Intracellular membrane traffic

Question 1

What molecules are delivered by exocytosis? Where are they transported to?

Answer 1

This secretory pathway delivers newly synthesised proteins, carbohydrates and lipids either to the PM or the intracellular space.

Question 2

What molecules are delivered by endocytosis? Where are they transported to?

Answer 2

Cells remove PM components and deliver them to components called endosomes, from where they can be recycled to same or different area of the PM or to be delivered to the lysosome for degradation.

Question 3

What are inner coat vesicles used for?

Answer 3

Needed to concentrate proteins present in the donor membrane so that vesicles which will be produced have high concentration of specific cargo.

Question 4

What are outer coat vesicles used for?

Answer 4

Needed for giving shape to the vesicle, usually a basket like lattice.

Question 5

Describe the structure of clathrin.

Answer 5

Each subunit consists of 3 large and 3 small polypeptide chains that together form a structure called a triskelion.

Question 6

What are the 3 main types of vesicles involved?

Answer 6

Clathrin, COPI, COPII

Question 7

Describe the process of clathrin-dependent vesicle budding.

Answer 7

A molecule is to be transported from the donor membrane. The cargo has to be loaded inside a clathrin coated vesicle to then be transported somewherreelse.

A cargo receptor is needed, as well as an adaptor protein called adaptin. This forms a complex with the cargo, where the inner layer is formed with the adaptor protein while the outer layer is formed by the triskelions of clathrin.

The membrane is budding until a complete vesicle is formed. This is then ready to travel to the target location. Once it arrives, it will shed away its adaptins and clathrins and fuse with the target membrane.

Question 8

What are surface markers?

Answer 8

High target specificity is obtained as all transport vesicles have a specific surface marker that identifies them, this tells the system their origin and type of cargo.

Question 9

How do target membranes recognize vesicles?

Answer 9

Target membranes show on their surface complementary receptors, which will recognise the surface markers on vesicles.

Question 10

What are the functions of Rab effector proteins?

Answer 10

Rab effector proteins are also called tethering proteins due to their function of docking the vesicle to the target membrane.

Question 11

Describe the connection between Rab GTPase and vescicles.

Answer 11

The Rab GTPase is located on the vesicle in active form as it is bound to GTP.

Question 12

Describe the ‘tethering’ part of vesicle and membrane fusion.

Answer 12

The first connection between the target membrane and the vesicle is produced due to the tethering proteins which are the effectors.

Question 13

Describe the ‘docking’ part of vesicle and membrane fusion.

Answer 13

The t-SNARE and v-SNARE pair to produce a SNARE complex. The docking of the vesicle is almost complete and fusion can take place.

This can only happen when GTP bound to Rab protein is hydrolyzed, it is released after this.

Question 14

Describe the ‘fusion’ part of vesicle and membrane fusion.

Answer 14

To achieve a membrane fusion, the bilayers of the 2 membranes need to be within 1.5 nm of each other.

The layer of water in between 2 membranes needs to be displaced, so fusion proteins that can overcome energy barriers are needed.

This is carried out by SNARE proteins, which use energy arriving with the vesicle to fuse the 2 membranes. Otherwise this will not fuse, as the presence of water is highly energetically unfavourable.

Question 15

What are COPII vesicles used for?

Answer 15

COPII coated vesicles are used to transport cargo from the ER to the GA.

Vesicles are budded and coated with the proteins, there are specific proteins exposed on the inner layer of these vesicles.

Question 16

How does the intermediate vesicular structure move from the ER to the cis golgi network?

Answer 16

The intermediate vesicular structures move from the ER to the cis golgi network by using ‘rails’ made of microtubules.

COP coated vesicles fuse to form vesicular tubular clusters, which travel in the cytoplasm toeards the cis golgi network using rails of microtubules.

Question 17

Describe the general structure of the Golgi Apparatus.

Answer 17

The GA is made of an ordered series of compartments. There is the cis face, which faces the inner part of the cell, while the trans faces faces outwards at the membrane.

In between is the cisterne which can be classified into the cis, medial or trans cisterne.

Question 18

Where are oligosaccharides added?

Answer 18

The first step of addition of oligosaccharide to proteins happens in the ER.

Question 19

Where are oligosaccharides processed?

Answer 19

The processing of oligosaccharides occurs in the GA.

Each site in the GA is used for a different step of this processing and each type of cisterna has its own mix of enzymes that carries out its function.

Question 20

What are oligosaccharides transformed to in the GA?

Answer 20

Transformed to either complex oligosaccharides or high mannose oligosaccharides.

Question 21

How do N-linked glycosylations promote protein folding?

Answer 21

Renders intermediates are more soluble, thereby preventing aggregation.

Establishes a glyco-code which marks the progression of the protein folding, as well as mediating the binding to chaperones.

Question 22

What else can N-linked glycosylation do apart from protein folding?

Answer 22

It also helps protect the protein from digestion by enzymes. Specific oligosaccharide trees exposed on proteins can modify a protein’s antigenic and functional properties.

Question 23

What are lysosomes?

Answer 23

Lysosomes are membrane enclosed organelles filled with specific hydrolytic enzymes, as their function is to digest macromolecules.

Question 24

Why can’t hydrolytic enzymes survive outside lysosomes?

Answer 24

All these enzymes are active because they are activated by the presence of acidic pH. If by chance all these enzymes are lost by the lysosomes, they will not be active as the pH outside will be neutral.

Question 25

How is the pH inside lysosomes maintained?

Answer 25

The pH pump (ATPase) pumps H+ inside lysosomes to maintain its pH.

Question 26

How are lysosomes derived?

Answer 26

Lysosomes are derived from endosomes. Late endosomes contain material derived from the PM by endocytosis as well as enzymes.

These late endosomes fuse with preexisting lysosomes to form structures called endolysosomes, which then fuses with each other.

Question 27

How does phagocytosis work?

Answer 27

Specific phagocytic cells called macrophages and neutrophils ‘ eat’ large particles. resulting in a phagosome.

Question 28

How does phagocytosis work?

Answer 28

Specific phagocytic cells called macrophages and neutrophils ‘ eat’ large particles. Resulting in a phagosome.

Question 29

What are the functions of macropinocytes?

Answer 29

Macropinocytes specializes in the nonspecific uptake of fluids, membranes and particles attached to the PM.

Question 30

Why is autophagy needed?

Answer 30

When cells differentiate, they need to change their shape and contents.

In some cases autophagy responds to environmental stimuli.

Autophagy can remove large molecules that are too large to be degraded by the proteasome. Otherwise diseases can be caused by malfunction of autophagy, like neurodegeneration and cancer.

Question 31

How does autophagy work?

Answer 31

Something induces autophagy, the parts that need to be degraded are engulfed in membranous structure called autophagosomes, which then fuses with the lysosome where the acid hydrolases are present.

Then the acid lysosomes degrade the contents of the autofibersomes.

Question 32

Describe selective autophagy.

Answer 32

Something chosen to be engulfed in the autophagosome and digested.

In this case the portion of cytosome contained in the autophagosome is not large, the shape usually reflects the contents. This is induced by specific needs of the cell.

Question 33

Describe non-selective autophagy.

Answer 33

A portion of the cytoplasm is engulfed in autophagosome, e.g. in stressful conditions when nutrients are lacking, the cell digests itself.

Question 34

What happens when lysosomes don’t work?

Answer 34

When lysosomes don’t work, this results in pathological consequences. The most severe is in the nervous system.

One or more than one gene encoding for a lysosomal hydrolase is mutated. Therefore cells parts unwanted material as they can’t digest it.

Question 35

Describe I cell disease.

Answer 35

A rare inherited metabolic disorder where almost all of the hydrolytic enzymes are missing from the lysosomes of many cell types.

Material is accumulated in the cells, resulting in large inclusions. Individuals with this condition rarely live beyond 6 or 7 years.

Question 36

Describe endocytosis.

Answer 36

First, vesicles fuse with the early endosome. The decision for the destination of the cargo is taken here. These vesicles stay this way inside then endosomes, as long as they are inside the bodies, they stay as vesicles. One possible destination is the lysosome.

The vesicles formed at the membrane can be clathrin formed vesicles. Some of them form a pit before budding away to form a vesicle. Many of these vesicles are continually produced from the PM and are very short-lived.

Question 37

Describe features of macropincytosis.

Answer 37

This is not dependent on clathrin.
This is not continuous and is induced in response to something.
Therefore there is a receptor on the cell membrane which is reached by a factor, this receptor activates a response which is the macropinocytosis response.

Question 38

Describe the process of macropinocytosis.

Answer 38

When the ligand binds to the receptor, it activates a signalling pathway which changes the state of the cytoskeleton, which means it changes acting dynamics and produces protrusions of the cell surface called ruffles. These ruffles form and collapse on the cell membrane, thus enclosing in vesicles being formed.

Macropinocytosis is a dedicated degradative pathway. There is an acidic pH which allows fusion with late endosomes, and from then on to lysosomes. Therefore what is contained in macropinosomes is sent to degradation. Although viruses’ genomes are not degraded.

Question 39

Explain consitutive secretory pathway.

Answer 39

Works continuously without the use of signal.

Question 40

Explain regulated secretory pathway.

Answer 40

A signal is needed (usually ligand binding to receptor) to switch on the signalling pathway.

Question 41

What is a third pathway between the 2 secretory pathways?

Answer 41

There is a third one that doesn’t end up at the PM and instead goes to lysosomes. This always occurs starting from the trans golgi network.

Question 42

Describe how clathrin works in forming secretory vesicles.

Answer 42

When passing through the trans golgi network, clathrin will assemble in some areas and change the shape of the membrane in that part.

Then an immature secretory vesicle is formed, the concentration of protein is high and becomes even higher due to the surface area of the secretory vesicle.

The final mature secretory vesicle is small and has lost volume, therefore the cargo had been concentrated with no clathrin. So clathrin is used to concentrate the secretory vesicle.

Question 43

How can the precursor proteins stored in vesicles be activated?

Answer 43

Proteolysis

A part can be cleaved off. These precursors are called pre pro-proteins. In some cases there are 2 parts that need to be cleaved, the signal peptide and another peptide.