The En Do Membrane System Chapter 12

8/3/2019 The En Do Membrane System Chapter 12

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The EndomembraneThe Endomembrane

SystemSystemChapter 12: The World of the CellChapter 12: The World of the Cell


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Endoplasmic ReticulumEndoplasmic Reticulum

EndoplasmicEndoplasmic-- within the cytoplasmwithin the cytoplasm

ReticulumReticulum networknetwork

FunctionFunction

Biosynthesis of proteins destined for incorporationBiosynthesis of proteins destined for incorporation

into plasma membraneinto plasma membrane

Synthesis of proteins destined for export from cellSynthesis of proteins destined for export from cell

Biosynthesis of lipids a) cholesterol b) plasmaBiosynthesis of lipids a) cholesterol b) plasma

membranemembrane

Incorporation into organelles of endomembraneIncorporation into organelles of endomembrane

systemsystem


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Smooth Endoplasmic ReticulumSmooth Endoplasmic Reticulum

No RibosomesNo Ribosomes

Found often in ovary and testesFound often in ovary and testes

Continuous with rough endoplasmicContinuous with rough endoplasmicreticulumreticulum

tubulartubular


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Example:Example:

Elimination of barbiturate drugsElimination of barbiturate drugs

Phenobarbital induces increase of barbituratePhenobarbital induces increase of barbiturate

detoxifying enzymes in liverdetoxifying enzymes in liver

Extensive formation of smooth ERExtensive formation of smooth ER

Also hydroxylates useful drugs such as antibioticsAlso hydroxylates useful drugs such as antibiotics


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Example:Example:

Member of Cytochrome PMember of Cytochrome P--450 protein family450 protein family

Aryl hydrocarbon hydroxylase involved inAryl hydrocarbon hydroxylase involved inmetabolizing polycyclic hydrocarbonsmetabolizing polycyclic hydrocarbons

Changes carcinogens to chemically activeChanges carcinogens to chemically active

form which results in tumorsform which results in tumors

Cigarette smoke is an inducer of arylCigarette smoke is an inducer of arylhydrocarbon hydroxylasehydrocarbon hydroxylase


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Carbohydrate MetabolismCarbohydrate Metabolism

Membrane of smooth ER of hepatocytesMembrane of smooth ER of hepatocytes

contain glucosecontain glucose--66--phosphase (breaks downphosphase (breaks down

glucoseglucose--66--phosphate into glucose)phosphate into glucose)

Glycogen is stored in liverGlycogen is stored in liver

Allows glucose to leave cell and travel intoAllows glucose to leave cell and travel into

blood systemblood system


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Figure 12-3


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Calcium StorageCalcium Storage

Sarcoplasmic Reticulum in muscle cells is anSarcoplasmic Reticulum in muscle cells is an

example of smooth ER that specializes inexample of smooth ER that specializes in

storage of calciumstorage of calcium

Lumen has ATP dependent calcium ATPasesLumen has ATP dependent calcium ATPases

which aids in muscle contractionwhich aids in muscle contraction


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Steroid BiosynthesisSteroid Biosynthesis

Cholesterol, cortisol, testosterone, estrogenCholesterol, cortisol, testosterone, estrogen

All share a 4All share a 4--ring structure but differ byring structure but differ byhydroxyl groups & carbons side chainshydroxyl groups & carbons side chains

PP--450 monooxygenases are important in450 monooxygenases are important in

steroid hormones by hydroxylationsteroid hormones by hydroxylation


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Rough Endoplasmic ReticulumRough Endoplasmic Reticulum

Transitional Elements (TE)Transitional Elements (TE) plays role inplays role information of vesiclesformation of vesicles

Transition VesiclesTransition Vesicles shuttle lipids and/orshuttle lipids and/orproteinsproteins

Flattened sacsFlattened sacs

Proteins enter lumen cotranslationallyProteins enter lumen cotranslationally

Proteins are anchored by hydrophobicProteins are anchored by hydrophobicinteractions or covalent attachmentsinteractions or covalent attachments

Soluble proteins are released into lumenSoluble proteins are released into lumen


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Phospholipid translocastors (flippases)Phospholipid translocastors (flippases)

move phospholipids from ER to theirmove phospholipids from ER to their

destination by fusing of membranedestination by fusing of membrane

In cases of no fusion, phospholipidIn cases of no fusion, phospholipid

exchange proteins recognize specificexchange proteins recognize specific

phospholipids and move them tophospholipids and move them to

destination (mitochondria, chloroplast,destination (mitochondria, chloroplast,peroxisome)peroxisome)


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Responsible forResponsible foraddition ofaddition ofcarbohydrates tocarbohydrates to

proteinsproteins Folding of proteinsFolding of proteins

Recognition andRecognition andremoval of misfoldedremoval of misfolded

proteinsproteins Assembly ofAssembly of

multimeric proteinsmultimeric proteins

Contains enzymes forContains enzymes forposttranslational andposttranslational andcotranslationalcotranslational

modification such asmodification such asdisulfide bondsdisulfide bonds

Quality controlQuality control ERERassociatedassociateddegradation proteinsdegradation proteinsERAD (cytoplasmicERAD (cytoplasmicproteases)proteases)


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Protein GlycosylationProtein Glycosylation

NN--linked glycosylationlinked glycosylation oligosaccharide unit tooligosaccharide unit to

attach to nitrogen on asparagineattach to nitrogen on asparagine

OO--linked glycosylationlinked glycosylation oligosaccharide unitoligosaccharide unit

to attach to oxygen on hydroxyl group ofto attach to oxygen on hydroxyl group of

serine or threonineserine or threonine


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Formation of Core Oligosaccharide occursFormation of Core Oligosaccharide occurs

in the Cytoplasmin the Cytoplasm

Dolichol Phosphate inserted into ERDolichol Phosphate inserted into ERmembranemembrane

22 NN--acetylglucosime (GlcNAc) are added toacetylglucosime (GlcNAc) are added to

dolicholdolichol

Addition of 5 mannose sugarsAddition of 5 mannose sugars


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NN--GlycosylationGlycosylation

Formation of Core OligosaccharideFormation of Core Oligosaccharide

2 GlcNAc units2 GlcNAc units 9 Mannose9 Mannose

3 Glucose3 Glucose


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Formation of Core Oligosaccharide occursFormation of Core Oligosaccharide occurs

in the lumen of the ERin the lumen of the ER

Translocation by flippase into lumenTranslocation by flippase into lumen

Addition of 4 mannose sugars (total of 9)Addition of 4 mannose sugars (total of 9)

Addition of 3 glucose unitsAddition of 3 glucose units

Transfer of core to nitrogen of asparagineTransfer of core to nitrogen of asparagine

(oligosaccharyl transferase)(oligosaccharyl transferase)

Modification of core oligosaccharide byModification of core oligosaccharide by

removing 3 glucose and 1 mannose sugarsremoving 3 glucose and 1 mannose sugars


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Oligosaccharide isOligosaccharide isusually addedusually addedcotranslationallycotranslationally

Promotes properPromotes properfoldingfolding

Proteins such asProteins such ascalnexin (CNX) andcalnexin (CNX) and

calreticulin (CRT)calreticulin (CRT)binds to glycoproteinbinds to glycoproteinto form a complex forto form a complex fordisulfide bridgedisulfide bridge

Thiol oxidoreductaseThiol oxidoreductase(ERp57) promotes(ERp57) promotesdisulfide bondsdisulfide bonds

If a protein is missingIf a protein is missinga sugar, UGGT (UDPa sugar, UGGT (UDP--glucose: glycoproteinglucose: glycoproteinglucotransferase) willglucotransferase) willadd a sugar soadd a sugar soCNX/CRT willCNX/CRT willpromote formation ofpromote formation ofdisulfide bonddisulfide bond


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Golgi ApparatusGolgi Apparatus

Posttranslational ModificationPosttranslational Modification

Terminal glycosylationTerminal glycosylation removal of a fewremoval of a few

carbohydrates of core oligosaccharidecarbohydrates of core oligosaccharide

OR/BOTHOR/BOTH

Addition ofAddition ofNN--acetylglucosamine and otheracetylglucosamine and other

monosacharides such as galactose, sialicmonosacharides such as galactose, sialic

acid and fucoseacid and fucose Galactosyl transferase is found exclusively inGalactosyl transferase is found exclusively in

Golgi which adds on galactoseGolgi which adds on galactose


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Golgi has 2 major classes of enzymesGolgi has 2 major classes of enzymes

Glucan synthetasesGlucan synthetases catalyzes formation ofcatalyzes formation of

oligosaccharide from monosaccharideoligosaccharide from monosaccharide

Glycosyl transferasesGlycosyl transferases attach carbohydratesattach carbohydrates

to proteinsto proteins

Question:Question: How are the lumenal side of ER andHow are the lumenal side of ER andexterior surface of plasma membrane similar?exterior surface of plasma membrane similar?


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The Golgi ComplexThe Golgi Complex

(1898) Camillo Golgi(1898) Camillo Golgi

Flattened sacs or cisternaeFlattened sacs or cisternae CisCis (CGN) or(CGN) orTransTrans face of Golgi (TGN)face of Golgi (TGN)

BetweenBetween ciscis && transtrans is medial cisternae ofis medial cisternae of

Golgi where processing occursGolgi where processing occurs


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Two Models depect flow of lipids & proteinsTwo Models depect flow of lipids & proteins

Stationary Cisternae ModelStationary Cisternae Model shuttle vesicles bind &shuttle vesicles bind &

fusefuse

Cisternal Maturation ModelCisternal Maturation Model Golgi cisternaeGolgi cisternaetransform from CGN (with aid of ER vesicles) andtransform from CGN (with aid of ER vesicles) and

accumulate specific enzymes. CGN transform toaccumulate specific enzymes. CGN transform to

medial cisternae then trans cisternae Golgi networkmedial cisternae then trans cisternae Golgi network

with new enzymes.with new enzymes.

Time elapsed fluorescence microscopy supportTime elapsed fluorescence microscopy support

maturation modelmaturation model


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Anterograde & Retrograde TransportAnterograde & Retrograde Transport

AnterogradeAnterograde -- vesicles from ER fuse withvesicles from ER fuse with

Golgi then plasma membraneGolgi then plasma membrane

RetrogradeRetrograde -- flow from plasma to Golgi to ERflow from plasma to Golgi to ER


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Golgi Apparatus/ER (rough)Golgi Apparatus/ER (rough)

Protein Trafficking from ER to GolgiProtein Trafficking from ER to Golgi

MembraneMembrane--bound & soluble proteins must bebound & soluble proteins must bedirected to a variety of intracellular locationsdirected to a variety of intracellular locations

including ER, Golgi, endosomes, andincluding ER, Golgi, endosomes, andlysosomeslysosomes

Each protein has a tag for a particular vesicleEach protein has a tag for a particular vesicle

Tag may be amino acid sequence,Tag may be amino acid sequence,

oligosaccharide chain, hydrophobic domainoligosaccharide chain, hydrophobic domain Membrane lipids may be tagged byMembrane lipids may be tagged by

phosphate group by a kinasephosphate group by a kinase


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Golgi Apparatus/ER (rough)Golgi Apparatus/ER (rough)

Retention TagsRetention Tags

RXR (arginine, any, arginine) tag is placed onRXR (arginine, any, arginine) tag is placed on

protein and stays in ERprotein and stays in ER

Retrieval tagsRetrieval tags placed on proteins thatplaced on proteins that

should return to ER. Receptors on Golgi willshould return to ER. Receptors on Golgi will

bind to KDEL (Lysbind to KDEL (Lys--AspAsp--GluGlu--Leu) or KKXXLeu) or KKXX

(Lys(Lys--LysLys--anyany--any) and return proteins to ERany) and return proteins to ERQuestion:Question: If proteins are returned to ER, whyIf proteins are returned to ER, why

go to Golgi?go to Golgi?


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Proteins of GolgiProteins of Golgi

Some have retention or retrieval tags attached toSome have retention or retrieval tags attached to

proteins so they will remain or come back to Golgiproteins so they will remain or come back to Golgi

Specific proteins are integral proteins spanning theSpecific proteins are integral proteins spanning themembrane (hydrophobic regions)membrane (hydrophobic regions)

Hydrophobic regions is a third way to keep proteins inHydrophobic regions is a third way to keep proteins in

GolgiGolgi

Proteins will migrate from CGN to TGN until theProteins will migrate from CGN to TGN until thethickness of membrane exceeds length ofthickness of membrane exceeds length of

hydrophobic membranehydrophobic membrane--spanning domainspanning domain


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Endosome PathwayEndosome Pathway

Lysosomal specific proteins haveLysosomal specific proteins have

mannosemannose--66--phophate formed fromphophate formed from

mannose by 2 Golgi specific enzymesmannose by 2 Golgi specific enzymes

In TGN of Golgi are mannoseIn TGN of Golgi are mannose--6 phosphate6 phosphate

receptors (pH 6.4) where enzymes bindreceptors (pH 6.4) where enzymes bind

and packaged into transport vesicleand packaged into transport vesicle

These vesicles formThese vesicles form Early EndosomeEarly Endosome

which bind to plasma membranewhich bind to plasma membrane


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Endosome PathwayEndosome Pathway

Early Endosome matures to lateEarly Endosome matures to late

endosome (pH 5.5)endosome (pH 5.5)

Bound lysosomal enzymes release fromBound lysosomal enzymes release fromreceptors inreceptors in Late EndosomeLate Endosome. This. This

release prevents retrograde movementrelease prevents retrograde movement

back to Golgiback to Golgi

Late Endosome matures into lysosomesLate Endosome matures into lysosomes


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Secretory PathwaysSecretory Pathways

1967 James Jamieson & George Palade1967 James Jamieson & George Palade

labeled amino acids and watchedlabeled amino acids and watched

movementmovement

After 3 minutes found in ERAfter 3 minutes found in ER

After 7 minutes found in GolgiAfter 7 minutes found in Golgi

37 minutes into vesicles37 minutes into vesicles

117 minutes vesicles discharge protein into117 minutes vesicles discharge protein intoextracellular matrixextracellular matrix


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After budding from TGN, secretoryAfter budding from TGN, secretory

vesicles constantly move to cellvesicles constantly move to cell

membrane and release proteins bymembrane and release proteins by

exocytosisexocytosis

Experiment removed KDEL retrieval tagExperiment removed KDEL retrieval tag

from ER proteins and they did not return tofrom ER proteins and they did not return to

ER but were secreted.ER but were secreted.


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Regulated SecretionRegulated Secretion

Vesicles fuse with plasma membrane only inVesicles fuse with plasma membrane only in

response to extracellular signalsresponse to extracellular signals

Vesicles wait around plasmas membrane untilVesicles wait around plasmas membrane until

signal response tells it to fuse and releasesignal response tells it to fuse and release

Examples: release of insulin form pancreaticExamples: release of insulin form pancreatic

B cellsB cells OROR release of zymogens (precursorsrelease of zymogens (precursorsof hydrolytic enzymes)of hydrolytic enzymes)


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ExocytosisExocytosis

Release of cellular products destined forRelease of cellular products destined forsecretion (triggered by calcium orsecretion (triggered by calcium orhormones)hormones)

Vesicles fuse with plasma membraneVesicles fuse with plasma membrane Discharge contentsDischarge contents

Inner surface of vesicle (lumen of ER andInner surface of vesicle (lumen of ER andGolgi) become outer surface of plasmaGolgi) become outer surface of plasma

membranemembraneAddition of lipids and proteins to cellAddition of lipids and proteins to cell

membranemembrane

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The En Do Membrane System Chapter 12