Endoplasmic Reticulum

What is the function of the endoplasmic reticulum?

Sites for protein and lipid biosynthesis

What is the form of the endoplasmic reticulum?

• Single membrane bilayer organelle, the largest in eukaryotic cells (typically over 50% of total membrane of an average animal cell)

• Exist as tubules and sheets that are interconnected as an extensive network stretching throughout the cell

• Membrane-bound tubules and sheets enclose a single internal space known as the ER lumen (occupied >10% cell volume)

What are the functions of the rough ER?

• Protein biosynthesis

• Protein target and translocation into the ER

What are the functions of the smooth ER?

• Lipid biosynthesis

• Contain ER exit site, involved in the ER-Golgi trafficking

What is the form of the rough ER?

Exists as interconnected layers of flattened sheets

What is the form of the smooth ER?

Exists as tubules

What happens when cells are homogenized to the rough ER?

Breaks up into small microsomes

• Still functional (ribosomes are still on)

• Easily purified (density is greater than other membranous organelles)

Is the morphology of the ER in an animal and plant cell similar or different?

Morphology is very similar - the organelle is well conserved in eukaryotes

Is the ER dynamic or stagnant?

Consistently being reorganized: some connections being broken, while new ones being reformed

How are the ER tubules moved?

Pulled out and moved on microtubules by motor proteins then fuse with another one to make a network

What are the two models for ER motility on microtubules?

1. Growth with the tip on the + end with the +TIP proteins

2. Slide along microtubules with a molecular motor

What is the function of reticulons?

Generate the curvature necessary formation of ER tubules and edges of ER sheets

What ais the forms of reticulons?

W-shaped structure

What is the function of Atlastins?

• Dynamin-like large GTPases that undergo a GTP-dependent oligomerization important for fusion of different tubules

• Have a transmembrane domain and they are large proteins

• Brings together the two tubules

What is the function of CLIMP63?

Serves as an ER luminal spacer for sheet formation - to space it out

How are proteins targeted to the ER?

Targeted to the ER together with ER-bound ribosomes

Where does the ER-bound ribosomes end up?

Within the ER lumen

When microsomes are treated with detergent and a protease is added what happens?

Digestion of secretory protein because the detergent disrupts the membrane

How are secretory proteins targeted to the ER?

ER signal sequence - once emerged guides the ribosome-mRNA -nascent peptide complex to the ER

What happens when the ribosome-mRNA-nascent peptide complex is properly attached to the ER?

The peptide is then translated and translocated into the lumen of the ER → Co-translational

How many proteins are cytosolic? HOw are they different than the ones targeted to the memrbrane?

>50% are cytosolic and do not need to be secreted many proteins are targeted to the ER for secretion. Cytosolic proteins do not have the ER signal sequence if the signal is not present, it will remain cytosolic

What is the ER signal sequence?

Around 25 ss long stretch of hydrophobic amino acids

How were co-translational ER signal sequence in secretory proteins discovered?

• Cell-free protein synthesis; no microsomes present

• Cell-free protein synthesis; microsomes present

• Without signal peptide, microsomes present

• Without the N-terminal signal peptide, no incorporation→ the signal sequence is necessary for ER import

• Imported proteins had their N-terminus signal peptide → ER signal sequence is cleaved off

• Import only occurred when microsomes were present while synthesis took place → co-translational

What is the signal sequence first recognized by?

Signal recognition particle (SPP)

What does SRP do?

Docks the nascent polypeptide-ribosome-mRNA onto the ER by its interaction with SRP receptor

What happens after the nascent polypeptide-ribosome-mRNA is docked?

The translocon closes

What happens once the nascent polypeptide-ribosome-mRNA is properly docked?

Both P54 of SRP and the alpha subunit of the SRP receptor hydrolyze their bound GTP that destabilizes the interaction interface so they dissociate. Also, ER signal sequence opens the translocation and enters the channel

What happens once the polypeptide translocates into the ER lumen as mRNA is translated?

• The ER signal sequence is cleaved off by the signal peptidase then degraded

• Once translocation completes: 1) The translocator closes 2) The ribosome is released and 3) The translocated protein is folded within the lumen of the ER