From gene to phenotype - Institut national de la recherche ... · DESNOUES Elsa, BALDAZZI...

From gene to phenotype: genetic control and modeling

of sugar metabolism during peach fruit development

DESNOUES Elsa,

BALDAZZI Valentina, GÉNARD Michel,

LAMBERT Patrick, CONFOLENT Carole, HEURTEVIN Laure,

QUILOT-TURION Bénédicte

Sugar concentration is an important factor of fruit quality

In peach fruit:

Sucrose Glucose Fructose Sorbitol

Moriguchi et al. 1990, Cantin et al. 2009

Sugar concentration depends on:

- environment- genotype

2Kanayama et al. 2005; Moriguchi et al. 1990

0

5

10

15

20

0 5 10 15 20

Fru

cto

se (

mg.

gFW

-1)

Glucose (mg.gFW-1)

Introduction | Phenotyping | Model Simulation | Genetic control | Conclusions

3

Sucrose

FructoseUDP-Glucose Glucose

SuSyNI

Sorbitol

SDH SO

Fructose 6P Glucose 6P

FK HK

PGI

PGMGlucose 1P

Sucrose 6P

SPS

Fructose 1,6 BisP

PFK F1,6 BisPase

UGPase

SPP

PFP

Sucrose

Fructose Glucose

Sorbitol

AI

VacuoleCytosol

Knowledge from literature is incompleteEspecially the compartmentation is poorly described

LoBiancoet al. 1999

Kobashiet al. 2000

Kanayamaet al. 2005

Lombardoet al. 2011

Morandi et al. 2008

Borsaniet al. 2009


Proteomics

Metabolomics

4

Sucrose


SuSyNI

Sorbitol

SDH SO


FK HK

PGI

PGMGlucose 1P

Sucrose 6P

SPS

Fructose 1,6 BisP

PFK F1,6 BisPase

UGPase

SPP

PFP

Sucrose

Fructose Glucose

Sorbitol

AI

VacuoleCytosol

No study on sugar carriers in peach

Proteomics

Metabolomics

Data from literature


5Dirlewanger et al. 1999; Etienne et al. 2002; Quilot et al. 2004

1 2 3 4 5 6 7 8

Schematic representation of the QTL controlling sugars in peach at maturity

Genomics

Metabolomics

Sucrose Sorbitol GlucoseFructose

What is the genetic control of sugar metabolism in peach ?

No study on the genetic control:

- during fruit development- on enzymatic capacities


6

Study the metabolism of sugars from metabolic, enzymatic and genetic points of view

Understanding the mechanisms underlying the phenotypic variability of sugar concentrations in peach

Genomics

Metabolomics

Proteomics

Approach

Characterize the phenotypic variability (metabolites and enzymatic capacities)

Develop a metabolic model taking into account the enzymatic capacities and compartmentation

Analyze the genetic control of sugar metabolism


7

Prunus davidiana

P. persica

Summergrand X

SD40 P. persica

Summergrand

X

X

BC1 P. persica

Zéphir

BC2

1 major gene

QTL

0 10 20 30

010

2030

glucose (mg/g MF)

fruct

ose

(mg/

g M

F)

Glucose (mg/gMF)

Fru

ctos

e (

mg/

gMF

)


Quilot et al. 2004

8

- 6 developmental stages- 106 genotypes- 9 metabolites - 12 enzymatic capacities

Metabolome Facility of Bordeaux Functional Genomics Center, IBVM, INRA, F-33140 Villenave d’Ornon France

PFP

Sucrose


SuSy

NIAI

Sorbitol

SDH SO


FK HK

PGI

PGMGlucose 1P

Sucrose 6P

SPS

Fructose 1,6 BisP

PFK F1,6 BisPase

UGPase

SPP

Citrate

Malate

We developped enzymatic assays for peach fruit analyses


A quasi complete description of sugar metabolism

9

040

080

012

00P

GM

a

010

3050

F16

BP

ase

b

010

020

030

0S

uSy

c

5010

020

0F

K

d

020

060

0U

GP

ase

e

5015

025

0H

K

f

020

4060

SP

S

g

010

030

0S

DH

h

010

020

030

0S

O

i

30 50 70 90

05

1015

20P

FK

j

30 50 70 90

010

020

030

0N

I

k

30 50 70 90

050

100

150

AI

l

Fruit age (% of fruit development)

Enz

yme

capa

city

(nm

ol g

FW

-1 m

in-1

)

010

20

30G

luco

se

a

05

10

1520

Fru

ctos

e

b

30 50 70 90

04

08

01

20S

ucro

se

c

30 50 70 90

02

46

812

Sor

bito

l d

Fruit age (% of fruit development)

Co

nce

ntra

tion

(mg

gFW

-1)

Enzymatic capacities are relatively stable over time and between genotypes


Desnoues et al. BMC Plant Biology 2014

Metabolite profiles Profiles of enzymatic capacities

standard phenotype

‘low-fructose’ phenotype

10

-1

-0.75

-0.5

-0.25

0

0.25

0.5

0.75

1S

orbi

tol

Suc

rose

Glu

cose

Fru

ctos

e

PG

MF

16B

Pas

eS

uSy

FK

UG

Pas

e

HK

PF

K

SP

SS

DH

SO

NI

SucroseGlucoseFructose

PGMF16BPase

SuSyFK

UGPaseHK

PFKSPSSDH

SONIAI

no correlation between the rate of accumulation of sugars and directly linked enzymatic capacities


The relationships between enzyme capacities and metabolites are not direct and are difficult to assess using conventional statistics

11

SucCyt

FruCyt GluCyt

SorCyt

HexP

Other compounds

Cytosol

FruVac GluVac

SucVac

Vacuole

Respiration

AISOSDHNI

SuSy

HKFK

SPS + SPP

F9F10

F11 F12

F14

F15

F16

F13 F13F9 F10

F17

F18

F1F2

F3F3

F4

SorVac

F19

F6

F5

F8

F7

- compartmentation between cytosol and vacuole

- enzymatic reactions

Metabolic model simulating the concentrations of sugars during fruit development


12

FruCyt

Cytosol VacuoleF3


13

FruCyt

Cytosol Vacuole

SorCyt

SDH

F11

F3

Enzymatic reactions:irreversible Michaelis-Menten equations


14

FruCyt

Cytosol Vacuole

SorCyt

SDH

F11

F3

Vmax : enzymatic capacities


15

FruCyt

Cytosol Vacuole

SorCyt

SDH

F11

F3

Km : enzyme affinity for the substrate


16

FruCyt

Cytosol VacuoleF3

SucCyt

NI

SuSyF10

SDH

F11

F9

SorCyt


17

FruCyt

Cytosol VacuoleF3

FruCyt

SucCyt

NI

SuSyF10

SDH

F11

F9

SorCyt

HexP

F14FK


18

FruCyt

Cytosol VacuoleF3

FruCyt

SucCyt

NI

SuSyF10

SDH

F11

F9

SorCyt

F14FK

HexP

FruVac

F7

Active transport: irreversible Michaelis-Menten equation


19

FruCyt

Cytosol VacuoleF3

SucCyt

NI

SuSyF10

SDH

F11

F9

SorCyt

F14FK

HexP

FruVac

F7

F8

Passive transport: proportional to concentration gradient


20

A set of 9 coupled differential equations



21

SucCyt

FruCyt GluCyt

SorCyt

HexP

Other compounds

Cytosol

FruVac GluVac

SucVac

Vacuole

Respiration

AISOSDHNI

SuSy

HKFK

SPS + SPP

F9F10

F11 F12

F14

F15

F16

F13 F13F9 F10

F17

F18

F1F2

F3F3

F4

SorVac

F19

F6

F5

F8

F7


19 fluxes and 30 parameters



Predictions of sugar concentration during fruit growth

22


Desnoues et al. submitted


Simulation of genetic variability

23




24

Simulation of genetic variability





25


Desnoues et al. J Experimental Botany 2016

00Sugar_transporter_x2745863

Sugar_transporter2736240SDH4114761Invertase4119839Sugar_transporter5023003Sugar_transporter6695852Invertase_inhibitor8477731Invertase8938072Invertase_inhibitor_x69686184SuSy10373098Invertase_inhibitor_x310389594Sugar_transporter10545923Sugar_transporter_x211381808Sugar_transporter_x1012363662SPS12709396Invertase_inhibitor15307207Sugar_transporter_x217203398FK Sugar_transporter18771430

Sugar_transporter23536346G6PDH23636824Sugar_transporter25772752Invertase_inhibitor26159866Sucrose_transporter Sugar_transporter27849252FBPase30673980PGM31271164PGM31359328Invertase_inhibitor_x231362452Invertase33539832HK33567152AGPase34099232Sugar_transporter34202992PGI34319936Sugar_transporter34385936PFK37632744Sugar_transporter40219568UGPase40693560Sugar_transporter_x341488424PGI43241936Invertase_inhibitor_x243302152Sucrose_transporter44281716Sugar_transporter44347180Invertase45471252F16BPase45788260Sugar_transporter47454368

FK

_1

Fru_123456

FW

_56

Glu_12

HK

_4

HK

_345_Z

Mal_12

PG

M_2

Sor_1

Suc_56

UG

Pase_1

UG

Pase_45_Z

1

00

Sugar_transporter_x53861624Invertase_inhibitor_x27242234Invertase_x213288728Invertase_inhibitor_x315600150UGPase16498039Invertase_inhibitor17347456Sugar_transporter_x317790424Invertase_inhibitor_x319757308FK20600680Sugar_transporter22328702Invertase_x223089256Sugar_transporter23647160Invertase_inhibitor24339340PGM24494820SuSy26137574Sugar_transporter26275988F16BPase26281136UGPase26687712Sugar_transporter27382916PFK27579252Sugar_transporter_x227659540Invertase27865048Invertase_inhibitor_x527937884Sugar_transporter28196656PGM28289944FK Sugar_transporter28301506SDH28365856Sugar_transporter_x229229556Invertase_inhibitor29387138Sugar_transporter_x230205096

Cit_345

Mal_345

Sor_6_Z

2

00Invertase_inhibitor_x5474999Invertase606163SuSy1010886UGPase1069857Sugar_transporter2015795Invertase_inhibitor2300979Sugar_transporter2550134Invertase3405921PFK4017399HK4096447Sugar_transporter_x35103365

Sugar_transporter15757313Invertase_inhibitor16038192Sugar_transporter16048486Sugar_transporter_x216644957FK17922916AGPase20466452G6PDH20821072Sugar_transporter21629154PFK21648144

Sugar_transporter25751272PGI26516740G6PDH26531676F26BPase27071116

FW

_45F

16BP

ase_45

FK

_5

UG

Pase_6_Z

3

4 5

00Invertase563185FBPA2294425F26BPase3770137Sugar_transporter4154356Sugar_transporter4844200Invertase4977474

FK

_2

6

uc_1

00Invertase_inhibitor131826PFK164366Sugar_transporter426241PFK671854Invertase_inhibitor1195495Sugar_transporter_x32015166Sugar_transporter3536606UGPase3599032Sugar_transporter4096771UGPase6203523Sugar_transporter_x36987171F16BPase7583583Sugar_transporter_x28922957Sugar_transporter12106768Invertase12882369SDH15817263

HK17960292Sugar_transporter18369144

FW

_5F

K_56_Z

Fru_12345

Fru_12_Z

FW

_2_Z

Glu_234

Glu_56_Z

HK

_6_Z

Sor_3456_Z

4uc_

Ci

PG

M_2

00SPP311315PFK464208Sugar_transporter696032Sugar_transporter717190Invertase2681142Sugar_transporter3092369PGM4993486Invertase_inhibitor_x25183213Invertase8955357Invertase_inhibitor_x29094726Sugar_transporter_x410124114Invertase_inhibitor11636050Sugar_transporter11755403Sugar_transporter12303785Sugar_transporter_x213449100Sugar_transporter14922045Invertase_inhibitor15581665FBPA17778792SuSy18195912Sugar_transporter18204020

Cit_56_Z

FK

_12

FK

_35

Glu_1235

HK

_34

Sor_2 S

uc_1

5

00Invertase563185FBPA2294425F26BPase3770137Sugar_transporter4154356Sugar_transporter4844200Invertase4977474

PFK8914315Invertase9128385

Sugar_transporter13074689

Sugar_transporter19048482Sugar_transporter19552856PFK20389672Invertase_inhibitor20457400HK22040650Invertase23318824Invertase25563296Sugar_transporter_x225911446Invertase26864548G6PDH27709052Invertase_inhibitor_x227789478Invertase27815656Invertase28061726Invertase_inhibitor_x228269920Sugar_transporter_x430511084

FK

_2F

K_56

FW

_123

HK

_56

PG

M_56

Suc_23

6

uc_1

00UGPase400727

SPP12296588Invertase_x313293841FBPA14159140FBPA14164351Sugar_transporter16096653FBPA16968676Sugar_transporter_x317993352Invertase_inhibitor_x418250054SuSy18350216Invertase_inhibitor_x218416980PFK19574976HK19677704Sugar_transporter_x520283858Sugar_transporter21041784SuSy21151882Sugar_transporter21354476AGPase Sugar_transporter22198854

FW

_456

Glu_1234

Mal_2345

NI_3

PG

M_1_Z

SO

_6

Sor_2

Suc_12356

7

00FK98079SPS302873Sugar_transporter1630354Sugar_transporter1834704F16BPase2566771Invertase_inhibitor3725038Sugar_transporter_x25814548F16BPase10340602Sugar_transporter11181030Sugar_transporter11690795FBPA11812456AGPase12662374Sugar_transporter_x313225762Sugar_transporter_x413658102SDH15994437SDH15999040Invertase16872814Sugar_transporter_x217164258Sugar_transporter20532424Sugar_transporter_x221474404Sugar_transporter21697140Invertase_inhibitor_x422159222SuSy22179196Invertase_inhibitor22322926

Mal_34

PF

K_345_Z

Suc_56

8

uc_1

QTL detection for metabolites and enzymatic capacities at different stages

26



We detected QTLs for metabolites and enzymatic capacities

Dynamic QTLs

27


genotypes with the wild allele are in red

Prunus davidiana alleles display changing effects during development


28



00Sugar_transporter_x2745863

Sugar_transporter2736240SDH4114761Invertase4119839Sugar_transporter5023003Sugar_transporter6695852Invertase_inhibitor8477731Invertase8938072Invertase_inhibitor_x69686184SuSy10373098Invertase_inhibitor_x310389594Sugar_transporter10545923Sugar_transporter_x211381808Sugar_transporter_x1012363662SPS12709396Invertase_inhibitor15307207Sugar_transporter_x217203398FK Sugar_transporter18771430

Sugar_transporter23536346G6PDH23636824Sugar_transporter25772752Invertase_inhibitor26159866Sucrose_transporter Sugar_transporter27849252FBPase30673980PGM31271164PGM31359328Invertase_inhibitor_x231362452Invertase33539832HK33567152AGPase34099232Sugar_transporter34202992PGI34319936Sugar_transporter34385936PFK37632744Sugar_transporter40219568UGPase40693560Sugar_transporter_x341488424PGI43241936Invertase_inhibitor_x243302152Sucrose_transporter44281716Sugar_transporter44347180Invertase45471252F16BPase45788260Sugar_transporter47454368

FK

_1

Fru_123456

FW

_56

Glu_12

HK

_4

HK

_345_Z

Mal_12

PG

M_2

Sor_1

Suc_56

UG

Pase_1

UG

Pase_45_Z

1

00

F

4

00

5

uc_1

00UGPase400727

SPP12296588Invertase_x313293841FBPA14159140FBPA14164351Sugar_transporter16096653FBPA16968676Sugar_transporter_x317993352Invertase_inhibitor_x418250054SuSy18350216Invertase_inhibitor_x218416980PFK19574976HK19677704Sugar_transporter_x520283858Sugar_transporter21041784SuSy21151882Sugar_transporter21354476AGPase Sugar_transporter22198854

FW

_456

Glu_1234

Mal_2345

NI_3

PG

M_1_Z

SO

_6

Sor_2

Suc_12356

7

uc_1

Co-locations between annotated genes, QTLs for enzyme activities and QTLs controlling metabolite concentrations

QTLs for capacity of enzymes <-> gene controlling the enzymeQTLs for sugars <-> genes controlling linked enzymesQTLs for sugars <-> genes coding for sugar transporters or enzyme inhibitors

29


We progressed in the description of - the sugar pathway in peach- the kinetics during peach development- the genetic control of metabolites and enzymatic capacities

We developed a metabolic model that can be used as a tool to explore sugar metabolismWe highlighted lack of knowledge on sugar carriers in fruit

1 major gene

QTL

0 10 20 30

010

2030

glucose (mg/g MF)

fruct

ose

(mg/

g M

F)

Glucose (mg/gMF)

Fru

ctos

e (

mg/

gMF

)

We are currently working on the genetic control of ‘low-fructose’ phenotype

INRA GAFL - UR 1052 INRA PSH UR - 1115

INRA BFP UMR - 1332

Yves GibonDuyen ProdhomePatricia Ballias

Michel GénardValentina Baldazzi

Doriane BancelGilles Vercambre

Bénédicte Quilot-TurionMathilde Causse

Véronique SignoretLaure HeurtevinCarole ConfolentEsther Pelpoir

Fabrice FlamainG2IP team

Lee SweetloveBernadette Gehl

Barbara GoublePatrice Reling

Fabien ChardonMagali Bedu

INRA SQPOV UMR - 408

Thanks to Elsa Desnoues

From gene to phenotype - Institut national de la recherche ... · DESNOUES Elsa, BALDAZZI...

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Transcript of From gene to phenotype - Institut national de la recherche ... · DESNOUES Elsa, BALDAZZI...