Advanced Molecular Cell Biology - Lecture 14

mutations in Sec23A cause changes in ER ___________

morphology

Changes in the morphology of the ER can be confirmed by?

electron microscopy, which allow luminal expansions to be observed

binding of ______ Sec23A to liposomes is unaffected as long as GTP-bound Sar1 is present on the liposome, therefore it was found via assay that the actually issue lies within _____ _________

mutant, coat recruitment

Sec23 F382L is not effective at recruiting?

coat

Patients with CLSD do not have problems with digestion or insulin secretion. Why are only some tissues affected?

because they have other components that they can use for COP2, and there are other paralogs for Sec23/24 present in other tissues

what kind of cargo would be affected in CLSD patients?

extracellular matrix, i.e. collagen

how is collagen packaged?

transported in carriers which are tubulated versions of COP2 vesicles (the coat that covers this cargo is similar to smaller cargo coats)

electron microscopy showed _______ sites that are _______ dense in the sarcoplasmic reticulum

contact, electron

many different organelles are in contact with each other via?

membrane contact sites

membrane contact sites provide platforms for:

• intracellular signalling

• lipid metabolism

• organelle biogenesis

• organelle division

• motor protein-mediated membrane dynamics

In the case of membrane contact sites, what is there no evidence of?

membrane insertion or fusion

ER contact sites occur where?

in the smooth ER (SER)

describe the structure of ER contact sites:

• Ribosomes excluded from contact sites

• Membranes very close (3-15nm)

• ER contacts can be short- or long-lived

how do membrane contact sites participate in intracellular signalling?

exploit differences in calcium concentration in different intracellular organelles

describe what the sarcoplasmic reticulum of muscle cells is:

Specialised ER for handling Ca2+ transients required for muscle contraction

when the signal from the motor neuron axon arrives at the neuromuscular junction what happens?

There is a signal at the cell surface mediated through T-tubules, resulting in the release of calcium from intracellular stores; this allows muscle proteins to participate in muscle contraction.

After muscle contraction, how does the ER replenish its intracellular calcium stores?

• mediated by contact sites

• STIM1 is a monomeric calcium sensor distributed throughout the ER

• when it detects low calcium concentrations, via its calcium-binding domain, it forms an oligomer

• it then migrates to certain areas where the ER is in contact with the plasma membrane; these areas are enriched in PIP2

• STIM1 oligomer associates with Orai channels in the plasma membrane; this allows uptake of extracellular calcium

endosomes can _____ the ER

drag

why is the pulling of ER via endosomes important?

Functionally important in EGFR (epidermal growth factor receptor, a type of tyrosine kinase) trafficking and signalling, the membrane contact site with the early endosome helps regulate EGFR movement into multivesicular bodies. PTP1B: protein tyrosine 1B (located in the ER membrane) dephosphorylates EGFR, switching it off and inactivating it, to make sure it is degraded in lysosomes by being incorporated into a multivesicular body.

membrane contact site proteins often form ______ linking the 2 membranes

bridges

For the MCS proteins that bridge linking 2 membranes to be brought together, you need to have?

a membrane anchored protein with a very big cytoplasmic domain

describe membrane contact sites in lipid transfer:

• ER is the site of membrane lipid synthesis

• Contact sites provide non-vesicular transfer of lipid

• Lipid transfer is unidirectional

• Lipid transfer proteins may use concentration gradients of lipid to promote lipid transfer

what are the ways that membrane contact sites facilitate lipid transfer between organelles?

• MCS protein, which has a hydrophobic groove and can extract a lipid from the donor membrane, flips around and delivers it to the ER

• A hydrophobic conduit can deliver a lipid from the donor membrane to the ER membrane

• A lipid transfer protein can move cholesterol from the ER to the Golgi, moving from an area of low to high concentration. A molecule of phosphoinositide (in abundance in the Golgi) is moved to the ER; it is exchanged for a cholesterol molecule (PI4P is kept in low concentration in the ER by being converted rapidly into PI in the ER)

defects in MCS give rise to ________

disease

what is niemann pick disease C?

An accumulation of cholesterol in the lysosomes

• When cholesterol is taken from the diet, it stops the cell from producing cholesterol; therefore, cholesterol can end up in the endosome or lysosome. There are membrane contact sites which are important in the transfer of cholesterol into the ER; if defective, this disease can be a result

• knock-on effects include neurological defects

describe how membrane contact sites allow mitochondrial fission:

MCS is often the ER winding itself around the mitochondria, enabling it to divide into 2. This is important for cell division and mitochondrial function in interphase cells and when the cells divide