Cell Biology Exam 3

Some secretory proteins and some integral membrane proteins must pass through a membrane. A ribonucleoprotein known as the signal recognition particle (SRP) guides these proteins either to the plasma membrane (in bacteria) or the endoplasmic reticulum (in eukaryotes). What are the steps of the membrane translocation of eukaryotic secretory proteins?

1. The signal peptide exits the ribosome during translation.

2. SRP binds to the signal peptide and translation stops.

3. SRP moves the ribosome to the ER membrane.

4. Ribosome docks to a translocon.

5. Translation resumes.

6. The growing polypeptide crosses the ER membrane through translocon.

7. Signal peptidase removes the signal peptide from the growing polypeptide.

8. The remainder of the polypeptide is moved into the ER lumen.

9. Protein is transported out of the ER.

Protein X is an unknown membrane protein found in erythrocytes that can be extracted primarily from plasma membranes using a concentrated salt solution. What type of membrane protein is protein X?

Since protein X is extracted from the membrane by altering the ionic strength of the extraction solution, it is a peripheral membrane protein.

To determine the location of protein X in the plasma membrane, erythrocytes were subjected to two different treatments. First, intact erythrocytes were treated with proteolytic enzymes. After the cells were lysed and membrane components extracted, protein X was intact. In the second experiment, the erythrocyte cells were permeabilized prior to treatment with proteolytic enzymes. When the plasma membrane components were then extracted, protein X was extensively fragmented. What do these observations suggest about the location of protein X in the plasma membrane?

Since proteolytic enzymes cannot fragment protein X in intact erythrocytes, protein X is located inside the cell and interacts with the inner surface of the membrane.

Proteins imported to what organelle do NOT have their signal sequence cleaved?

Peroxisomes

In what way does peroxisomal protein import differ from mitochondrial protein import?

Ubiquitination is a key step in receptor recycling.

Where would you find the precursor of a mitochondrial matrix protein in the following types of mitochondrial mutants:

(a) a mutation in the Tom22 signal receptor;

(b) a mutation in the Tom70 signal receptor;

(c) a mutation in the matrix Hsp70;

(d) a mutation in the matrix signal peptidase?

(a) cytosol;

(b) cytosol;

(c) inner membrane;

(d) matrix

What is true of secretory protein translocation?

1. Type I membrane proteins contain two signal sequences; one targets the protein to the ER and one is a stop transfer anchor sequence.

2. Type IV membrane proteins contain a mixture of both stop transfer anchor sequences and signal anchor sequences, the order of which determines the protein's orientation in the membrane.

3. Hydropathy profiles are used to computationally identify secreted proteins.

Which of the following are required for cotranslational translocation of a secretory protein into the ER?

1. hydrophobic signal sequence

2. signal recognition particle

3. signal recognition particle receptor

4. GTP

5. translocon

What is the source of energy needed for unidirectional translocation across the membrane in

(a) cotranslational translocation into the endoplasmic reticulum (ER); (b) post-translational translocation into the ER; and

(c) translocation into the mitochondrial matrix?

(a) GTP hydrolysis;

(b) ATP hydrolysis;

(c) ATP hydrolysis

As part of an early experiment on the secretory system, an in vitro translation system consisting only of mRNA and ribosomes resulted in secretory proteins that were larger than the identical protein when translated in a cell. Why?

Signal sequence cleavage requires ER entry and action of ER-resident proteins.

Proteins have their signal sequence cleaved when they are imported to which organelles?

chloroplasts, mitochondria, and endoplasmic reticulum

What are important ER-resident enzymes that modify proteins in the secretory pathway?

1. oligosaccharyl transferase,

2. protein disulfide isomerase, and

3. BiP

Which molecular mechanisms are required for the unfolded protein response?

1. competitive binding,

2. nuclear import, and

3. endonuclease activity

Nuclear import of proteins from the cytoplasm occurs via _______ using the nuclear cargo protein _______.

nuclear pore complexes (NPC); importin

The function of FG-nucleoporins within the nuclear pore complex is to:

shield large, hydrophilic, unchaperoned proteins from entering the nucleus.

Release of the cargo protein from its nuclear cargo protein into the cytoplasm is triggered by:

the binding of importin to Ran-GTP.

Ultimately, the entire process of nuclear protein import is driven by:

GTP hydrolysis

Preventing GEF activity in the nucleus would prevent:

the exchange of GDP for GTP on Ran

What can be shared by both secreted soluble proteins and plasma membrane proteins that are synthesized by ribosomes docked on ER translocons?

an N-terminal ER signal sequence

Which of the following are activities of a Signal Recognition Particle (SRP)?

1. arresting protein translation,

2. binding an ER signal sequence, and

3. interacting with an SRP receptor to dock the ribosome on a translocon

What is the final topology of a protein in the ER that is synthesized with an N-terminal signal sequence and one stop-transfer anchor sequence?

N-terminus in the ER lumen, one transmembrane domain, C-terminus in the cytosol

If a mutation is introduced into I-ĸBα that inhibits its phosphorylation by the IKK complex, this would prevent:

translocation of NF-ĸB into the nucleus.

Exocytosis is one of the processes living organisms use to move molecules. What are examples of exocytosis?

1. A nerve cell releases peptides used as messengers into the extracellular space;

2. A single‑celled eukaryotic organism excretes digestive waste into the surrounding water;

3. Pancreatic cells release digestive enzymes into the small intestine.

Which situations involve a cell using endocytosis?

1. A white blood cell engulfs a bacterium, brings the bacterium inside the cell, and uses digestive enzymes to destroy the bacterium.

2. A cell wraps a portion of its membrane around a droplet of nutrient‑rich water and brings the droplet inside the cell.

3. LDL cholesterol attaches to cell membrane proteins, triggering the membrane to fold around the cholesterol and bring it into the cell.

What is the correct order for secretory protein synthesis and export from the cell? Meaning once a secretory protein is made, what path will it take as it is released from the cell?

ribosome ->

rough ER ->

transport vesicle ->

Golgi apparatus ->

transport vesicle ->

plasma membrane

What is clathrin?

peripheral membrane protein that mediates endocytosis

Which statements about SNARE‑mediated membrane fusion are correct?

1. The lipid bilayers of the fusing membranes undergo rearrangement involving changes in membrane curvature.

2. SNAREs from both the cell plasma membrane and the vesicle combine to form a complex that drives membrane fusion.

Wolman disease is a lysosomal storage disease that results in the buildup of lipids (fats) in the spleen, liver, bone marrow, small intestine, adrenal glands, and lymph nodes. What is the probable cause?

A gene mutation results in a shortage of lipid‑digesting enzymes in the lysosome.

Rheumatoid arthritis (RA) is an autoimmune disease marked by inflammation and cartilage destruction. Although the causes are not known, it may be linked to lysosomal proteases (protein‑digesting enzymes). How may lysosomal proteases be linked to RA?

Excess lysosomal proteases are produced and released extracellulary, where they degrade collagen.

What mechanistic features are shared by the formation of multivesicular endosomes via budding into the interior of an endosome and the outward budding of HIV virus vesicles at the cell surface?

ubiquitinylated membrane proteins

What are some differences between phagocytosis and autophagy?

1. Phagocytosis is specific to a few cell types, while autophagy is a common cellular degradation mechanism.

2. Only one of the two processes is a type of endocytosis.

3. Autophagy degrades intracellular material, while phagocytosis degrades extracellular material.

What is true about LDL internalization?

1. Endosomal pH is crucial for dissociation of LDL from its receptor after endocytosis.

2. The LDL receptor transits the secretory pathway.

3. Mutations in the LDL receptor cause familial hypercholesterolemia.

How does lysosomal pH contribute to lysosomal protein sorting?

The mannose-6-phosphate receptor has altered affinity for M6P under acidic pH conditions.

What are some facts about apical and basolateral protein sorting?

1. GPI-linked proteins are sorted specifically to the apical membrane of most polarized epithelial cells.

2. Transcytosis is a mechanism for the movement of basolaterally secreted proteins to the apical membrane.

3. Infection of epithelial cells with specific viral proteins has helped elucidate the mechanisms of polarized protein secretion.

Which of the following statements regarding clathrin-coated vesicles are TRUE?

1. Adapter complexes provide the specificity of vesicle cargo.

2. Dynamin is required for pinching off clathrin-coated vesicles.

3. Clathrin-coated vesicle assembly is mediated by the GTPase ARF.

Which of the following statements are TRUE regarding signal sequences?

1. The di-arginine sorting sequence can be located anywhere in the cytoplasmic domain of an ER-resident protein.

2. The KDEL receptor contains a C-terminal Lys-Lys-X-X sequence.

3. A protein with a KDEL sequence localizes to the ER via COPI retrieval.

Which of the following statements about the function of a vesicle fusion protein is correct?

Rab proteins are responsible for docking of vesicles on target membranes.

Which of the following statements is TRUE regarding COP vesicle budding?

1. COPI and COPII vesicles are composed of completely distinct coat proteins.

2. Both Sar1 and ARF undergo conformational changes to insert into the ER or Golgi membranes in order to carry out their functions in vesicle assembly and budding.

3. COPII vesicles travel from the endoplasmic reticulum to the cis-Golgi.

What are some techniques commonly used for the detection of secretory proteins to follow their movement within the cell?

1. Western blotting

2. microscopy of fluorescently labeled protein

3. pulse-chase labeling and autoradiography

The KDEL sequence on the C-termini of ER luminal proteins is necessary for:

retrieval of the protein back to the ER.

The KDEL receptor is expressed in which of the following?

COPI and COPII vesicles and the cis-Golgi network

The binding affinity of the KDEL receptor to KDEL sequences is dependent on:

pH only

If the pH of the rough ER were higher than normal, what might be expected of the fate of proteins bearing the KDEL sequence?

They would not be released from the KDEL receptor of vesicles returning to the ER.

_______ protein complexes are responsible for _______ transport from the ER to cis-Golgi.

COPII; anterograde

Fusion of a vesicle to the membrane of either the ER or cis-Golgi is mediated directly by:

SNARE protein interactions on both the vesicle and ER/cis-Golgi membranes.

What molecules are involved in vesicle formation in anterograde vesicle transport?

1. COPII coat proteins

2. V-SNARES

3. membrane cargo proteins

4. soluble cargo proteins

Retrograde vesicle transport, mediated by COPI coat proteins, serves the function of:

recycling certain proteins from the cis-Golgi to ER.

Where in the cell would you expect to find a protein that contains the following amino acid sequence within its polypeptide chain Proline-Lysine-Lysine-Lysine-Arginine

the nucleus

The figure below shows the topology of an ER membrane protein. If inserted into the cell membrane, the N-terminus of this protein will be

on the cytosolic side

proteins are targeted to the endoplasmic reticulum through:

signal sequences in the polypeptide

Which organelles are bound by two membranes?

mitochondria AND the nucleus

You fuse a mitochondrial matrix targeting sequence to the cytosolic protein DHFR to study if the sequence can target a non-mitochondrial protein to the mitochondria. DHFR is transported to the mitochondria when it is fused to the targeting sequence. However, when you treat the cells with a drug that forces DHFR to fold into its native conformation, DHFR is no longer able to get into the mitochondria. Why not?

The protein must be unfolded to translocate through the TOM complex

Proteins translocation to the mitochondria occurs:

Post-translationally

What type of ribosomes are responsible for synthesizing proteins that are transported through the TOM complex?

Free cytosolic ribosomes

proteins are translocated into the nucleus as:

completely folded polypeptides

T/F: The nuclear pore complex is a hereto-dimer composed of nuclear export protein and Ran GTPase.

FALSE, it is made of nucleoporins and is not a hereto-dimer

Which of the following strategies would allow you to follow VSV-G over time as it moves along the secretory pathway in live cells?

Express GFP-tagged VSV-G in cells and use fluorescence microscopy

Deletion of the gene X results in accumulation of secretory proteins in the Golgi. Deletion of the gene Y results in accumulation of the protein at ER. Where would you expect secretory proteins to accumulate in the double mutant that lacks both X and Y ?

the endoplasmic reticulum

COPII-coated vesicles mediate the transport of cargo proteins from:

the ER to the Golgi

What sorting signal directs soluble lysosomal proteins from trans Golgi to the lysosomal lumen?

Mannose 6 phosphate

Ubiquitination is an example of:

post-translational protein modification