Endoplasmic Reticulum

What is the ER?

This is the starting point for secretory and biosynthetic pathways across to other organelles

What are the struictures of the ER?

They have sheets and tubes

What isn the nuclear membrane?

A shingle sheet of ER that is structurally maintained by cytoskeleton and LINK complex spanning the inner and outer membrane maintaining distance

Where is the NE most curved?

At the NPCs

What are the curved strucutres of the ER called?

Cortical ER, they are highly curved and made of both sheets and tubes, the outside membrane portion has no ribosomes but the cytoplasm facing side have many ribosomes

How are ER matrices and nanoholes made?

Dynamic structures in the ER membrane, membrane shaping and fusion proteins are localized here for their formation, involves reticulons and atlastins to form them

What are the role of ER matrices and nanoholes?

May help ER to rapidly change shape during cell migration, tubular junctions at ER matrices provide increased surface area for lipid synthesis, nanoholes may allow for rapid movement of membrane shaping proteins like reticulons

What is membrane curvature needed for?

Vesicle formation, nuclear pore synthesis, membrane fusion and repair

How do you create membrane curvature?

Uniform lipids make a flat membrane, in order to curve it you need more lipids with helical structure or those with longer tails and different charges, we use modification of local lipid composition, wedge-shaped proteins and interactions of TM proteins with lipids and clustering

What are wedge shaped proteins?

These are proteins that are wedge-shaped and cause membrane curvature creating asymetry through their TMDs

What way do positive and negative charged lipids cause bending

+ve bends towards the cytoplasm and -ve bends away from cytoplasm

How does modification of local lipid composition cause bending?

Adding lipids of different charges and lipids of different head and tail size cause asymmetry that leads to bending

What are examples of intrinsic membrane bending mechanisms?

amphipathic helix insertion can causes membrane bending

What are some extrinsic methods of membrane bending?

Protein can be partly hydrophobic and it can bind to the hydrophobic ER membrane and if protein is curved it will pull and bend the membrane, protein clustering can also cause bending as they bind at different sides of each other to curve the membrane, or proteins can be helical and cause bending away from cytoplasm

Can the cytoskeleton cause membrane bending?

Yes

What is clathrin mediated endocytosis involved in?

Vesicle formation

How does clathrin mediated endocytosis work?

A membrane receptor will bind to a lignad and cause a conformational change, the receptor will move into a pit and can now bind to AP2, AP2 attracts clathrin and clathrin will cause membrane bending and vesicle formation, microtubles will then cut off the vesicle at the memebrane, this is extrinsic membrane bending, clathrin must oligimerize to cause bending, multiple clathrins bind each other and bend the membrane

Where does the receptor localize when it binds to cargo? What are the characteristics of this location?

Localizes to a pit and the pit is full of specialized lipids, they have large heads and are highly charged so they spread out and form the pit

How is the vesicle seperated in cathrin mediated vesicle formation?

Dyeinin pinches off the membrane by hydrolyzing GFP

How would you show that dyeinin needs to hydrolyze GTP to pinch off the vesicle?

use a non-hydrolysable form of GTP and see if vesicles still form

How do membranes bend at NPCs?

Intrinsic factors include reticulons, they accumulate at bent membranes and stabilize them, extrinsic bending is due to NUPs forming a coat and bend each membrane

What is responsible for maintaining flat membrane in the NE?

The LINK complex

What stabilizes NPC bending?

Reticulons and amphipathic helices

Why do reticulons sense membrane curvature?

They cause membrane bending but they also bind to bent membranes to stabilize them, we want them to sense specific areas of bending so they only stabilize bending at say an NPC and not a hole where ESCRT3 is repairing

How do reticulons sense membrane curvature?

Reticulons have an amphipathic helix that sense membrane curvature, it is negatively charged on one side and hydrophobic on the other side, when the amphipathic helix is negatively charged it can only bind to curved membranes so this acts as a sensor for the reticulon, amphipathic helix can be positive or negatively charged but reticulons only have negative ones, positive ones can bind to flat membranes and cause curvature

What is the shape of a reticulon?

They are wedge shaped and in duce curvature and also stabilize the membrane

What causes the rough ER to form sheets instead of tubules?

Ribsosmes on the surface stabilize the structure, there are proteins that act as spacers between the sheets and the N term binds to surface proteins and the Cterm oligamerizes with other proteins to help form sheets, reticulons are on OM bending it while proteins like CLIMP are in-between sheets connecting them

What filaments does the ER travel along?

Microtubules

How does the ER travel along microtubules?

The microtubules have adaptors that bind to the ER, the ER doesn’t really travel along it but is made and extended as the microtubule is extended

What are the four types of movement the ER can do along microtubules?

Sliding mechanism, TAC (TIP attachment complex), ring rearrangements, hitchhiking

What is the sliding mechanism?

A new ER tubule is pulled out (made) from an existing one along an existing microtubule

What is the TAC mechanism?

The TIP of the ER tubule is linked to the + end of the microtubule via a protein complex called Tip attachment complex (TAC), the ER tubule will then extend with the microtubule as it polymerizes or it will shirt as the microtubule depolymerizes

What is the ring rearrangement mechanism?

This is a variation of the ER sliding mechanism, this is ER tubules curved into ring structures and move along a microtubule filament, usually they have a mitochondria or endosome inside the ring and this is usually where a mitochondria or endosome will divide (fission)

What is hitchhiking?

This is when an organelle such as a peroxisome will bind to the ER and move with it as the ER moves along a microtubule (this saves energy)

How do new microtubule filaments form?

They bud off of each other (budding) in a branching direction or by homotypic fusion, this is when a microtubule will fuse to another one to create a branch, this requires the cytoskeleton

What is the mechanism of homotypic fusion?

Atlastins

What are atlastins?

Have a long N-terminal GTPase domain, a short linker domain, 3HB domain structure, transmembrane domains and then a C-terminal end that sticks out of the cell) they have roles in membrane fusion

What does the linker domain of atalstin do?

Separates fragments of the protein from each other and brings them close depending on the proteins conformation

How are atlastins similar to reticulins?

They also bind curved membranes, they use curvature sensing through negative charges to bind microtubules and bent membranes

How do atlastins cause membrane fusion?

two atlastins in different memebranes will be close together, the GTPase N-term is close to the 3HB and is tcuked away by the bent linker, when the GTPase domains dimerize they come close and the linkers straighten out (conformational change), the GTPase doamins will move freely and twirl together at the N-term and the linkers will come close together, this pulls the 3HB bundles close together and they fuse, the GTPase doamins then release GDP and the proteins resent now in the same membrane

What is different about the ER domains?

Smooth is tubules and is involved in lipid synthesis, rough is covered in ribosomes

What is the ERES?

These are vesicle budding sites from the ER that go to the golgi

What are MCS (mitochondrial contact sites)?

This is a part of the ER that wraps around mitochondria, mitochondria will require lipids from the ER and they cant be transfered by vesicles so it must be direct, the ER also acts as a scaffold for the mitochondria as they divide

What is the difference between Mitophagy and Autophagy?

Mitophagy is destruction of damaged mitochondria, autophagy is getting rid of unwanted cells

What controls mitochondrial volume in a cell?

Muscle cells have more mitochondria, the ER send fission (more) and fusion (less) signals for division, the lipids secreted by Er to mitochondria are incorperated in the surface and they attract proteins, if you wnat fission they will attract dyenin

What promotes fission of mitochondria?

Mitochondria will make Cdk proteins that attract ER lipids aswell as phosphorylate Drp1 that binds proteins to recruit dyeinin

What is the relationship between the ER and peroxisomes?

Peroxisomes are in the ER framework, peroxisomes are tehtered to the ER with persoxisome and ER specific proteins, the thethering involves proteins from both organelles (MSP major sperm protein) for peroxisome and FFAT on the ER, cutting the tethering will also stop material transfer