endomembrane system

chapter 15

internal membranes• endomembrane system

• functions as a coordinated unit• divide cytoplasm into distinct

compartments• controls exocytosis and

endocytosis• movement of molecules which

cannot pass through membrane

transport of proteins• protein synthesis

• begins on a ribosome in cytoplasm

• sorting signals• protein transport

• nuclear transport• protein translocators• vesicular transport

pathways• endocytic

• endosomes & lysosomes• exocytic

• biosynthetic pathway• secretory pathway

• constitutive secretion• regulated secretion

• secretory granules

exocytic endocytic

golgi complex

rough ER

secretory granule

regulated secretory pathway

constitutive secretory pathway

endocytic pathway

endoplasmic reticulum• rough ER

• cisternae -• many bound ribosomes • polarity

endoplasmic reticulum• smooth ER

• functions• synthesis of steroid hormones• liver processing of toxins• sequestration of Ca+

• sarcoplasmic reticulum

biosynthesis in RER• synthesis of proteins

• free ribosomes• cytosolic proteins• peripheral proteins• nuclear proteins

• bound ribosomes• secretory proteins• integral proteins

Completed polypeptide

Incoming ribosomal subunits

Start of mRNA (5′ end)

End of mRNA (3′ end)

Polyribosome

Ribosomes

mRNA

0.1 μm

Growing polypeptides

biosynthesis in RER• bound ribosomes

• bind to ER as needed• SRP (signal recognition particle)• binds to SRP receptor

• translocon

Ribosome

mRNA

Signal peptide

SRP

1

SRP receptor protein

Translocation complex

ER LUMEN

2

3

45

6

Signal peptide removed

CYTOSOL

Protein

ER membrane

biosynthesis in RER• translocon

• SRP interacts with a SRP receptor• ribosome interacts with the translocon• SRP is released

• facilitated by G-proteinsRibosome

mRNA

Signal peptide

SRP

1

SRP receptor protein

Translocation complex

ER LUMEN

2

3

45

6

Signal peptide removed

CYTOSOL

Protein

ER membrane

biosynthesis in RER• Processing

• signal sequence cleaved by signal peptidase• Carbohydrates added

• oligosaccharyltransferase• RER lumen

• chaperones assist in folding• protein disulfide isomerase adds disulfide bonds to

cysteines.

• integral membrane proteins• hydrophobic region generated

• translocon opens laterally• expels transmembrane segment• synthesis continues

biosynthesis in RER

start transfer sequence cleaved

1

2

3

4

biosynthesis in RER• glycosylation

• begins in the RER• membrane bound glycosyltransferases (GTs)

• lipid carrier (dolichol phosphate)• oligosaccharyltransferase -> transfers to polypeptide

• removal of 2 of 3 terminal glucose residues• quality control

• glycoprotein binds with calnexin (chaperone)• terminal glucose removed (by glycosidase II)

• GTs recognize malfolded proteins• tag with glucose --> refolded or destroyed

• transferred to golgi

golgi bodies• golgi apparatus

• functionally distinct cisternae - • cis• trans

• differences in composition between cis and trans face

trans-Golgi network (TGN)

trans cisternae

medial cisternae

cis cisternae

cis- Golgi network

movement of materials• movement across Golgi

• vesicular transport model• vesicles shuttle cargo from

CGN to TGN • cisternal maturation model

• cisternae mature as they move through the apparatus

• combination?• retrograde transport via

vesicles cisternal vesicular

glycosylation• carbohydrate assembly

• glycolipids and glycoproteins• assembled across golgi by glycosyltranferases

vesicle shuttles• vesicle-mediated transport

• donor compartment• acceptor compartment• direction

• anterograde transport• toward plasma membrane

• retrograde transport• away from plasma membrane

• protein coats• two main functions

• cause membrane to curve• select components to be carried

vesicle shuttles• steps in vesicle transport

• (1) cargo selection• sorting mechanisms• coat proteins

• COPI, COPII, clathrin• bulk transport

• proteins trapped

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vesicle shuttles• steps in vesicle transport

• (2) budding• coat proteins interact with

adaptor proteins• COPII - sec23/24,

sec13/31, Sar1• Sar1

• coat recruitment GTPase• GTP bound Sar1 is

membrane bound

COPII-coated vesicle

sec23/24

cargo receptors

donor membrane

sec13/31

outer coatinner coat

assembly of protein coat1. sec12 (a GEF) in membrane bound to cargo receptor

• cargo receptor binds to cargo2. GDP-bound SAR1 binds to sec12, exchanges GDP for GTP

• sec12 is acting as a GEF here3. SAR1-GTP exposes tail which anchors it to ER membrane4. sec23/24 dimer binds to SAR1-GEF complex (assembles

inner coat)• sec13/31 dimer binds to inner coat • curving of membrane may increase affinity for SAR1

vesicle shuttles• steps in vesicle transport

• (3) scission• scission proteins excise bud

• (4) uncoating• coat proteins detach and get

reused

protein cargo

vesicle shuttles• steps in vesicle transport

• (5) transport • microtubule transport via

motor proteins• dynactin

• (6) tethering• Rab proteins bind to tether

proteins on acceptor membrane

Rab

tethering proteins

x

vesicle shuttles• steps in vesicle transport

• (7) docking• soluble NSF attachment

protein receptor• v-SNAREs• t-SNAREs

• (8) fusion • (9) disassembly

• NSF and SNAP proteins

exocytic pathway• TGN

• proteins sorted• secretory (constitutive &

regulated)• lysosomal

• constitutive secretion• plasma membrane proteins

• polarized cells - apical / basal proteins segregated in TGN

• exocytosis• triggered by increase in Ca2+ • vesicle fuses with plasma

membrane

non-lysosomal proteins• regulated secretion

• aggregate as dense granules at TGN• secretory vesicles (granules)• 2 main functions

• timing, amount

lysosome enzyme sorting• sorting proteins at TGN

• lysosomal proteins tagged• phosphorylated mannose• recognized by MPRs

• released in clathrin-coated vesicles

lysosomes• digestive organelles

• contain acid hydrolases• digest a wide variety of biological

molecules• intracellular digestion

• lysosomal storage disorders• absent or defective enzymes

• buildup of undigested materials• Tay-Sachs

lysosomal disorders• Tay-Sachs disease

• autosomal recessive disorder• probably with multiple origins

• deficiency in hexosaminidase A• buildup of ganglioside GM2

• symptoms include• mental retardation, blindness, death

uptake of materials• phagocytosis• endocytosis

• pinocytosis• receptor-mediated

endocytosis

RME• receptor-mediated

endocytosis• clathrin coat

arranged in geometric pattern

lipoproteins on cell surface

clathrin polygons on

inner surface

invagination of clathrin pit

clathrin structure• clathrin

• triskelion• three heavy chains• three light chains

• binds to AP2 adaptor• binds to cargo receptors

clathrin structure• clathrin

• triskelions and adaptors geometrically arranged• dynamin-ring

• promotes pinching off of coated pit into vesicle• uses GTP to tighten around neck of vesicle

• Hsc70 uses ATP and frees clathrin coat

endocytic pathway• endosomes

• formed from fusion of endocytes• sorts materials arriving from PM

• early endosomes (rab5)• derived from PM vesicles

• fuse with early endosome• proteins dissociate - pH affects

conformation• proton pumps lower pH of

endosome• late endosome (rab7, rab9, MPRs)

• as pH goes down, MPRs release hydrolases - recycle to golgi

• lysosome

lysosomal pathways• lysosomes

• autophagy• digestion of organelles

• autophagosome• organelle wrapped in

double membrane• fuses with lysosome

• autophagolysosome

• phagocytosis• engulfing large particles• phagosome

• fuses with lysosome

plant cell vacuoles• vacuoles

• aqueous storage organelles• tonoplast - water vacuole in plants

• contains active transport system for K+ ions • aquaporins