Er to golgi transport
Download
1 / 49

ER-to-Golgi Transport - PowerPoint PPT Presentation


  • 128 Views
  • Uploaded on

ER-to-Golgi Transport. Pathway Maturation De Novo Golgi Biogenesis Mitotic Assembly/Disassembly Post-ER Processing. Pathway Diagram. Sorting, Retention, Retrieval. ER. ERGIC. Golgi. The Golgi is the central processing and sorting station of the secretory pathway. ER. ERGIC. Golgi.

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about ' ER-to-Golgi Transport' - byron


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
Er to golgi transport
ER-to-Golgi Transport

Pathway

Maturation

De Novo Golgi Biogenesis

Mitotic Assembly/Disassembly

Post-ER Processing




ER

ERGIC

Golgi

The Golgi is the central processing and sorting station of the secretory pathway

ER

ERGIC

Golgi







The Golgi maintains its complex structure in the presence of continuous membrane traffic

trans

medial

cis


3d view of golgi stack
3D View of Golgi Stack continuous membrane traffic


By what mode does golgi traffic occur

By what mode does Golgi traffic occur? continuous membrane traffic


Vesicle maturation models
Vesicle & Maturation Models continuous membrane traffic


Enzymes in copi
Enzymes in COPI? continuous membrane traffic

Table II. Relative lateral distribution of Golgi proteins and VSV-G within the Golgi complex

Golgi stack (cisternae) Lateral rims (uncoated) Coated buds and vesicles

Man II 68.9 ± 3.9 13.6 ± 2.7 17.5 ± 2.8

Giantin 58.2 ± 2.6 18.3 ± 2.8 23.5 ± 2.7

KDELr 35.8 ± 4.3 8.9 ± 2.0 55.3 ± 4.5

rBet1 25.6 ± 4.9 15.7 ± 4.0 58.7 ± 5.6

VSV-Gts045–GFP (20 min, 32°C)

anti-GFP 93.1 ± 1.4 4.3 ± 1.3 2.6 ± 0.6

anti–VSV-G–lum

94.1 ± 0.8 3.4 ± 0.6 2.5 ± 0.5

------------------------------------------------------------------------

Numbers represent the percentages (mean ± SEM) of the total labeling over the distinct membrane

categories and were obtained by analyzing 50 Golgi complexes for each antibody.


Enzymes not in copi
Enzymes not in COPI? continuous membrane traffic


Copi vesicles concentrate enzymes not cargo
COPI Vesicles Concentrate Enzymes not Cargo continuous membrane traffic

------------------------------------------------------------------------

Markers Amount of vesicles (% of starting membranes)

------------------------------------------------------------------------

GTP GTPS arf-1 Q71L

------------------------------------------------------------------------

Golgi enzymes

Mann II 13.2 ± 1.3 6.1 ± 1.4 4.2 ± 1.1

NAGT I 25.8 ± 2.7 3.1 ± 1.0 3.0 ± 0.8

GalT 11.0 ± 2.0 1.7 ± 0.4 1.8 ± 0.4

Anterograde cargo

pIgR  4.6 ± 1.0 3.6 ± 1.4 3.3 ± 1.1

Phospholipids  2.7 ± 0.7 3.1 ± 0.7 2.5 ± 0.8

------------------------------------------------------------------------


How is the golgi formed
How is the Golgi Formed? continuous membrane traffic


Trafficking may be sufficient for golgi self assembly
Trafficking may be sufficient for Golgi self-assembly continuous membrane traffic

Golgi

ERGIC

cis

medial

trans

Trans Golgi Network

Homotypic fusion

MT

COP I

COP I

COP I

ER

COP II


Can the golgi self assemble

Can the Golgi self-assemble? continuous membrane traffic

Idea: Use a reversible ER export block

-Brefeldin A to collapse Golgi

-H89 to block ER export and collapse ERGIC



Gm130 co fractionates with er membranes

GM130 co-fractionates with ER membranes continuous membrane traffic

Golgi

ER

untreated

BFA H89


Re emergence of the golgi from the er

min after H89 washout continuous membrane traffic

0’

4’

Re-emergence of the Golgi from the ER

GM130

10’

30’


Reassembled golgi membranes are stacked
Reassembled Golgi membranes are stacked continuous membrane traffic

BFA H89

30’ washout


Early compartmentalization of golgi membranes after er exit
Early compartmentalization of Golgi membranes after ER exit continuous membrane traffic

5’

Giantin

GPP130



De novo biogenesis continued
De novo biogenesis continued continuous membrane traffic


Transport paths consequences of their regulation
Transport paths & consequences of their regulation continuous membrane traffic

Mitotic: vesicles=>block in docking

Osmotic:ER=>ER export block


M phase golgi vesiculation
M-phase Golgi Vesiculation continuous membrane traffic



Anti-p115 injection induces dispersion of the Golgi continuous membrane traffic in vivo



What are the essential p115 interactions? continuous membrane traffic

(i.e. candidates for regulation at mitosis)

acidic tail

head domain

coiled-coil domain

Rab1

Giantin GM130

PLCg1

SNAREs

phosphorylation site

GBF1


Beauty is in the rnai of the beholder
Beauty is in the RNAi of the Beholder continuous membrane traffic


100 continuous membrane traffic

80

60

40

mock

24 h

48 h

72 h

96 h

20

0

mock

96 h

p115 siRNA induces time-dependent depletion of p115

GPP130

p115

p115

mock

%intact

p115 levels

siRNA


siRNA-induced p115 depletion results in Golgi fragmentation continuous membrane traffic

p115

GPP130

mock

siRNA


20 continuous membrane traffic

40

60

80

100%

f

+p115

Mismatched p115 rescues siRNA-induced Golgi fragmentation

inj. marker

p115

GalNac-T2

% injected cells with intact Golgi


O continuous membrane traffic

The giantin/GM130-binding domain is NOT required for Golgi biogenesis in vivo

WT

S941A

Dtail

Acidic Tail

20

40

60

80

100%

Targeted?

+

WT

+

S941A

+

Dtail

% expressing cells with intact Golgi


O continuous membrane traffic

WT

Dcc1

The SNARE domain of p115 is required for Golgi biogenesis in vivo

cc1

+

+

+

+

+

?

+

Targeted?


Working model for mechanism of action of p115 continuous membrane traffic

recruitment of p115 by Rab1

binding to

t-SNARE

catalysis of SNARE pairing

p115

v-SNARE

Rab1

t-SNARE

=> dissect SNARE mechanism w/ further mutagenesis

=> test other p115 interactions-> distinct phenotypes?


rbet1 continuous membrane traffic

syntaxin5

membrin

sec22

Sub-regions of the cc1 domain show homology to distinct SNAREs

rbet1

cc1

core complex domain

syntaxin5

membrin

p115 homology region

sec22


Part 1 post er processing
Part 1 Post-ER Processing continuous membrane traffic

Modification of carbohydrate side-chains

Proteolytic activation of precursors



Branched carbohydrate continuous membrane traffic


Evidence for enzyme compartmentalization continuous membrane traffic


Processing Reactions continuous membrane traffic

endoH sensitive

endoH resistant

cis

trans


Example of proteolytic activation
Example of proteolytic activation continuous membrane traffic


Proton pumps reduce lumenal ph
Proton pumps reduce lumenal pH continuous membrane traffic


ad