostasis during nutrient depletion. Several studies have shown that the CDK inhibitor p27Kip1 promotes starvation-induced autophagy. However, the underlying mechanism remains unknown. Here, we report that in amino acid deprived cells, p27 controls autophagy via an mTORC1-dependent mechanism. During prolonged amino acid starvation, a fraction of p27 is recruited to lysosomes where it interacts with LAMTOR1, a component of the Ragulator complex required for mTORC1 lysosomal localization and activation. p27 binding to LAMTOR1 prevents Ragulator assembly and function and subsequent mTORC1 activation, thereby promoting autophagy. Conversely, upon amino acid withdrawal, p27−/− cells exhibit elevated mTORC1 signaling, impaired lysosomal activity and autophagy, and resistance to apoptosis. This is associated with sequestration of TFEB in the cytoplasm, preventing the induction of lysosomal genes required for lysosomal function. Silencing of LAMTOR1 or mTOR inhibition restores autophagy and induces apoptosis in p27−/− cells. Together, these results reveal a direct, coordinated regulation between the cell cycle and cell growth machineries.

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Astragaloside IV Alleviates Tacrolimus-Induced Chronic Nephrotoxicity via p62-Keap1-Nrf2 Pathway

Frontiers in Pharmacology ◽

10.3389/fphar.2020.610102 ◽

2021 ◽

Vol 11 ◽

Author(s):

Ping Gao ◽

Xiaoyi Du ◽

Lili Liu ◽

Hua Xu ◽

Maochang Liu ◽

...

Keyword(s):

Oxidative Stress ◽

Renal Function ◽

Nuclear Translocation ◽

Underlying Mechanism ◽

Protective Effects ◽

Astragaloside Iv ◽

Nrf2 Pathway ◽

Ros Accumulation ◽

Related Proteins

Tacrolimus-induced chronic nephrotoxicity (TIN) hinders its long-term use in patients. However, there are no drugs available in the clinic to relieve it at present. Astragaloside IV (AS-IV) is a saponin extract of the Astragalus which is widely used in the treatment of kidney disease. This study aimed to investigate the effect of AS-IV on TIN and its underlying mechanism. Herein, C57BL/6 mice were treated with tacrolimus and/or AS-IV for 4 weeks, and then the renal function, fibrosis, oxidative stress and p62-Keap1-Nrf2 pathway were evaluated to ascertain the contribution of AS-IV and p62-Keap1-Nrf2 pathway to TIN. Our results demonstrated that AS-IV significantly improved renal function and alleviated tubulointerstitial fibrosis compared with the model group. The expression of fibrosis-related proteins, including TGF-β1, Collagen I and α-SMA, were also decreased by AS-IV. Furthermore, AS-IV relieved the inhibition of tacrolimus on antioxidant enzymes. The data in HK-2 cells also proved that AS-IV reduced tacrolimus-induced cell death and oxidative stress. Mechanistically, AS-IV markedly promoted the nuclear translocation of Nrf2 and the renal protective effects of AS-IV were abolished by Nrf2 inhibitor. Further researches showed that phosphorylated p62 was significantly increased after AS-IV pretreatment. Moreover, AS-IV failed to increase nuclear translocation of Nrf2 and subsequent anti-oxidative stress in HK-2 cells transfected with p62 siRNA. Collectively, these findings indicate that AS-IV relieve TIN by enhancing p62 phosphorylation, thereby increasing Nrf2 nuclear translocation, and then alleviating ROS accumulation and renal fibrosis.

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Sepsis-Associated Encephalopathy: Insight into Injury and Pathogenesis

CNS & Neurological Disorders - Drug Targets ◽

10.2174/1871527319999201117122158 ◽

2020 ◽

Vol 19 ◽

Author(s):

Lina Zhao ◽

Yanxia Gao ◽

Shigong Guo ◽

Xin Lu ◽

Shiyuan Yu ◽

...

Keyword(s):

Amino Acid ◽

Mitochondrial Dysfunction ◽

Blood Brain Barrier ◽

Intestinal Flora ◽

Underlying Mechanism ◽

Brain Barrier ◽

Balance Disorders ◽

Amino Acid Balance ◽

Insight Into

: Sepsis-associated encephalopathy causes long-term health problems in patients with sepsis. This review ex-plores the pathogenesis of sepsis-associated encephalopathy, including its effects on the blood-brain barrier, microglia ac-tivation, mitochondrial dysfunction, the inflammatory medium and neurotransmitters and its roles in amino acid balance disorders, hyperammonemia, and intestinal flora imbalance. Understanding the etiology of sepsis-associated encephalopa-thy may allow the development of adjunctive therapies targeting its underlying mechanism and help develop preventative strategies.

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The mTOR regulated RNA-binding protein LARP1 requires PABPC1 for guided mRNA interaction

Nucleic Acids Research ◽

10.1093/nar/gkaa1189 ◽

2020 ◽

Author(s):

Ewan M Smith ◽

Nour El Houda Benbahouche ◽

Katherine Morris ◽

Ania Wilczynska ◽

Sarah Gillen ◽

...

Keyword(s):

Cell Growth ◽

Rna Binding ◽

Binding Protein ◽

Mammalian Target Of Rapamycin ◽

Rna Binding Protein ◽

Mtor Inhibition ◽

Growth And Survival ◽

Mrna Targets ◽

Terminal Oligopyrimidine ◽

Rna Interaction

Abstract The mammalian target of rapamycin (mTOR) is a critical regulator of cell growth, integrating multiple signalling cues and pathways. Key among the downstream activities of mTOR is the control of the protein synthesis machinery. This is achieved, in part, via the co-ordinated regulation of mRNAs that contain a terminal oligopyrimidine tract (TOP) at their 5′ends, although the mechanisms by which this occurs downstream of mTOR signalling are still unclear. We used RNA-binding protein (RBP) capture to identify changes in the protein-RNA interaction landscape following mTOR inhibition. Upon mTOR inhibition, the binding of LARP1 to a number of mRNAs, including TOP-containing mRNAs, increased. Importantly, non-TOP-containing mRNAs bound by LARP1 are in a translationally-repressed state, even under control conditions. The mRNA interactome of the LARP1-associated protein PABPC1 was found to have a high degree of overlap with that of LARP1 and our data show that PABPC1 is required for the association of LARP1 with its specific mRNA targets. Finally, we demonstrate that mRNAs, including those encoding proteins critical for cell growth and survival, are translationally repressed when bound by both LARP1 and PABPC1.

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FoxO1 inhibits autophagosome-lysosome fusion leading to endothelial autophagic-apoptosis in diabetes

Cardiovascular Research ◽

10.1093/cvr/cvz014 ◽

2019 ◽

Vol 115 (14) ◽

pp. 2008-2020 ◽

Cited By ~ 4

Author(s):

Hui Zhang ◽

Song Ge ◽

Kesuai He ◽

Xin Zhao ◽

Ya Wu ◽

...

Keyword(s):

Endothelial Cells ◽

Underlying Mechanism ◽

Diabetic Patients ◽

Aortic Endothelial Cells ◽

Glycation End Products ◽

Induced Apoptosis ◽

Related Proteins ◽

Aortic Intima ◽

Human Aortic Endothelial Cells

Abstract Aims Inadequate autophagy contributed to endothelial dysfunction in diabetic patients. We aimed to investigate the relationship between inadequate autophagy and endothelial cells (ECs) apoptosis in diabetes and its underlying mechanism. Methods and results Aortic intima and ECs were isolated from diabetic patients. Cultured human aortic endothelial cells (HAECs) were stimulated with advanced glycation end products (AGEs). The expression of autophagy and apoptosis-related proteins were determined by western blotting. Autophagosomes were observed by electron microscopy. The fusion of autophagosome and lysosomes was detected by immunofluorescence. Compared with non-diabetic subjects, the levels of LC3-II, p62, FoxO1, and Ac-FoxO1 were increased in ECs from diabetic patients, accompanied by the decreased expressions of Atg14, STX17, and co-localization of LC3-II/LAMP2 and Atg14/STX17. Long-term stimulation with AGEs up-regulated LC3-II and p62 expression and the number of autophagosomes with decreased level of Atg14, STX17, Ras-related protein 7 (Rab7), and co-localization of LC3-II/LAMP2 and Atg14/STX17 in HAECs. The apoptosis rates were increased with elevated cleaved-caspase-3 and declined Bcl-2 expression. Inhibition of autophagy with 3-methyladenine could reduce long-term AGEs-induced apoptosis. Higher levels of FoxO1, Ac-FoxO1, and Ac-FoxO1 binding to Atg7 were detected in AGEs-treated HAECs. AGEs-induced FoxO1 enhanced Akt activity, decreased SIRT1-deacetylase activity by phosphorylation and elevated Ac-FoxO1. Knockout of FoxO1 reduced AGEs-induced autophagy and promoted the expression of Atg14 and the co-localization of LC3-II/LAMP 2 and Atg14/STX17. Conclusion Inadequate autophagy with impaired autophagosome-lysosomal fusion exists in aortic intima and ECs from diabetic patients. FoxO1 mediates AGEs-induced ECs autophagic apoptosis through impairing autophagosome-lysosomes fusion by inhibiting Atg14 expression.

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SIRT1 attenuates sepsis-induced acute kidney injury via Beclin1 deacetylation-mediated autophagy activation

Cell Death and Disease ◽

10.1038/s41419-021-03508-y ◽

2021 ◽

Vol 12 (2) ◽

Author(s):

Zhiya Deng ◽

Maomao Sun ◽

Jie Wu ◽

Haihong Fang ◽

Shumin Cai ◽

...

Keyword(s):

Acute Kidney Injury ◽

Protective Effect ◽

Cell Model ◽

Kidney Injury ◽

Underlying Mechanism ◽

Autophagy Induction ◽

Promising Therapeutic Approach ◽

Related Proteins ◽

Caecal Ligation And Puncture

AbstractOur previous studies showed that silent mating-type information regulation 2 homologue-1 (SIRT1, a deacetylase) upregulation could attenuate sepsis-induced acute kidney injury (SAKI). Upregulated SIRT1 can deacetylate certain autophagy-related proteins (Beclin1, Atg5, Atg7 and LC3) in vitro. However, it remains unclear whether the beneficial effect of SIRT1 is related to autophagy induction and the underlying mechanism of this effect is also unknown. In the present study, caecal ligation and puncture (CLP)-induced mice, and an LPS-challenged HK-2 cell line were established to mimic a SAKI animal model and a SAKI cell model, respectively. Our results demonstrated that SIRT1 activation promoted autophagy and attenuated SAKI. SIRT1 deacetylated only Beclin1 but not the other autophagy-related proteins in SAKI. SIRT1-induced autophagy and its protective effect against SAKI were mediated by the deacetylation of Beclin1 at K430 and K437. Moreover, two SIRT1 activators, resveratrol and polydatin, attenuated SAKI in CLP-induced septic mice. Our study was the first to demonstrate the important role of SIRT1-induced Beclin1 deacetylation in autophagy and its protective effect against SAKI. These findings suggest that pharmacologic induction of autophagy via SIRT1-mediated Beclin1 deacetylation may be a promising therapeutic approach for future SAKI treatment.

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Methionine and Arginine Supply Alters Abundance of Amino Acid, Insulin Signaling, and Glutathione Metabolism-Related Proteins in Bovine Subcutaneous Adipose Explants Challenged with N-Acetyl-d-sphingosine

Animals ◽

10.3390/ani11072114 ◽

2021 ◽

Vol 11 (7) ◽

pp. 2114

Author(s):

Yusheng Liang ◽

Nana Ma ◽

Danielle N. Coleman ◽

Fang Liu ◽

Yu Li ◽

...

Keyword(s):

Adipose Tissue ◽

Amino Acid ◽

Insulin Signaling ◽

Elongation Factor ◽

In Vivo Studies ◽

Glutathione Metabolism ◽

Protein Abundance ◽

Related Proteins ◽

Subcutaneous Adipose

The objective was to perform a proof-of-principle study to evaluate the effects of methionine (Met) and arginine (Arg) supply on protein abundance of amino acid, insulin signaling, and glutathione metabolism-related proteins in subcutaneous adipose tissue (SAT) explants under ceramide (Ce) challenge. SAT from four lactating Holstein cows was incubated with one of the following media: ideal profile of amino acid as the control (IPAA; Lys:Met 2.9:1, Lys:Arg 2:1), increased Met (incMet; Lys:Met 2.5:1), increased Arg (incArg; Lys:Arg 1:1), or incMet plus incArg (Lys:Met 2.5:1 Lys:Arg 1:1) with or without 100 μM exogenous cell-permeable Ce (N-Acetyl-d-sphingosine). Ceramide stimulation downregulated the overall abundance of phosphorylated (p) protein kinase B (AKT), p-mechanistic target of rapamycin (mTOR), and p-eukaryotic elongation factor 2 (eEF2). Without Ce stimulation, increased Met, Arg, or Met + Arg resulted in lower p-mTOR. Compared with control SAT stimulated with Ce, increased Met, Arg, or Met + Arg resulted in greater activation of mTOR (p-mTOR/total mTOR) and AKT (p-AKT/total AKT), with a more pronounced response due to Arg. The greatest protein abundance of glutathione S-transferase Mu 1 (GSTM1) was detected in response to increased Met supply during Ce stimulation. Ceramide stimulation decreased the overall protein abundance of the Na-coupled neutral amino acid transporter SLC38A1 and branched-chain alpha-ketoacid dehydrogenase kinase (BCKDK). However, compared with controls, increased Met or Arg supply attenuated the downregulation of BCKDK induced by Ce. Circulating ceramides might affect amino acid, insulin signaling, and glutathione metabolism in dairy cow adipose tissue. Further in vivo studies are needed to confirm the role of rumen-protected amino acids in regulating bovine adipose function.

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Putative imbalanced amino acid metabolism in rainbow trout long term fed a plant-based diet as revealed by 1H-NMR metabolomics

Journal of Nutritional Science ◽

10.1017/jns.2021.3 ◽

2021 ◽

Vol 10 ◽

Author(s):

Catherine Deborde ◽

Blandine Madji Hounoum ◽

Annick Moing ◽

Mickaël Maucourt ◽

Daniel Jacob ◽

...

Keyword(s):

Amino Acids ◽

Rainbow Trout ◽

Amino Acid ◽

Amino Acid Content ◽

Proton Nuclear Magnetic Resonance ◽

Free Form ◽

Metabolomic Analysis ◽

Nmr Metabolomics ◽

H Nmr

Abstract The long-term effect of a plant (P)-based diet was assessed by proton nuclear magnetic resonance (1H-NMR) metabolomics in rainbow trout fed a marine fish meal (FM)–fish oil (FO) diet (M), a P-based diet and a control commercial-like diet (C) starting with the first feeding. Growth performances were not heavily altered by long-term feeding on the P-based diet. An 1H-NMR metabolomic analysis of the feed revealed significantly different soluble chemical compound profiles between the diets. A set of soluble chemical compounds was found to be specific either to the P-based diet or to the M diet. Pterin, a biomarker of plant feedstuffs, was identified both in the P-based diet and in the plasma of fish fed the P-based diet. 1H-NMR metabolomic analysis on fish plasma and liver and muscle tissues at 6 and 48 h post feeding revealed significantly different profiles between the P-based diet and the M diet, while the C diet showed intermediate results. A higher amino acid content was found in the plasma of fish fed the P-based diet compared with the M diet after 48 h, suggesting either a delayed delivery of the amino acids or a lower amino acid utilisation in the P-based diet. This was associated with an accumulation of essential amino acids and the depletion of glutamine in the muscle, together with an accumulation of choline in the liver. Combined with an anticipated absorption of methionine and lysine supplemented in free form, the present results suggest an imbalanced essential amino acid supply for protein metabolism in the muscle and for specific functions of the liver.

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Upregulation of 5′-terminal oligopyrimidine mRNA translation upon loss of the ARF tumor suppressor

Scientific Reports ◽

10.1038/s41598-020-79379-8 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Kyle A. Cottrell ◽

Ryan C. Chiou ◽

Jason D. Weber

Keyword(s):

Protein Synthesis ◽

Tumor Cells ◽

Ribosomal Proteins ◽

Human Cancer ◽

Mrna Translation ◽

Ribosome Profiling ◽

Translational Efficiency ◽

Gene Encoding ◽

Terminal Oligopyrimidine ◽

Ribosome Export

AbstractTumor cells require nominal increases in protein synthesis in order to maintain high proliferation rates. As such, tumor cells must acquire enhanced ribosome production. How the numerous mutations in tumor cells ultimately achieve this aberrant production is largely unknown. The gene encoding ARF is the most commonly deleted gene in human cancer. ARF plays a significant role in regulating ribosomal RNA synthesis and processing, ribosome export into the cytoplasm, and global protein synthesis. Utilizing ribosome profiling, we show that ARF is a major suppressor of 5′-terminal oligopyrimidine mRNA translation. Genes with increased translational efficiency following loss of ARF include many ribosomal proteins and translation factors. Knockout of p53 largely phenocopies ARF loss, with increased protein synthesis and expression of 5′-TOP encoded proteins. The 5′-TOP regulators eIF4G1 and LARP1 are upregulated in Arf- and p53-null cells.

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Global versus Local Regulatory Roles for Lrp-Related Proteins: Haemophilus influenzae as a Case Study

Journal of Bacteriology ◽

10.1128/jb.183.13.4004-4011.2001 ◽

2001 ◽

Vol 183 (13) ◽

pp. 4004-4011 ◽

Cited By ~ 20

Author(s):

Devorah Friedberg ◽

Michael Midkiff ◽

Joseph M. Calvo

Keyword(s):

Amino Acid ◽

Haemophilus Influenzae ◽

Regulatory Protein ◽

Enteric Bacteria ◽

Regulatory Role ◽

Regulatory Function ◽

Ecological Niches ◽

E Coli ◽

Sequence Identity ◽

Related Proteins

ABSTRACT Lrp (leucine-responsive regulatory protein) plays a global regulatory role in Escherichia coli, affecting expression of dozens of operons. Numerous lrp-related genes have been identified in different bacteria and archaea, includingasnC, an E. coli gene that was the first reported member of this family. Pairwise comparisons of amino acid sequences of the corresponding proteins shows an average sequence identity of only 29% for the vast majority of comparisons. By contrast, Lrp-related proteins from enteric bacteria show more than 97% amino acid identity. Is the global regulatory role associated withE. coli Lrp limited to enteric bacteria? To probe this question we investigated LrfB, an Lrp-related protein fromHaemophilus influenzae that shares 75% sequence identity with E. coli Lrp (highest sequence identity among 42 sequences compared). A strain of H. influenzae having anlrfB null allele grew at the wild-type growth rate but with a filamentous morphology. A comparison of two-dimensional (2D) electrophoretic patterns of proteins from parent and mutant strains showed only two differences (comparable studies withlrp + and lrp E. coli strains by others showed 20 differences). The abundance of LrfB in H. influenzae, estimated by Western blotting experiments, was about 130 dimers per cell (compared to 3,000 dimers per E. colicell). LrfB expressed in E. coli replaced Lrp as a repressor of the lrp gene but acted only to a limited extent as an activator of the ilvIH operon. Thus, although LrfB resembles Lrp sufficiently to perform some of its functions, its low abundance is consonant with a more local role in regulating but a few genes, a view consistent with the results of the 2D electrophoretic analysis. We speculate that an Lrp having a global regulatory role evolved to help enteric bacteria adapt to their ecological niches and that it is unlikely that Lrp-related proteins in other organisms have a broad regulatory function.

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