scholarly journals The E3 ubiquitin ligase ZNRF2 is a substrate of mTORC1 and regulates its activation by amino acids

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Gerta Hoxhaj ◽  
Edward Caddye ◽  
Ayaz Najafov ◽  
Vanessa P Houde ◽  
Catherine Johnson ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Ubiquitin Ligase ◽  
Protein Phosphatase ◽  
E3 Ubiquitin Ligase ◽  
Mechanistic Target Of Rapamycin ◽  
Rag Gtpases ◽  
Intracellular Amino Acid ◽  
Amino Acid Levels ◽  
Amino Acid Sensing

The mechanistic Target of Rapamycin complex 1 (mTORC1) senses intracellular amino acid levels through an intricate machinery, which includes the Rag GTPases, Ragulator and vacuolar ATPase (V-ATPase). The membrane-associated E3 ubiquitin ligase ZNRF2 is released into the cytosol upon its phosphorylation by Akt. In this study, we show that ZNRF2 interacts with mTOR on membranes, promoting the amino acid-stimulated translocation of mTORC1 to lysosomes and its activation in human cells. ZNRF2 also interacts with the V-ATPase and preserves lysosomal acidity. Moreover, knockdown of ZNRF2 decreases cell size and cell proliferation. Upon growth factor and amino acid stimulation, mTORC1 phosphorylates ZNRF2 on Ser145, and this phosphosite is dephosphorylated by protein phosphatase 6. Ser145 phosphorylation stimulates vesicle-to-cytosol translocation of ZNRF2 and forms a novel negative feedback on mTORC1. Our findings uncover ZNRF2 as a component of the amino acid sensing machinery that acts upstream of Rag-GTPases and the V-ATPase to activate mTORC1.

scholarly journals Deletion of RTS1, Encoding a Regulatory Subunit of Protein Phosphatase 2A, Results in Constitutive Amino Acid Signaling via Increased Stp1p Processing

2006 ◽  
Vol 5 (1) ◽  
pp. 174-179 ◽  
Author(s):  
Nadine Eckert-Boulet ◽  
Katrin Larsson ◽  
Boqian Wu ◽  
Peter Poulsen ◽  
Birgitte Regenberg ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Phosphatase ◽  
Protein Phosphatase 2A ◽  
Regulatory Subunit ◽  
Mutant Library ◽  
Amino Acid Permease ◽  
Phosphatase 2A ◽  
Amino Acid Sensing ◽  
Amino Acid Signaling

ABSTRACT In Saccharomyces cerevisiae, extracellular amino acids are sensed at the plasma membrane by the SPS sensor, consisting of the transporter homologue Ssy1p, Ptr3p, and the endoprotease Ssy5p. Amino acid sensing results in proteolytic truncation of the transcription factors Stp1p and Stp2p, followed by their relocation from the cytoplasm to the nucleus, where they activate transcription of amino acid permease genes. We screened a transposon mutant library for constitutively signaling mutants, with the aim of identifying down-regulating components of the SPS-mediated pathway. Three isolated mutants were carrying a transposon in the RTS1 gene, which encodes a regulatory subunit of protein phosphatase 2A. We investigated the basal activity of the AGP1 and BAP2 promoters in rts1Δ cells and found increased transcription from these promoters, as well as increased Stp1p processing, even in the absence of amino acids. Based on our findings we propose that the phosphatase complex containing Rts1p keeps the SPS-mediated pathway down-regulated in the absence of extracellular amino acids by dephosphorylating a component of the pathway.

scholarly journals Amino Acid-Mediated Intracellular Ca2+ Rise Modulates mTORC1 by Regulating the TSC2-Rheb Axis through Ca2+/Calmodulin

2021 ◽  
Vol 22 (13) ◽  
pp. 6897
Author(s):  
Yuna Amemiya ◽  
Nao Nakamura ◽  
Nao Ikeda ◽  
Risa Sugiyama ◽  
Chiaki Ishii ◽  
...  
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Growth Regulator ◽  
Inhibitory Effect ◽  
Environmental Cues ◽  
Gtpase Activating Protein ◽  
Mechanistic Target Of Rapamycin ◽  
Rag Gtpases ◽  
Mtorc1 Activation

Mechanistic target of rapamycin complex 1 (mTORC1) is a master growth regulator by controlling protein synthesis and autophagy in response to environmental cues. Amino acids, especially leucine and arginine, are known to be important activators of mTORC1 and to promote lysosomal translocation of mTORC1, where mTORC1 is thought to make contact with its activator Rheb GTPase. Although amino acids are believed to exclusively regulate lysosomal translocation of mTORC1 by Rag GTPases, how amino acids increase mTORC1 activity besides regulation of mTORC1 subcellular localization remains largely unclear. Here we report that amino acids also converge on regulation of the TSC2-Rheb GTPase axis via Ca2+/calmodulin (CaM). We showed that the amino acid-mediated increase of intracellular Ca2+ is important for mTORC1 activation and thereby contributes to the promotion of nascent protein synthesis. We found that Ca2+/CaM interacted with TSC2 at its GTPase activating protein (GAP) domain and that a CaM inhibitor reduced binding of CaM with TSC2. The inhibitory effect of a CaM inhibitor on mTORC1 activity was prevented by loss of TSC2 or by an active mutant of Rheb GTPase, suggesting that a CaM inhibitor acts through the TSC2-Rheb axis to inhibit mTORC1 activity. Taken together, in response to amino acids, Ca2+/CaM-mediated regulation of the TSC2-Rheb axis contributes to proper mTORC1 activation, in addition to the well-known lysosomal translocation of mTORC1 by Rag GTPases.

scholarly journals Electrospun Microfibers Modulate Intracellular Amino Acids in Liver Cells via Integrin β1

2021 ◽  
Vol 8 (7) ◽  
pp. 88
Author(s):  
Tianjiao Huang ◽  
John A. Terrell ◽  
Jay H. Chung ◽  
Chengpeng Chen
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Liver Cells ◽  
Urea Synthesis ◽  
Integrin Β1 ◽  
Flat Substrate ◽  
Intracellular Amino Acid ◽  
Vital Functions ◽  
Cytochrome P450 Activity ◽  
Amino Acid Levels

Although numerous recent studies have shown the importance of polymeric microfibrous extracellular matrices (ECMs) in maintaining cell behaviors and functions, the mechanistic nexus between ECMs and intracellular activities is largely unknown. Nevertheless, this knowledge will be critical in understanding and treating diseases with ECM remodeling. Therefore, we present our findings that ECM microstructures could regulate intracellular amino acid levels in liver cells mechanistically through integrin β1. Amino acids were studied because they are the fundamental blocks for protein synthesis and metabolism, two vital functions of liver cells. Two ECM conditions, flat and microfibrous, were prepared and studied. In addition to characterizing cell growth, albumin production, urea synthesis, and cytochrome p450 activity, we found that the microfibrous ECM generally upregulated the intracellular amino acid levels. Further explorations showed that cells on the flat substrate expressed more integrin β1 than cells on the microfibers. Moreover, after partially blocking integrin β1 in cells on the flat substrate, the intracellular amino acid levels were restored, strongly supporting integrin β1 as the linking mechanism. This is the first study to report that a non-biological polymer matrix could regulate intracellular amino acid patterns through integrin. The results will help with future therapy development for liver diseases with ECM changes (e.g., fibrosis).

scholarly journals Functional Amino Acids and Autophagy: Diverse Signal Transduction and Application

2021 ◽  
Vol 22 (21) ◽  
pp. 11427
Author(s):  
Chunchen Liu ◽  
Linbao Ji ◽  
Jinhua Hu ◽  
Ying Zhao ◽  
Lee J. Johnston ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Amino Acids ◽  
Amino Acid ◽  
Mammalian Target Of Rapamycin ◽  
Comprehensive Overview ◽  
Related Disease ◽  
Signal Relay ◽  
Amino Acid Levels ◽  
Amino Acid Sensing

Functional amino acids provide great potential for treating autophagy-related diseases by regulating autophagy. The purpose of the autophagy process is to remove unwanted cellular contents and to recycle nutrients, which is controlled by many factors. Disordered autophagy has been reported to be associated with various diseases, such as cancer, neurodegeneration, aging, and obesity. Autophagy cannot be directly controlled and dynamic amino acid levels are sufficient to regulate autophagy. To date, arginine, leucine, glutamine, and methionine are widely reported functional amino acids that regulate autophagy. As a signal relay station, mammalian target of rapamycin complex 1 (mTORC1) turns various amino acid signals into autophagy signaling pathways for functional amino acids. Deficiency or supplementation of functional amino acids can immediately regulate autophagy and is associated with autophagy-related disease. This review summarizes the mechanisms currently involved in autophagy and amino acid sensing, diverse signal transduction among functional amino acids and autophagy, and the therapeutic appeal of amino acids to autophagy-related diseases. We aim to provide a comprehensive overview of the mechanisms of amino acid regulation of autophagy and the role of functional amino acids in clinical autophagy-related diseases and to further convert these mechanisms into feasible therapeutic applications.

Activation mechanisms of the E3 ubiquitin ligase parkin

2017 ◽  
Vol 474 (18) ◽  
pp. 3075-3086 ◽  
Author(s):  
Nikhil Panicker ◽  
Valina L. Dawson ◽  
Ted M. Dawson
Keyword(s):  
Parkinson’S Disease ◽  
Amino Acid ◽  
Ubiquitin Ligase ◽  
Recent Literature ◽  
E3 Ubiquitin Ligase ◽  
Mitochondrial Damage ◽  
Cytosolic Protein ◽  
Mitochondrial Functions ◽  
Activation Mechanisms

Monogenetic, familial forms of Parkinson's disease (PD) only account for 5–10% of the total number of PD cases, but analysis of the genes involved therein is invaluable to understanding PD-associated neurodegenerative signaling. One such gene, parkin, encodes a 465 amino acid E3 ubiquitin ligase. Of late, there has been considerable interest in the role of parkin signaling in PD and in identifying its putative substrates, as well as the elucidation of the mechanisms through which parkin itself is activated. Its dysfunction underlies both inherited and idiopathic PD-associated neurodegeneration. Here, we review recent literature that provides a model of activation of parkin in the setting of mitochondrial damage that involves PINK1 (PTEN-induced kinase-1) and phosphoubiquitin. We note that neuronal parkin is primarily a cytosolic protein (with various non-mitochondrial functions), and discuss potential cytosolic parkin activation mechanisms.

Interrelationship between hepatic ureagenesis and gluconeogenesis in early sepsis

1991 ◽  
Vol 260 (3) ◽  
pp. E453-E458 ◽  
Author(s):  
Y. Ohtake ◽  
M. G. Clemens
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Substrate Preference ◽  
Hormone Sensitivity ◽  
Urea Production ◽  
Urea Release ◽  
Amino Acid Levels ◽  
Early Sepsis ◽  
Septic Group

This study was performed to investigate the interrelationship between gluconeogenesis and ureagenesis during sepsis. In isolated perfused livers, gluconeogenesis was assessed using either lactate or a combination of lactate, glutamine, and alanine as substrate. Ureagenesis was assessed using either NH4Cl or glutamine plus alanine as substrate. NH4Cl stimulated urea production in livers from both septic and sham-operated control rats. Urea release was approximately 1.2 and 2.0 mg urea nitrogen.g-1.h-1 for 1 and 5 mM NH4Cl, respectively, and was equal for both groups. With amino acids as substrate, urea production was significantly greater in livers from septic animals compared with controls. Phenylephrine stimulated urea production in the sham-operated group by about twofold, whereas in the septic group urea release was slightly inhibited. Gluconeogenesis from lactate was inhibited by NH4Cl (1 and 5 mM) in both groups, with no difference between groups. In contrast to enhanced ureagenesis from amino acids in septic rats, gluconeogenesis was decreased by approximately 24% (P less than 0.5). Similarly, phenylephrine (1 microM) stimulated gluconeogenesis by 13 +/- 1 mumol.g-1.h-1 in sham-operated rats but only by 9 +/- 1 mumol.g-1.h-1 in septic rats (P less than 0.02). These results suggest that hepatic gluconeogenic and ureagenic pathways are intact in sepsis but that altered substrate preference and hormone sensitivity may result in decreased gluconeogenesis in the presence of elevated amino acid levels.

scholarly journals Activation of the SPS Amino Acid-Sensing Pathway in Saccharomyces cerevisiae Correlates with the Phosphorylation State of a Sensor Component, Ptr3

2007 ◽  
Vol 28 (2) ◽  
pp. 551-563 ◽  
Author(s):  
Zhengchang Liu ◽  
Janet Thornton ◽  
Mário Spírek ◽  
Ronald A. Butow
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Amino Acid ◽  
Nuclear Translocation ◽  
Proteolytic Processing ◽  
Casein Kinase I ◽  
Positive Regulators ◽  
Amino Acid Permeases ◽  
Amino Acid Sensing ◽  
Amino Acid Sensor

ABSTRACT Cells of the budding yeast Saccharomyces cerevisiae sense extracellular amino acids and activate expression of amino acid permeases through the SPS-sensing pathway, which consists of Ssy1, an amino acid sensor on the plasma membrane, and two downstream factors, Ptr3 and Ssy5. Upon activation of SPS signaling, two transcription factors, Stp1 and Stp2, undergo Ssy5-dependent proteolytic processing that enables their nuclear translocation. Here we show that Ptr3 is a phosphoprotein whose hyperphosphorylation is increased by external amino acids and is dependent on Ssy1 but not on Ssy5. A deletion mutation in GRR1, encoding a component of the SCFGrr1 E3 ubiquitin ligase, blocks amino acid-induced hyperphosphorylation of Ptr3. We found that two casein kinase I (CKI) proteins, Yck1 and Yck2, previously identified as positive regulators of SPS signaling, are required for hyperphosphorylation of Ptr3. Loss- and gain-of-function mutations in PTR3 result in decreased and increased Ptr3 hyperphosporylation, respectively. We found that a defect in PP2A phosphatase activity leads to the hyperphosphorylation of Ptr3 and constitutive activation of SPS signaling. Two-hybrid analysis revealed interactions between the N-terminal signal transduction domain of Ssy1 with Ptr3 and Yck1. Our findings reveal that CKI and PP2A phosphatase play antagonistic roles in SPS sensing by regulating Ptr3 phosphorylation.

scholarly journals Amino acids whose intracellular levels change most during aging alter chronological lifespan of fission yeast

2020 ◽  
Author(s):  
Charalampos Rallis ◽  
Michael Mülleder ◽  
Graeme Smith ◽  
Yan Zi Au ◽  
Markus Ralser ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Fission Yeast ◽  
Yeast Cells ◽  
Total Amino Acid ◽  
Wild Type ◽  
Chronological Lifespan ◽  
Biomarkers Of Aging ◽  
Concentration Changes ◽  
Amino Acid Levels

AbstractAmino acid deprivation or supplementation can affect cellular and organismal lifespan, but we know little about the role of concentration changes in free, intracellular amino acids during aging. Here, we determine free amino-acid levels during chronological aging of non-dividing fission yeast cells. We compare wild-type with long-lived mutant cells that lack the Pka1 protein of the protein kinase A signalling pathway. In wild-type cells, total amino-acid levels decrease during aging, but much less so in pka1 mutants. Two amino acids strongly change as a function of age: glutamine decreases, especially in wild-type cells, while aspartate increases, especially in pka1 mutants. Supplementation of glutamine is sufficient to extend the chronological lifespan of wild-type but not of pka1Δ cells. Supplementation of aspartate, on the other hand, shortens the lifespan of pka1Δ but not of wild-type cells. Our results raise the possibility that certain amino acids are biomarkers of aging, and their concentrations during aging can promote or limit cellular lifespan.

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