Golgi complexGolgi complex

BIOLOGY, Faculty of Medicine and Dentistry

2015. 10. 05.

László KŐHIDAI, PhD., Assoc. Prof.Department of Genetics, Cell- and

ImmunobiologySemmelweis University

Multiphoton fluorescence image of cultured HeLa cells

Camillo GolgiCamillo Golgi(1843-1926)(1843-1926)

Nobel prize 1906Nobel prize 1906

"internal reticular apparatus"

StructureStructure

• SacculesSaccules• TubulesTubules• VesiclesVesicles

structural-functional unit: structural-functional unit: dictyosomedictyosome 4-6 saccles 4-6 saccles

the structure is polarized into sub-compartmentsthe structure is polarized into sub-compartments

cis Golgicis Golgi medial Golgimedial Golgi trans Golgi trans Golgi

cis Golgi network (CGN)cis Golgi network (CGN) trans Golgi network(TGN)trans Golgi network(TGN)

Dictyosomes of the plant cellsDictyosomes of the plant cells

Synthesis of complexSynthesis of complexpolysaccharidespolysaccharidesof the cell wallof the cell wall

(hemicellulose, pectins)(hemicellulose, pectins)

PositionPosition

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freefreeadhesionadhesion

pattern pattern inducedinducedadhesionadhesion

Relation to the cytoskeletonRelation to the cytoskeleton

Main functionsMain functions

• transporttransport

• sortingsorting

• transformationtransformation

• membrane wrappingmembrane wrapping

Golgi – „Traffic manager” of the cellGolgi – „Traffic manager” of the cell

Main pathways going in and out GolgiMain pathways going in and out Golgi

Pulse-chase techniquePulse-chase technique

dERdERCisternal Cisternal transporttransport

Materials are forwardedMaterials are forwarded together the cisternstogether the cisterns

VesicularVesicular transporttransport

Vesicles trasport substancesVesicles trasport substances from cystern to cisternfrom cystern to cistern

Significance of cytoskeletal networkSignificance of cytoskeletal network

CGNCGN• the peptides arrive from the ER in vesiclesthe peptides arrive from the ER in vesicles• they are N-glycosilatedthey are N-glycosilated• no sorting in the ERno sorting in the ER

Bidirectional transport of proteins:Bidirectional transport of proteins:

• soluble, soluble, endogeneous proteins of the ER endogeneous proteins of the ER recycledrecycled in transport vesicles - in transport vesicles - retention signal retention signal is requiredis required

• sorting and transport of sorting and transport of lysosomal enzymeslysosomal enzymes

Sorting and modification of Sorting and modification of lysosomal enzymeslysosomal enzymes

Mannose-6-phosphate (M-6-P) signaling:Mannose-6-phosphate (M-6-P) signaling:

• based on the recognition of based on the recognition of lysosomal hydrolaseslysosomal hydrolases• recognition of the recognition of the ““signal patchessignal patches” ” (proper 3D combination(proper 3D combination of amino acids) is requiredof amino acids) is required• main working enzyme: GlcNAc-phosphotransferasemain working enzyme: GlcNAc-phosphotransferase

Phosphorylation of the mannoses:Phosphorylation of the mannoses:

• promotes the promotes the sortingsorting of these enzymes of these enzymes • prevents the further modificationsprevents the further modifications

N- and O- glycosylationN- and O- glycosylation

Glycosilation in the GolgiGlycosilation in the GolgiModifications on the Modifications on the N-glycosilationN-glycosilation pattern pattern

• cis-Golgicis-Golgi::mannosemannose-type oligosaccharides-type oligosaccharidescomplexcomplex oligosaccharides oligosaccharides

• TGNTGN::substitution withsubstitution with sialic acids sialic acids - negatively charged - negatively charged

O-glycosilation:O-glycosilation:

• takes place mainly in the takes place mainly in the medial- and trans-Golgimedial- and trans-Golgi• sidechains of Ser and Thr are glycosilatedsidechains of Ser and Thr are glycosilated

© 2000 by Geoffrey M. Cooper

mannse phosphorylationmannse phosphorylation(lysosomal prot.)(lysosomal prot.)

Removal of mannoseRemoval of mannose

removal of mannoseremoval of mannose

labelling w galactoselabelling w galactose

Labelling w syalic acidLabelling w syalic acid

Labelling w GlcNAc Labelling w GlcNAc

sortingsorting

CGNCGN

cis-Golgicis-Golgi

medial-Golgimedial-Golgi

trans-Golgitrans-Golgi

TGNTGN

Structure of Structure of M-6-PM-6-P

Significance of M-6-P labellingSignificance of M-6-P labelling

Other modificationsOther modifications

• glucose-amino-glycane (GAG) chainsglucose-amino-glycane (GAG) chains

• sulphatation (proteoglycanes, Tyr res. of peptides) - TGNsulphatation (proteoglycanes, Tyr res. of peptides) - TGN

• proteolytic modifications - secretion vesicleproteolytic modifications - secretion vesicle

Synthesis of lipids in the GolgiSynthesis of lipids in the Golgi

ceramideceramideglycolipidsglycolipids

sphingomyelinsphingomyelin

Main transport pathways from Main transport pathways from TGNTGN

Main transport pathways from Main transport pathways from TGNTGN

• endosomal-lysosomal compartmentendosomal-lysosomal compartmentvia transport vesicles - via transport vesicles - M-6-P receptorsM-6-P receptors

• surface membrane - secretionsurface membrane - secretionconstitutive secretion constitutive secretion - transports - transports lipids lipids and and peptidepeptidecomponents of the surface membrane andcomponents of the surface membrane andthe extracellular matrixthe extracellular matrix

• exocytosis exocytosis regulated secretionregulated secretion

acid phosphatasetrans Golgi network

osmium reduction

cis-Golgi

unstained

Enzyme content of different compartments in GolgiEnzyme content of different compartments in Golgi

Modifications of secretory Modifications of secretory vesiclesvesicles

• selective aggregation - TGNselective aggregation - TGN

• further modifications and sortingfurther modifications and sorting

inactive precursor - active enzyme or hormoneinactive precursor - active enzyme or hormone

(e.g. preproinsulin - proinsulin - insulin)(e.g. preproinsulin - proinsulin - insulin)

• concentration - loss of waterconcentration - loss of water

• hydratation - e.g. proteoglyganshydratation - e.g. proteoglygans

• uptake some cytoplasmatic substances e.g. histamineuptake some cytoplasmatic substances e.g. histamine

Formation of the acrosome Acrosome is a large lysosome and found in

sperm

It contains enzymes e.g. hyaluronidase, acid phosphatase…

During the sperm differantiation several small vesicles (primary lysosomes) are formed from Golgi

thesmall vesicles are fused to form a single large lysosome - acrosome

Main pathway going out the GolgiMain pathway going out the Golgi

Alternative pathwaysAlternative pathways

• some molecules do not synthesized on the rERsome molecules do not synthesized on the rER (e.g. interleukin 1(e.g. interleukin 1 and 1 and 1 - IL1 - IL1-IL1-IL1, , basic fibroblast growth factor-bFGF) basic fibroblast growth factor-bFGF)

• these molecules transported by ABC-transportersthese molecules transported by ABC-transporters

• other roles of the alternative pathway:other roles of the alternative pathway:- elimination of toxic proteins- elimination of toxic proteins- regulation of protein concentrations in cytosol- regulation of protein concentrations in cytosol

Defects of sorting mechanismDefects of sorting mechanism

lysosomal enzymes do not enter the late endosomes lysosomal enzymes do not enter the late endosomes

BUTBUT

enzymes enter the constitutive secretory pathway and releasedenzymes enter the constitutive secretory pathway and released

I (=inclusion) cell disease:I (=inclusion) cell disease:- - the the M-6-P signal M-6-P signal is not formen on the enzymesis not formen on the enzymes- lysosomal enzymes “- lysosomal enzymes “escapeescape” from the cell” from the cell- deficient intracellular digestion- deficient intracellular digestion- the - the non-digestednon-digested substances form INCLUSIONS substances form INCLUSIONS

Golgi and the cell divisionGolgi and the cell division

Network of membran flow in Network of membran flow in eukaryotic cellseukaryotic cells

Organell-dependent metabolism Organell-dependent metabolism of lipidsof lipids

http://www.youtube.com/watch?v=u38LjCOvDZU