Amino acids regulate hepatic intermediary metabolism-related gene expression via mTORC1 ...

234 Pages·2015·9.28 MB·English

N ° d (cid:495) o r d r e :

THÈSE

Présentée a

L’UNIVERSITÉ DE PAU ET DES PAYS DE L’ADOUR
ÉCOLE DOCTORALE 211 - SCIENCES EXACTES ET LEURS APPLICATIONS

Weiwei DAI
par

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DOCTEUR

SPÉCIALITÉ : Sciences agronomiques, biotechnologies agro-alimentaires

Amino acids regulate hepatic intermediary metabolism-related gene
expression via mTORC1-dependent manner in rainbow trout
(Oncorhynchus mykiss)
************
Les acides aminés régulent l'expression des gènes du métabolisme
intermédiaire chez la truite par le biais de mTORC1
(Oncorhynchus mykiss)

Soutenue le: 27 October, 2015

Devant la commission d’examen formée de:

Mme. BONNIEU, Anne, Directrice de Recherche, INRA Rapporteur
M. MAZURAIS, David, Ingénieur de Recherche HDR, Ifremer Rapporteur
M. DAVAIL, Stéphane, Professeur, President
M. DUCHAMP, Claude, Professeur, Université de Lyon Examinatrice
Université de Pau et des Pays de l(cid:495)Adour
Mme. SKIBA-CASSY, Sandrine, Directrice de recherche, INRA Directeur de thèse

I dedicate this thesis to INRA, NuMeA—where I grew and made progresses

-----------------------Weiwei DAI

Table of Contents
LIST OF ABREVIATIONS ....................................................................................................................................... III
LIST OF FIGURES .................................................................................................................................................. VIII
LIST OF TABLES ........................................................................................................................................................ X
Chapter 1 INTRODUCTION AND REVIEW OF LITERATURE ....................................................................... 1
1.1 Aquaculture development: current status, trends and future prospects ................................................ 3
............................................... 3
1.1.2 Towards sustainable aquaculture ................................................................................................................... 4
(cid:883).(cid:883).(cid:883) Aquaculture(cid:495)s role in the global good system, trends and prospects
1.1.3 The importance of protein/amino acids research for aquaculture .................................................. 6
1.2 Cellular signaling ............................................................................................................................................................. 8
1.2.1 Insulin signaling in mammals ............................................................................................................................ 8
1.2.1.1 Overview of insulin signaling pathway and some critical nodes ............................................... 8
1.2.1.2 Insulin regulates glucose metabolism ................................................................................................ 11
1.2.1.3 Insulin stimulates lipid synthesis but inhibits lipid degradation ........................................... 13
1.2.1.4 Insulin signaling regulates mitochondrial metabolism .............................................................. 15
1.2.1.5 Impaired insulin action and insulin resistance .............................................................................. 16
1.2.2 AA signaling pathways: mTOR & AAR pathway in mammals ........................................................... 18
1.2.2.1 Overview of mTOR signaling pathway in mammals .................................................................... 18
1.2.2.2 mTORC1 signaling pathway in mammals ......................................................................................... 20
1.2.2.3 mTORC2 signaling pathway in mammals ......................................................................................... 30
1.2.2.4 AAR pathway in mammals ...................................................................................................................... 34
1.2.3 AAs & insulin secretion, insulin action and insulin resistance in mammals .............................. 35
1.2.4 AAs and insulin/mTOR signaling pathways in fish ............................................................................... 39
1.3 The regulation of hepatic intermediary metabolism in mammals and fish ........................................ 42
1.3.1 The regulation of hepatic glucose metabolism ....................................................................................... 42
1.3.1.1 Hepatocyte glucose uptake and phosphorylation ......................................................................... 44
1.3.1.2 Glycolysis and glycogen synthesis ....................................................................................................... 46
1.3.1.3 Glycogenolysis and gluconeogenesis .................................................................................................. 48
1.3.2 The regulation of hepatic lipid metabolism in mammals and fish ................................................. 50
1.3.2.1 Hepatic lipogenesis .................................................................................................................................... 50
1.3.2.2 FA catabolism/β-Oxidation ..................................................................................................................... 54
1.3.3 The regulation of hepatic amino acid catabolism in mammals and fish ...................................... 56
1.4 Rainbow trout as a model for nutrition and nutrient metabolism studies ......................................... 58
1.5 Hypothesis and objectives of the thesis ............................................................................................................. 60
Chapter 2 MATERIAL AND METHODS ............................................................................................................ 63
2.1 Experimental trials ...................................................................................................................................................... 65
2.1.1 Ethics statement ................................................................................................................................................... 65
I
2.1.2 In vivo experimental trial ................................................................................................................................. 65
2.1.2.1 Experimental and sampling procedures ........................................................................................... 65
2.1.2.2 Sampling procedure ................................................................................................................................... 67
2.1.3 In vitro experiments ........................................................................................................................................... 67
2.1.3.1 Animals ............................................................................................................................................................ 67
2.1.3.2 Primary cell culture of rainbow trout hepatocytes ...................................................................... 67
2.1.3.3 Stimulations and cell recovery .............................................................................................................. 69
2.2 Analytical methods ...................................................................................................................................................... 71
2.2.1 Plasma metabolites analysis ........................................................................................................................... 71
2.2.2 Western blot analysis......................................................................................................................................... 71
2.2.3 Gene expression analysis: real time RT-PCR ........................................................................................... 74
2.2.4 Enzyme activity analysis .................................................................................................................................. 76
2.2.5 Statistical analysis ............................................................................................................................................... 76
Chapter 3 RESULTS ............................................................................................................................................... 77
Chapter 4 GENERAL DISCUSSION ................................................................................................................... 155
4.1 mTOR activation, overactivation and inhibition ...........................................................................................157
4.2 mTORC1 activation regulates hepatic intermediary metabolism .........................................................161
4.3 Nutritional regulation of hepatic fatty acid biosynthesis in rainbow trout ......................................165
4.4 The regulation of hepatic gluconeogenesis .....................................................................................................170
4.5 The regulation of AA catabolism..........................................................................................................................173
Chapter 5 CONCLUSIONS AND PERSPECTIVES .......................................................................................... 177
BIBLIOGRAPHY .................................................................................................................................................... 189
ACKOWLEDGEMENT ........................................................................................................................................... 207
LIST OF PUBLICATIONS ..................................................................................................................................... 209

II
LIST OF ABREVIATIONS

Abbreviations Full name

9/SCD1 Stearoyl-CoA desaturase-1
Δ(cid:888)/D(cid:888)D Δ(cid:888) fatty acyl desaturase
4EBP1 eukaryotic initiation factor 4E (eIF4E)-binding protein 1
△
6PF1K 6-phosphofructo-1-kinase

AACE amino acid catabolizing enzymes
AAR amino acid response
AAs amino acids
ACC Acetyl-CoA carboxylase
ACLY ATP citrate lyase
AGC protein kinases A, G, and C
ALAT alanine transaminase
AMPK AMP-activated protein kinase
AS160 Akt substrate of 160 kDa
ASAT aspartate transaminase
ASNS asparagine synthetase
ATF activating transcription factor
Atg 4b autophagy-related protein 4B
Atg12 autophagy-related protein 12

BCKD branched- -ketoacid dehydrogenase
BCKDK branched- -ketoacid dehydrogenase kinase
chain (cid:573)
bHLH-LZ basic helix-loop-helix leucine zipper
chain (cid:573)
BNIP3 BCL2/adenovirus E1B 19 kDa protein-interacting protein 3
–
BSA bovine serum albumin
cAMP cyclic adenosine monophosphate
CARE (C/EBP-ATF) response elements
CAT cationic amino acid transporter 1
CBP CREB-binding protein
CHO carbohydrate
CHOP C/EBP homology protein
ChREBP carbohydrate responsive element-binding protein
CPT 1a carnitine palmitoyltransferase 1 alpha
CPT 1b carnitine palmitoyltransferase 1 beta
CREB cAMP response element-binding protein
CREBH ER-bound transcription factor
CRTC2 CREB regulated transcription coactivator
CRTCs CREB regulated transcription coactivators
DGAT diacylglycerol acyltransferase
III
DHA docosahexaenoic acid
DNL de novo lipogenesis
EAAs essential amino acids
eEF2K eukaryotic elongation factor 2 kinase
EF1 Eukaryotic translation elongation factor 1 alpha 1
EGF epidermal growth factor

Elovl2 elongation of very long chain fatty acids like-2
e)F(cid:884)(cid:573) translation initiation factor (cid:884)(cid:573)
ElovL5 elongation of very long chain fatty acids like-5
EPA eicosapentaenoic acid
ER endoplasmic reticulum
ERK extracellular signal-regulated kinase
estrogen-
ERR(cid:573) related receptor (cid:573)
FA fatty acids
FAD fatty acid desaturases
FAS or FASN fatty acid synthase
FBPase fructose-1,6-bisphosphatase
FKBP12 12 kDa FK506-binding protein
FKHR forkhead related protein
FM fish meal
FO fish oil
FOXO Forkhead box protein O
G6P glucose 6-phosphate
G6Pase I glucose-6-phosphatase I
G6Pase II glucose-6-phosphatase II
G6PDH glucose-6-phosphate dehydrogenase
Gab1 Grb2-associated binder-1
Gabarapl1 gamma-aminobutyric acid
GAP GTPase-activating protein
GCKR glucokinase regulatory protein
GCN2 general control non-derepressible 2
GEF guanine nucleotide exchange factor
GH growth hormone
GK or GCK glucokinase
GLS glutaminase
Glu glucose
GLUT glucose transporter
GP glycogen phosphorylase
GPCR G protein-coupled receptor
GRB10 growth factor receptor-bound protein 10
GRB2 growth factor receptor-bound protein 2
GS glycogen synthase
GSK3 glycogen synthase kinase 3
IV
GTT glucose tolerance test
HAA high level of amino acids
HBP hexosamine biosynthetic pathway
HERP homocysteine-induced ER protein
HIF- hypoxia-inducible factor 1-alpha
HK hexokinase
(cid:883)(cid:573)
hepatic nuclear factor-4-alpha
HNF6 hepatic nuclear factor 6
(NF(cid:886)(cid:573)
HOAD 3-hydroxyacyl-CoA dehydrogenase
HPHC high protein high carbohydrate
HPLC high protein low carbohydrate
IGF1 insulin-growth factor-1
IKK IkappaB kinase
IP intraperitoneal
IR insulin receptor
IRS insulin receptor substrates
JNK c-Jun-N-terminal kinase
LAA low level of amino acids
LATS large tumour suppressor homologue
LC3B light chain 3B
LCFA long-chain fatty acids
LiRiKO liver-specific rictor knockout
LPHC low protein high carbohydrate
LXR liver X receptor

MAP mitogen-activated protein
MCD Malonyl-CoA decarboxylase
ME malic enzyme
mLST8 mammalian lethal with SEC thirteen 8
mPEPCK metochonial phosphoenolpyruvate carboxykinase
MST1 mammalian sterile 20 like kinase-1
mTOR mechanistic target of rapamycin
–
mTORC1 mechanistic target of rapamycin complex 1
mTORC2 mechanistic target of rapamycin complex 2
MUFA monounsaturated fatty acid
NAFLD nonalcoholic fatty liver disease
O-GlcNAc O- -N-acetylglucosamine
OGT O-GlcNAc transferase
linked (cid:574)
p38MAPK mitogen-activated protein kinase p38
V
PBS phosphate buffered saline
PDCD4 programmed cell death 4
PDGF platelet-derived growth factor
PDK1 phosphoinositide dependent protein kinase 1
PEPCK tot phosphoenolpyruvate carboxykinase total
PGC- peroxisome proliferator-
PH pleckstrin-homology
(cid:883)(cid:573) activated receptor γ coactivator (cid:883)(cid:573)
PI3K phosphoinositide 3-kinase
PIP phosphatidylinositol-3,4,5-trisphosphate
3
PK pyruvate kinase
PKA protein kinase A
PKB or Akt protein kinase B
PKC protein kinase Ctheta
PKC- protein kinase C-
PML promyelocytic leukemia
(cid:573) (cid:573)
Pol 1 Polymerase I
PP2A protein phosphatase 2A
PPAR peroxisome proliferator-activated receptor
PPP pentose phosphate pathway
PRAS40 phosphorylation of proline-rich Akt substrate of 40 kDa
PRTE pyrimidine-rich translational element
PTB phosphotyrosine-binding
PtdIns3P phosphatidylinositol-3-phosphate
p- -oxidation
(cid:574)Ox peroxisomal (cid:574)
RAG RAS-related GTP-binding protein
Rapa rapamycin
RAPTOR regulatory-associated protein of mTOR
REDD1 regulation of DNA damage response 1
RHEB Ras homolog enriched in brain
RIA homologous radioimmunoassay
RICTOR rapamycin-insensitive companion of mTOR
ROK Rho kinase
ROS reactive oxygen species
rRNA ribosomal RNA
RSK p90 ribosomal S6 kinase
RTKs receptor tyrosine kinases
S or Ser serine residues
S6K1 p70 ribosomal S6 kinase 1
SCAP SREBP cleavage-activating protein
SD serine dehydratase
SGK1 protein kinase 1
SH2 Src-homology-2
SHC Src-homology-2-containing protein
SIK2 salt-inducible serine/threonine kinase 2
VI