生化課件老師-信號轉(zhuǎn)導(dǎo)課

1Signal-TransductionPathways信號轉(zhuǎn)導(dǎo)通路鄭利民:

zhenglm2012-11http://Robert

J.

LefkowitzBrian

K.

Kobilka“for

studies

of

G-protein-coupled

receptors”表彰他們對G蛋白耦聯(lián)受體的研究人高度緊張時(shí)“腎上腺素開始大量

”2012年，化學(xué)獎How

could

the

inside

of

the

cell

knowwhat

was

happening

on

the

outside?身體各部位，激素激活腎上腺腦：神經(jīng)信號警告

眼：瞳孔放大，視野變窄心臟：心率加速肺：氣管擴(kuò)張，呼吸頻率加快肌肉：血量增加，肌肉收縮肝臟：糖原分解，糖被到血液脂肪細(xì)胞：脂肪酸被 到血液腸胃：流入消化系統(tǒng)的血量減少Time

to

flee!髓質(zhì)皮質(zhì)醇腎上腺素去甲腎上腺素皮質(zhì)由于衰老和疾病，多種細(xì)胞或

需要修復(fù)；來源？

(胚胎干細(xì)胞)?干細(xì)胞分化：根據(jù)組織局部微環(huán)境的差異而分化成相應(yīng)的細(xì)胞Whether

the

specialisationof

cells

is

reversiblein

human?6干細(xì)胞分化：根據(jù)組織局部微環(huán)境的差異而分化成相應(yīng)的細(xì)胞2011年，Science

評出本世紀(jì)前十年十大科學(xué)成就：5.細(xì)胞重編程：將充分發(fā)育的細(xì)胞進(jìn)行“重編程”，使其成為多能干細(xì)胞-具有成為其身體中任何類型細(xì)胞的能力。John

B.

Gurdon

Shinya

YamanakaFor

the

discovery

that

mature

cells

can

bereprogrammed

to e

pluripotent“發(fā)現(xiàn)成熟細(xì)胞可以重新編程而獲得多能性”2012年，生理/醫(yī)學(xué)獎中樞神經(jīng)軟骨肌肉脂肪上皮畸胎瘤實(shí)驗(yàn)iMiceiPS-derived

mice引入關(guān)鍵體細(xì)胞重新編程iPS,

induced

pluripotent

stem

cell誘導(dǎo)多能干細(xì)胞iPS在疾病治療中的應(yīng)用優(yōu)勢誘導(dǎo)簡單，容易操作可從自體細(xì)胞中獲得，免疫反應(yīng)少無 問題需要探討的問題載體轉(zhuǎn)染的安全性10植物的生長發(fā)育是在環(huán)境因子的影響下正確進(jìn)行時(shí)空表達(dá)的過程How

could

the

inside

of

thecell

know

what

washappeningon

the

outside?signal

transduction!12信號

受體

反應(yīng):

手觸摸就是刺激(信號)，小葉合攏就是反應(yīng)。偶聯(lián)刺激到反應(yīng)之間的生化和分子途徑,就是這個(gè)反應(yīng)的信號轉(zhuǎn)導(dǎo)通路觸摸

含羞草后,小葉合攏細(xì)胞信號轉(zhuǎn)導(dǎo)網(wǎng)絡(luò)的簡單模式（信號輸入）（信號輸出）Signal

Transduction

Pathway:

Complicated1415Important

roles

of

biosignaling

Functional

integration

of

distant

organs,tissues

and

cells

requires

communication;

Signaling

is

perhaps

a

primal

requirement

torespond

to

our

environment;

The

foundation

of

any

complex

responsepathway

lies

with

cellularbiochemicals.BiosignalingIntercellular（細(xì)胞間）&

Intracellular（細(xì)胞內(nèi)）常見四種類型：Endocrine

(內(nèi)

）Paracrine

(旁

）Autocrine

(自

）Membrane

attached

proteinIntercellular

signaling（細(xì)胞間信號）16Four

schemes

of

intercellular

signaling

(1)17Four

schemes

of

intercellular

signaling

(2)1819Intracellular

signaling（細(xì)胞內(nèi)信號）Electron-micrograph

of

macrophage

(pink)attacking

Escherichia

coli

(green)2021信號轉(zhuǎn)導(dǎo)要素：信號或配體,受體,信號放大(產(chǎn)生第二信使),

應(yīng)答和反饋調(diào)節(jié)22Signals

for

phagocytosisFcR

CR3Ca++srcPI3kPKCMAPKgelsolinRhoGTPaseArp2/3PLCPLDActin

rearrangementPhagocytosis; Oxidative

activationSignalReceptorAmplificationTransductionResponsessecondmessengers23PARTⅠBasic

characteristics

of

signal

transductionFour

general

types

of

signal

transducersPARTⅡRegulatory

mechanisms

Some

diseases

caused

by

defects

inthebiosignaling

pathways1

Four

basic

characteristics:SpecificityAmplificationDesensitization/AdaptationIntegration24SpecificitySignal

molecule

fits

binding

siteon

its

complementary

receptor;other

signals

do

not

fit.thrombin25凝血酶often

short-lived&

low

concentration26Desensitization/AdaptationReceptor

activationtriggers

afeedbackcircuit

that

shuts

offthe

receptor

or

removesit

from

the

cell

surfaceProduce

a

rapid

and

major

cellular

responseto

a

transient

signal27Integration28When

two

signals

haveopposite

effects

on

ametabolic

characteristicsuch

as

concentrationof

a

second

messengerX,

or

the

membranepotential

Vm,

theregulatory

eresults

from

theintegrated

input

fromboth

receptors2.

Four

types

of

signal

transducers29Gated

Ion

ChannelsLigand-gated

ion

channelsVoltage-gated

ion

channels2.22.3Receptor

EnzymesG

Protein–Coupled

Receptorsand Second

Messengers2.4 Steroid

Receptors30Four

generaltypes

of

signaltransducers31Why

Ion

Channels

?

Resting

potential:

Asymmetric

ion-distributionActing

potentialGated

Ion

ChannelsLigand-gated

ion

channelsVoltage-gated

ion

channelsResting

potential3233Ion

Conc.

in

Mammalian

Cells

and

Serum

(mM）IonCytoplasmBlood

SerumK+1404Na+12145Cl-4116protein

charges1389Mg+20.81.5Ca+2<0.00021.8Why

Ion

Channels: asymmetric

ion-distributionasymmetric

ion-distribution34Resting

potentialActing

potential35Acting

potentialVoltage-gatedNa+

channels&

K+

channels36pump

and

ion

leak

channelsCl

-leakchannel372.1.1

Ligand-gated

ion

channel:Binding

of

some

small

molecule

forces

anallosteric

transition

in

protein,

open/closechannel.

acetylcholine(乙酰膽堿)receptorion

channel2.1.2

Voltage-gated

ion

channelA

charged

protein moves

relative

tothe

membrane

in

response

to

a

change

intransmembrane

electrical

potential.(voltage-gated

Na+，

Ca2+，K+

channels)38乙酰膽堿受體離子通道

1AAc

Chh39乙酰膽堿受體離子通道240Closed

OpenBinding

of

ACh

to

R

cause

conformationalchange.

As

M2

helices

twist

slightly,

the

Leuresidues

(yellow)

rotate

away

from

the

channeland

are

replaced

by

smaller

polar

residues(blue).

This

gating

mechanism

opens

channel,allowing

passage

of

Ca,

Na,

or

KVoltage-gated

Na+

channels

141Voltage-gatedNa+

channels

2422.

Four

types

of

signal

transducers43Gated

Ion

ChannelsLigand-gated

ion

channelsVoltage-gated

ion

channelsReceptor

EnzymesG

Protein–Coupled

Receptorsand Second

MessengersSteroid

Receptors2.2

Receptor

enzymesA

ligand-binding (胞外)and

an

enzymeactive

site

on

cytosolic

side,connected

by

asingle

transmembrane

segment.Commonly

a

kinase

that

phosphorylates

Tyrresidues

in

specific

proteins（insulinreceptor）Other:

synthesize

the

i.c.

second

messengercGMP

in

response

to

ex.c.

signals

(

thereceptor

for

atrial

natriuretic

factor)44Activation

of

receptor

tyrosine

kinases45Insulin

receptor

tyrosine

kinaseInsulin

structure46anInsulin

receptor

bindsinIsSnuslHiun2li

teoorrfgoGersb2

auptbhoionpsdhpsohsotprohyolPar–tyTelaystrIioRonfSo-1nitosIncRiatSrs-bT1oy.xrySlr-oetsseibdrimuneidnssa.

ltoTyGr

rrbe2s,idtuheesn.

to

Ras,Activated

RasbindscRaaufs-in1g

pGhDosPprheolreyalsaetesEPRhKoasaMnmpndEhdoKovGarecToystnlPaiivnttabetwtiodneodsESinRlekgar1ft-oj1o.

instShReFRrnaeutscoildesuutesims,aunlaadctteivtahteing

it.pthroasnMpshcEorKirpyptlhiaotnsepsahnodrylates

NtruacnlEesRalaKrtitoornnanaosfcTrahiprste&iotnaofTyr

fgaecnteroesrssnidesueuedc,hedaacsftoiEvralkct1ien,lgl

it.adcitvisaiotnin.gthem.47Activation

of

glycogen

synthase

by

insulinRegulation

of

blood

glucoselevel492.

Four

types

of

signal

transducers50Gated

Ion

ChannelsLigand-gated

ion

channelsVoltage-gated

ion

channelsReceptor

EnzymesG

Protein–Coupled

Receptorsand Second

MessengersSteroid

Receptors512.3 GPCR

and

Second

messengersThree

essential

components:a

plasma

membrane

receptor

with

seventransmembrane

helical

segmentsan

enzyme

inthe

plasma

membrane

thatgenerates

an

intracellular

2nd

messengera

guanosine

nucleotide–binding

protein

(Gprotein)GPCR：感受物化刺激，包括多種神經(jīng)遞質(zhì)、肽類激素和趨化因子受體；超過半數(shù)的現(xiàn)代藥物靶向GPCRNobel

Prize

in

Physiology

and

Medicine

1994"for

their

discovery

of

G-proteins

and theroleof

these

proteinsin

signaltransduction

incells"Alfred

G.

Gilman1941-Martin

Rodbell1925-199852Three

essential

components

ofG

Protein–Coupled

Receptors5354A

protein

binds

Guanine

nucleotides

(GDP,

GTP);activated

in

GTP-form,

inactivated

in

GDP-formIntegral

membrane

protein,

heterotrimers

();Have

similar

&

subunits,

but

differ

in

-subunitWhen

G-protein

is

activated,

the

subunitdissociates

to

interact

with

an

enzymes

thatgenerate

second

messengers

(e.g.

cAMP)Others:

small

G-proteins

(~20-25

kDa),

e.g.

Ras,Rho,

Rac,etc,

are

not

membranebound.G

protein

(GTP-binding

protein)55G

protein

(discovery)M.

Rodbell:56a

transducerprovided

the

link

betweenreceptor

andamplifier.A.

G.

Gilman:

identify

&purifythe

G

protein.System:

Mutated

lymphomacells

containing

areceptor

andnormalcAMP-generating

enzyme,

was

yetunable

to

respond

(producecAMP),

since

it

lacked

thetransducermutated

cellnormalcell“ON-OFF”

switch

is

regulated

by

GTP

or

GDPbound

form.

All

G-proteins

has

intrinsicGTPase

activity,

release

Pi

and

inactivated.Activation:

release

of

GDP

and

replaced

by

GTP57The

association

ofactive

Gswithadenylyl

cyclasestimulatesthe

cyclase

tocatalyze

cAMPsynthesis58Adenosine

3’,5’-cyclicmonophosphate(cAMP)59Two

major

systems:THE

PKA

SYSTEM(cAMP

as

the

second

messenger)The

-Adrenergic

Receptor

SystemTHE

PKC

SYSTEM

(DAG,IP3

andCa2+

as

the

second

messengers)Robert

J.

LefkowitzBrian

K.

Kobilka“for

studies

of

G-protein-coupled

receptors”表彰他們對G蛋白耦聯(lián)受體的研究人高度緊張時(shí)“腎上腺素開始大量

”2012年，化學(xué)獎612Illustration

of

Kobilka’s

crystal

structure

of

an

activated

β-adrenergic

receptor

(blue).

A

hormone

(orange)

attaches

tothe

outside

and

a

G-protein

(red)couples

on

the

inside.G蛋白耦聯(lián)受體：感受物化刺激，包括多種神經(jīng)遞質(zhì)、肽類激素和趨化因子受體超過半數(shù)的現(xiàn)代藥物靶向63synthesized

in

adrenal

medulla;belongs

to

catecholamines(兒茶酚胺）;

cells

include

liver,

skeletal

muscle,

heartmuscle

and

adipose;released

in

response

to

acute

stressEpinephrine

腎上腺素signal64Epinephrine

腎上腺素signalingpathwaycAMP65Epinephrine

腎上腺素signaling

pathway

(2)66Activation

of

cAMP-dependent

protein

kinase

(PKA)sInactive

PKA:Regulatory

(R)subunits:

auto-inhibitorybRuriseudbucantitasly:tics(Ca)utsouibnuhniibtsit:

orys

buriedAsPeK-AbindingC

ictksedopbeynRssuubbusntirtaste

bindingsites67Acatalytic

subunit

ofPKAATPPotent

inhibitorpeptide

(PKI):Arg-Arg-Gln-Ala-Ile(consensus

sequencerecognized

by

PKAexcept

Ala)68x

分子Epinephrine

triggersa

series

of

reactionsin

hepatocytes

inwhich

catalystsactivate

catalysts,resulting

in

great“amplification”

ofthe

signal10,000

x

分子69PKA

regulates

a

number

of

enzymesThe

proteins

phosohorylated

by

PKA

sharea

region

of

sequence

similarity

aroundthe

Ser

or

Thr

residue

that

undergoesphosphorylation,

a

sequence

thatmarksthem

for

regulation

by

PKA.The

catalytic

site

of

PKA

interacts

withseveral

residues

near

the

Thr

or

Serresidue

inthe protein,

whichdefine

the

substrate

specificity.707172Desensitization

of the

PKA

systemdesensitizing

β-Adrenergic

Receptordegrading

the

second

messengerGsbg

recruitsbARK

to

themembrane,

whereit

phospho-

SerattheC-terminus

ofRecpt.arr

binds

to

thepi-

C-terminusofRecpt.Receptor-arrestincomplex

enters

thecell

byendocytosis.β-Arrestin

uncouples

receptor

from

its

G

protein

andbrings

together

3

enzymes

of

MAPK

cascade.

(Onestimulus

triggers

two

distinct

pathways:

the

pathactivated

by

G

protein

and

MAPK

cascade)74degrading

thesecondmessenger7576Regulation

of

transcription

by

steroid

hormonesSteroid

receptor77PARTⅠBasic

characteristics

of

signal

transductionFour

general

types

of

signal

transducersPARTⅡRegulatory

mechanisms

Some

diseases

caused

by

defects

inthebiosignaling

pathwaysBiosignaling:

How

are

they

regulated?78Integration79When

two

signals

haveopposite

effects

on

ametabolic

characteristicsuch

as

concentrationof

a

second

messengerX,

or

the

membranepotential

Vm,

theregulatory

eresults

from

theintegrated

input

fromboth

receptorsSopEPKB/AktRac,CDC42PTKPI-3KR調(diào)理后吞噬病原侵襲信號生存信號細(xì)胞凋亡和粘膜屏障損壞細(xì)胞存活細(xì)胞凋亡是由：細(xì)胞內(nèi)“

/生存”信號之間的精密平衡來決定干擾該平衡就可改變病原對細(xì)胞凋亡的最終影響病原侵襲和吞噬對巨噬細(xì)胞凋亡的影響及其機(jī)制80細(xì)胞存活細(xì)胞凋亡細(xì)胞接受到“ 信號”，不一定就會若同時(shí)也接受到“生存信號”，就可繼續(xù)存活The

balance

between

pro-

andanti-apoptoticgenes/signals

determine

the

cell

fate81Regulatory

mechanisms

of

Bio-signals82Phosphorylation

as

aregulatorymechanismRegulation

of

transcription

bysteroidhormonesRegulation

of

the

cell

cycle

byprotein

kinasesRegulation

of

Signaling

PathwaysExternalsignalsSecondmessengersModulatorproteinsFunctionproteinslHormonesOdorantsDrugsLightMitogenichormonesGrowthfactorsPlasma

membranecAMPcGMPCa2+DAG

protein

structural

metabolicIP3

kinase

proteins

orAnd

And

physiologicaTyrosine

phosphatase

enzymes

responseskinases83Nobel

Prize

in

Physiology

and

Medicine

1992Eldwin

G.

KrebsEdmond

H.“Reversible

protein

phosphorylationas

a

biological

regulatory

mechanism"J.

Biol.

Chem.

216:121-132,

195584Protein

PhosphorylationNobel

PrizeNobel

Prize

2000Edmond

H.and

Edwin

G.Krebs

J.

Biol.

Chem.216:121-132,1955Arvid

Carlsson,

PaulGreengard

and

Eric

KandelProtein

Kinases

1:

GPCRs

(5%),

2:

Kinases

(2.8%

genome)~

25%

of

the

mammalian

intracellular

proteinsundergoes reversible

phosphorylation86Protein

kinases

(534

human

kinases)Ser/thrKinasescomprise

80%of

allprotein

kinasesAGC87Phosphorylation

&

de-phosphorylationare

the

most

common

regulatorymechanisms, mediated

by

proteinkinase

and

phosphatase,

respectively蛋白激酶n

NTP蛋白質(zhì)n

NDP蛋白質(zhì)-n

Pin

Pi蛋白磷酸酶H2O88蛋白激酶（protein

kinase，PK）

植物和酵母中～2%的 編碼蛋白激酶；而在哺乳動物中，高達(dá)5－8％。

根據(jù)磷酸化靶蛋白的氨基酸殘基的種類不同，蛋白激酶有絲氨酸/蘇氨酸激酶、酪氨酸激酶和組氨酸激酶等三類。部分蛋白激酶具有雙重底物特異性，既可使絲氨酸或蘇氨酸殘基磷酸化，又可使酪氨酸殘基磷酸化8990Calcium

Is

a

Second

Messenger

inMany

Signal

TransductionsNormally,

[Ca2]i

is

~100

nM(而細(xì)胞外：>1

mM)Hormonal,

neural,

or

other

stimuli

cause

eitheran

influx

of

Ca2+

into

the

cell

through

specificCa2+

channels

in

the

plasma

membrane

or

therelease

of

Ca2+

from

ER

or

mitochondria,Changes

in

[Ca2]i

are

detected

by

Ca2+-bindingproteins

that

regulate

a

variety

of

Ca2-dependent

enzymes-Calmodulin

(CaM)cAMPPKAPhosporylatingcellularproteinsResponse91Activation

of

G

Protein–Coupled

ReceptorsGEF：Guanine

nucleotide

Exchange

FactorGAP：GTPase

Activating

Protein92Activation

of

Ras

proteinSHCGEFRas

-

GTPFRas

-

GDPFPi93GAPMEKERKPLA2ELK1RSKGSK3BADAKTPDK1p70S6KPKCRACPLDRALPKCCa2+FORKHEADFORKHEADCell-cycle

progressionTranscriptionSurvivalTranscriptionCytoskeletal

signalsTranslationTranscriptionVesicle

transportCell-cycle

progressionCalciumsignallingRasRAFPI3KRAC-GEFsRALGDSPLCsMEKERKPLA2

ELK1

RSKGSK3BADAKTp70S6KPDK1

RACCa2+FORKHEADPKCPLD

RAL

FORKHEADPKCCell-cycle

progressionTranscriptionSurvivalTranscriptionCytoskeletal

signalsTranslationTranscriptionVesicle

transportCell-cycle

progressionCalciumsignallingRas蛋白的上游和下游信號通路BADFKHRFKHRp110

p85AktBADCaspase9CREBCREBCREBNFκBFKHRFKHRFKHR14-3-3IKKαIκBIκB

NFκBPDK1Fas-LBim生存cIAP1cIAP1生存Bcl-2mcl-1CytCCaspasecascadeReceptorPI3K－AKT途徑對細(xì)胞生存的調(diào)控機(jī)制9495Most

of

these

complicatedsignaling

pathways

are

transducedby

the

phosphorylation

an