Lecture 12. The ER as a Portal and Stress Sensor Flashcards
What are the five major interdependent strategies of ER selective transport?
Cargo capture: receptor-mediated export of proteins from the ER to the Golgi complex in coatamer protein II (COPII) vesicles
Bulk Flow: some proteins and lipids are included in COPII vesicles by default
Retention: Prevents proteins from entering the transport vesicles
Retrieval: retrograde transport from the ER-Golgi intermediate compartment (ERGIC)/early Golgi back to the ER
ERAD: Cytosolic elimination of ER proteins that fail quality control
What happens with cargo capture and anterograde transport?
- In the ER, secretory cargo is loaded into COPII (coatamer protein II) transport vesicles at ER exit sites (ERES). This requires export signals in fully folded client proteins and cargo receptor proteins in the vesicle membrane
- COPII vesicles fuse to form the ER-Golgi intermediate compartment (ERGIC). When COPII vesicles are close to the cis-Golgi membrane, they shed their coats - COPII components are recycled
- The receptors usually return to the ER by retrieval pathways
What happens with bulk flow and retention?
- Export by bulk flow does not require receptors or export signals. Some soluble and membrane proteins (and membrane lipids) enter COPII vesicles by default
- There is a biotechnological benefit: foreign proteins directed into the ER are often secreted into the growth medium as soluble proteins that are relatively easy to purify (because they lack species-specific recognition signals)
- Retention: some proteins are selectively excluded from COPII vesicles
What happens in the COPI retrieval pathway?
COPI coated vesicles retrieve transport machinery, cargo receptors, lipid membrane, and escaped ER-resident proteins from ERGIC and the cis-Golgi and delivers it back to the ER
What do the retrieved proteins and soluble proteins from COPI coated vesicles have?
Retrieved membrane proteins typically possess a C-terminal dilysine motif (KKXX) or a close variant (e.g OST (human) …EKEKSD)
Retrieved soluble proteins typically have a C-terminal ‘LDEL’ motif (HDEL in yeast and K/HDEL in plants)
How is Rab6 involved in retrieval and retrograde transport?
Rab6 organises the return via long tubular elements of membrane proteins transferred forward
Not currently well-characterised (used by bacterial Shiga toxin that binds as a lipid receptor)
What are the two categories of stresses that increase misfolding/stimulate unfolding?
Abiotic stresses and biotic stresses
What are examples of abiotic stresses?
Heat stress (in plants, fungi and poikilothermic animals especially, but also in
homeothermic animals)
Osmotic stress (Plants: drought, high salinity – can severely impact crop plants)
High light intensity (plants)
What are examples of biotic stresses?
Infection (plants, animals, fungi)
Stress related hormones (salicylic acid secreted by a competitor plant, abscisic acid)
How can the ER act as a stress sensor in the unfolded protein response (UPR)?
The protein folding capacity of the endoplasmic reticulum (ER) is tightly regulated
by a network of signalling pathways - the unfolded protein response (UPR)
UPR sensors monitor the ER folding status of proteins in the ER
Following sensing, the UPR adjusts the folding capacity of the ER according to need
What are the multiple sensing mechanisms mediated by/what are three stress sensors?
Ire1 (Inositol-requiring Enzyme 1) – animals, plants, fungi
PERK (PRKR-like endoplasmic reticulum kinase) – animals and fungi
ATF6 (activating transcription factor 6) – animals and fungi (bZIP28 and bZIP17 – plants)
What is the structure of Ire1 and what causes Ire1 to dimerise/multimerise?
Ire1 is a transmembrane protein and is bound to BiP in the ER lumen that keeps it inactive
As ER stress occurs and more unfolded/misfolded clients emerge, BiP unbinds from Ire1 stress sensor to bind to the the clients
Ire1 dimerises (and also binds to unfolded proteins)
How is Ire1 activated?
Freeing Ire1 from the inactivating effects of BiP allows Ire1 molecules to activate each other through auto-transphosphorylation, cytosolic domains activate each other by kinase activity
Level of stress correlated with level of Ire1
How does activated Ire1 splice a specific RNA Ire1 not just a kinase but also an RNase)?
Recognises very specific, highly conserved cytosolic RNA that has two stem loops
Ire1 cuts this RNA with association from a specific ligase, resulting in the two exons being spliced together, leaving an unusual cytosolic intron
The splicing of the two exons creates an open reading frame that’s translated to make a transcription factor which is transported to the nucleus to reduce ER stress through activation of multiple pathways (ERAD, folding and trafficking pathways all upregulated)
What is the difference between splicing done by the spliceosome and Ire1?
Most eukaryotic splicing requires two transesterifications coordinated by snRPs, and occurs in the nucleus. Ire1 has RNAse activity that recognises a specific cytosolic RNA, and which removes a highly conserved intron