Cell Biology Exam 1 Notes

Cell Biology Exam 1 Notes

Multiple Choice Questions

Question 1

• Question: Which statement best describes the function of a promoter in an expression vector?
• (a) It controls the initiation of transcription for the inserted gene
• (b) It ensures the plasmid is stably maintained in the host cell
• (c) It provides antibiotic resistance to allow for selection
• (d) It enhances the replication rate of the plasmid
• Explanation: Promoters are DNA sequences that define where transcription of a gene by RNA polymerase begins. In expression vectors, the promoter drives transcription of the inserted gene.

Question 2

• Question: Which of the following statements is not a principle of cell theory?
• (a) All living organisms are composed of one or more cells
• (b) The cell is the basic unit of structure and function in living organisms
• (c) All cells arise from pre-existing cells
• (d) Cells can form spontaneously from non-living materials under normal conditions
• Explanation: One of the key tenets of cell theory is that cells only arise from pre-existing cells. Spontaneous generation has been disproven.

Question 3

• Question: Which of the following statements correctly differentiates Western blot analysis from immunostaining?
• (a) Western blot requires protein separation by gel electrophoresis, whereas immunostaining detects proteins in their original cellular context
• (b) Immunostaining is used to detect specific proteins, while Western blot detects all proteins in a sample
• (c) Western blot can only be performed on live cells, whereas immunostaining requires lysed cells
• (d) Immunostaining is more quantitative than Western blot because it measures absolute protein concentration
• Explanation: Western blots involve separating proteins by size using gel electrophoresis, transferring them to a membrane, and then using antibodies to detect a specific protein. Immunostaining detects proteins in cells or tissues in their native context.

Question 4

• Question: Which of the following would be most likely to result from a mutation that reduces the GTPase activity of a G protein?
• (a) The G protein would remain in the GDP-bound inactive state, preventing its function
• (b) The G protein would stay active for a prolonged period
• (c) The G protein would rapidly exchange GDP for GTP, causing constitutive activation and depletion of GTP
• (d) The G protein would not be affected by the mutation
• Explanation: G proteins are active when bound to GTP and inactive when bound to GDP. GTPase activity hydrolyzes GTP to GDP, inactivating the G protein. Reduced GTPase activity would cause the G protein to remain in the active, GTP-bound state for longer.

Question 5

• Question: Signal sequences that direct proteins to ER or nucleus are _.
• (a) added to proteins through post-translational modification.
• (b) added to proteins by a protein glycosylation.
• (c) encoded in the amino acid sequence in a protein
• (d) always removed once a protein is at the correct destination.
• Explanation: Signal sequences are specific amino acid sequences within a protein that target it to a particular cellular location, such as the ER or nucleus.

Question 6

• Question: Which statement is correct regarding the nuclear localization sequence (NLS) of a nuclear protein?
• (a) It is a hydrophobic sequence that enable the protein to bind to the nuclear pore complex.
• (b) It has high binding affinity for Ran-GTP
• (c) It prevents protein folding until the protein is thread through nuclear pores.
• (d) It dissociates from importin in the presence of Ran-GTP
• Explanation: The NLS is recognized by importin, which facilitates the protein's entry into the nucleus. In the nucleus, Ran-GTP binds to importin, causing it to release the cargo protein.

Question 7

• Question: How does phosphorylation commonly regulate nuclear import?
• (a) It always enhances the binding of importins to the nuclear localization signal (NLS)
• (b) It can either promote or inhibit nuclear import by modulating NLS accessibility
• (c) It causes proteins to be rapidly degraded, preventing nuclear import
• (d) It permanently sequesters proteins in the cytoplasm
• Explanation: Phosphorylation can change the conformation of a protein, either exposing or masking the NLS, which affects its ability to interact with importins and thus regulate nuclear import.

Question 8

• Question: Your friend works in a biotechnology company and has discovered a drug that blocks the function of Ran-GAP. Which step within the nuclear transport is mostly likely be affected by the drug?
• (a) Importin binding to cargo proteins.
• (b) Importin exiting the nucleus.
• (c) Importin releasing the cargo in the nucleus.
• (d) Importin's interaction with nuclear pore proteins
• Explanation: Ran-GAP is located in the cytoplasm and hydrolyzes GTP bound to Ran, creating a high concentration of Ran-GDP in the cytoplasm. This gradient is essential for nuclear transport. Blocking Ran-GAP would disrupt this gradient and affect the ability of importin to release its cargo in the nucleus and exit.

Question 9

• Question: If you isolate a nuclear protein, then inject it into to the cytosol, it is likely to
• (a) remain in the cytosol until it is degraded.
• (b) be taken up into the nucleus
• (c) be secreted through channels in the plasma membrane.
• (d) be taken up by the ER, concentrated in secretory vesicles, and transported to the nucleus.
• Explanation: Nuclear proteins contain a nuclear localization signal (NLS) that will be recognized by importins, leading to their active transport into the nucleus.

Question 10

• Question: Which of the following organelles is not part of the endomembrane system?
• (a) the Golgi.
• (b) the early endosome.
• (c) the plasma membrane
• (d) the peroxisome
• Explanation: The endomembrane system includes the ER, Golgi, endosomes, lysosomes, and plasma membrane. Peroxisomes are not part of this system; they import proteins post-translationally.

Question 11

• Question: Which of the following statements about proteins inserted in the ER lumen during co-translational import is false?
• (a) Some of these proteins may be taken up by the Golgi
• (b) Some of these proteins may be inserted in the lysosome membrane
• (c) Some of these proteins may be packaged into COPII coated vesicles.
• (d) Some of these proteins may be secreted out of the cell.
• Explanation: Proteins that enter the ER lumen can proceed through the Golgi and be secreted or end up in the lysosome lumen, but they don't typically become integral membrane proteins of the lysosome.

Question 12

• Question: Which of the following organelles does not receive proteins by posttranslational import of proteins synthesized by cytoplasmic ribosomes?
• (a) Lysosome
• (b) Nucleus
• (c) Mitochondria
• (d) Peroxisomes
• Explanation: The nucleus imports proteins post-translationally. Proteins destined for the lysosome arrive via the endomembrane system and are not directlyimported from the cytoplasm after translation. Mitochondria and peroxisomes import proteins post-translationally.

Question 13

• Question: Imagine a protein that has been engineered to contain a nuclear localization signal on the N-terminus of the protein and an ER signal sequence in the middle of the protein. With both signals present on the same protein, where would you expect to find the protein after its synthesis?
• (a) the nucleus
• (b) the endoplasmic reticulum (ER)
• (c) it will first enter the nucleus then recognized by the SRP and enter the ER
• (d) the cytosol
• Explanation: The ER signal sequence will be recognized first during translation, leading the ribosome to the ER membrane for co-translational import. Thus, the protein will end up in the ER.

Question 14

• Question: Proteins that are fully translated in the cytosol and lack a protein sorting (targeting) signal will end up in ____.
• (a) the cytosol.
• (b) the interior of the nucleus.
• (c) the mitochondria.
• (d) secreted out of the cells
• Explanation: Proteins lacking a specific targeting signal remain in the cytosol after translation.

Question 15

• Question: After isolating the rough endoplasmic reticulum from the rest of the cytoplasm, you purify the RNAs attached to it. Which of the following proteins is not likely to be encoded by the RNA?
• (a) Insulin
• (b) Insulin receptor
• (c) BiP
• (d) Sar1
• (e) Lysosome H+ pump
• Explanation: Insulin, the insulin receptor, BiP, and Sar1 are all related to the ER and secretory pathway. The lysosome H+ pump is unlikely to be synthesized directly on the rough ER; it is more likely to be synthesized on free ribosomes and then sorted.

Question 16

• Question: Ribosomes that do not anchor on the surface of the rough endoplasmic reticulum during translation most probably
• (a) have a defect in ribosomal proteins that allow attachment to the ER surface.
• (b) have a signal peptidase error.
• (c) are synthesizing cytoplasmic or nuclear proteins.
• (d) do not make the appropriate anchor protein.
• Explanation: Ribosomes synthesizing cytoplasmic or nuclear proteins do not have a signal sequence that directs them to the ER, so they remain free in the cytosol.

Question 17

• Question: What would be the most likely consequence of a mutation that prevents GTP hydrolysis in SRP or its receptor?
• (a) SRP would fail to recognize signal peptides, leading to inefficient protein targeting
• (b) SRP and its receptor would remain bound together, thus sequestering cytosolic SRPs
• (c) SRP would be unable to bind GTP, leading to uncontrolled ribosome targeting to the ER
• (d) SRP would degrade prematurely, preventing its reuse in subsequent protein targeting events
• Explanation: GTP hydrolysis is required for SRP to detach from its receptor after delivering the ribosome to the ER. If GTP hydrolysis is blocked, SRP and its receptor would remain bound, preventing SRP from being recycled.

Question 18

• Question: Consider a transmembrane protein with the topology in the ER… How would the protein topology look like on the plasma membrane after the modification?
• (a) [Image A: N-terminus extracellular, C-terminus cytosolic, helix 1 cytosolic, helix 2 extracellular]
• (b) [Image B]
• (c) [Image C]
• (d) [Image D]
• Explanation: Adding an N-terminal ER signal sequence will cause the N-terminus to be translocated into the ER lumen. The first transmembrane domain (helix 1) will then be inserted with the N-terminus on the lumenal side (later the extracellular side). The second transmembrane domain (helix 2) will be inserted with the C-terminus on the cytosolic side

Question 19

• Question: Your friend has just joined a lab that studies coated vesicle budding… Which of the following protein do you think might be defective in this cell line?
• (a) Sar1-GAP
• (b) Sar1-GEF
• (c) Arg1-GEF
• (d) tethering proteins
• Explanation: Sar1-GAP is responsible for hydrolyzing GTP on Sar1, causing the coat to disassemble. If Sar1-GAP is defective, the coat proteins would remain bound to the vesicle.

Question 20

• Question: You have a protein that normally is constitutively secreted out of the cells. You want to modify the protein to make it accumulate in the ER. What would you do?
• (a) Fuse it to GFP
• (b) Add an N-terminal ER signal sequence
• (c) Add a KDEL sequence
• (d) Remove the N-terminal ER signal sequence
• Explanation: Adding a KDEL sequence (ER retention signal) will cause the protein to be retrieved from the Golgi back to the ER.

Question 21

• Question: Most proteins destined to enter the endoplasmic reticulum _.
• (a) are transported across the ER membrane after their synthesis is complete.
• (b) are synthesized on free ribosomes in the cytosol.
• (c) the mRNAs begin to cross the ER membrane while still being transcribed.
• (d) reside within or travel through the endomembrane system.
• Explanation: Proteins destined for the ER are synthesized on ribosomes that are initially free in the cytosol. Once the signal sequence is recognized, the ribosome is targeted to the ER membrane.

Question 22

• Question: Which of the following statements about the unfolded protein response (UPR) is true?
• (a) Activation of the UPR is initiated by binding of misfolded proteins to cytoplasmic domains of the UPR receptors.
• (b) Activation of the UPR decreases general protein translation.
• (c) Activation of the UPR slows down production of Bip proteins
• (d) Prolonged activation of the UPR increases cell survival
• Explanation: The UPR is activated by the accumulation of unfolded proteins in the ER lumen. Activation of the UPR leads to a decrease in general protein translation to reduce the burden on the ER.

Question 23 - 25

• Instructions: Indicate which of the following descriptions better applies to COPI, COPII or clathrin-coated vesicles (2 points each)

• Question 23: Their coat assembly occurs on the Golgi membrane.

  • (a) COPI
  • (b) COPII
  • (c) Clathrin
  • Explanation: COPI vesicles are involved in retrograde transport from the Golgi to the ER and within the Golgi itself, and their assembly occurs on the Golgi membrane.

• Question 24: Their coated vesicles are involved in endocytosis.

  • (a) COPI
  • (b) COPII
  • (c) Clathrin
  • Explanation: Clathrin-coated vesicles are involved in endocytosis at the plasma membrane.

• Question 25: Their coat assembly requires the activity of Arf1

  • (a) COPI
  • (b) COPII
  • (c) Clathrin
  • Explanation: COPI vesicle assembly requires Arf1, a small GTPase.

Question 26

• Question: On which of the following intracellular locations does clathrin organize a coat and form vesicles?
• (a) Lysosomes
• (b) Plasma membrane
• (c) Endoplasmic network
• (d) Inner membrane of mitochondria
• (e) Regulated secretory vesicles
• Explanation: Clathrin is involved in vesicle formation at the plasma membrane during endocytosis and at the TGN for transport to endosomes.

Question 27

• Question: What would most likely occur if a tethering protein was defective or dysfunctional in a cell?
• (a) The vesicle would become incapable of recognizing its target membrane, resulting in failed fusion and misdirected transport
• (b) The vesicle would fuse with any available membrane, potentially causing random cargo release throughout the cell
• (c) The vesicle would remain tightly bound to the donor membrane, preventing its release and transport
• (d) The vesicle would fuse without the need for any SNARE complex formation, leading to uncontrolled fusion events
• Explanation: Tethering proteins are critical for the initial recognition and binding of vesicles to their target membranes. Without functional tethering proteins, vesicles would not be able to find and bind to the correct target membrane.

Question 28

• Question: Which of the following statements about vesicular membrane fusion is false?
• (a) Membrane fusion does not always immediately follow vesicle docking.
• (b) Action of tethering protein needs to occur before the action of SNARE proteins.
• (c) The GTP hydrolysis of the Rab proteins provides the energy for membrane fusion.
• (d) The interactions of the v-SNAREs and the t-SNAREs pull the vesicle membrane and the target organelle membrane together
• Explanation: Membrane fusion is driven by the interaction of v-SNAREs and t-SNAREs, which bring the two membranes close together. Rab proteins are involved in vesicle targeting and docking, but GTP hydrolysis by Rab is not the direct energy source for fusion. Fusion itself is energetically favorable once the membranes are in close proximity.

Question 29

• Question: How does the acidic environment in the endosome contribute to the dissociation of LDL from its receptor?
• (a) The acidic pH promotes the binding of LDL to the LDL receptor in the endosome
• (b) The acidic pH causes a conformational change in the LDL receptor, reducing its affinity for LDL
• (c) The acidic pH increases the affinity of LDL for its receptor, enhancing internalization
• (d) The acidic pH causes LDL to degrade, leading to its release from the receptor
• Explanation: The acidic pH in the endosome induces a conformational change in the LDL receptor, decreasing its affinity for LDL and causing the LDL to be released.

Question 30

• Question: What is the significance of LDL receptor recycling in maintaining cholesterol homeostasis?
• (a) Recycling ensures that the receptor is available for repeated uptake of LDL, supporting continuous cholesterol absorption
• (b) Recycling reduces the need for LDL uptake, limiting cholesterol entry into cells
• (c) Recycling decreases the efficiency of cholesterol uptake, leading to increased plasma LDL levels
• (d) Recycling has no impact on cholesterol homeostasis, as it only affects LDL receptor turnover
• Explanation: Recycling of the LDL receptor allows cells to continuously take up LDL from the bloodstream, ensuring a constant supply of cholesterol while keeping LDL levels in the blood under control.

Question 31

• Question: What is the primary reason why the low pH of lysosomes prevents damage when enzymes are confined to the organelle but not when released into the cytoplasm?
• (a) The low pH inactivates the enzymes in the lysosome, preventing them from reacting with cellular components
• (b) The low pH promotes enzyme stability in the lysosome, but does not affect their activity outside the lysosome
• (c) Lysosomal enzymes are only active in an acidic environment, so they would not function properly if released into the neutral cytoplasm
• (d) The acidic environment in the lysosome neutralizes the enzymes’ damaging effects, ensuring no harm occurs even if released into the cytoplasm
• Explanation: Lysosomal enzymes are acid hydrolases, meaning they function optimally at acidic pH. If released into the neutral cytoplasm, their activity is greatly reduced, preventing them from digesting cellular components.

Question 32

• Question: (2 points) Insulin is a secretory protein made by the β cells in the pancreas. Sort the following events to reflect the order of signal transduction pathway that leads to insulin secretion.
  • Glucose uptake by the beta cells -> ( ) -> ( ) -> ( ) -> ( ) -> insulin secretion
  • A. Influx of calcium into the cell
  • B. Depolarization of the plasma membrane
  • C. Activation of ATP-sensitive K+ channel
  • D. Glycolysis
• Answer: Glucose uptake by the beta cells -> (D) Glycolysis -> (C) Activation of ATP-sensitive K^+ channel -> (B) Depolarization of the plasma membrane -> (A) Influx of calcium into the cell -> insulin secretion
• Explanation:
  1. Glucose enters beta cells.
  2. Glycolysis increases ATP levels.
  3. ATP binds to and closes ATP-sensitive K^+ channels, causing membrane depolarization.
  4. Depolarization opens voltage-gated calcium channels.
  5. Calcium influx triggers insulin secretion.

Question 33

• (8 points) You set up an in vitro translation system containing the entire translation machinery (ribosomes, amino acids, tRNA, ATP etc). To this system, you add mRNA encoding a 20 kD secretory protein. You perform in vitro translation, with or without addition of ER microsomes or SRP. After completion of the protein translation, you separate the protein products by SDS-PAGE, and visualize the proteins by Western blot analysis. The result is also shown on the right. The presence or absence of ER microsomes and SRP in each reaction is indicated at the top by + and –, respectively. The numbers on the left indicate the apparent molecular mass (kD) of spots on the gel. Briefly explain the molecular mechanism underlying the size of the protein product that you obtained in each reaction. For example, in lane 1, why having mRNA and SRP, but not ER microsome in the reaction results in a protein product of 8 kD? You need to explain based on the related concept that we learned in class. If not, there won’t be any points.

• Lane 1: mRNA + SRP, no ER microsomes - 18kD product

  • The protein is synthesized with its signal sequence because SRP is present, however, because ER microsomes that contain signal peptidase are absent, the signal sequence is not cleaved off, resulting in a slightly larger protein (18kD instead of the expected 16kD).

• Lane 2: mRNA, no SRP, no ER microsomes - 20kD product

  • In the absence of SRP, the signal sequence is not recognized, and the protein is fully translated (20kD) but remains in the cytosol.

• Lane 3: mRNA + ER microsomes, no SRP - 20kD product

  • Without SRP, the ribosome won't be targeted to the ER membrane; therefore the protein will be of size 20kD

• Lane 4: mRNA + SRP + ER microsomes - 16kD product

  • SRP targets the ribosome to the ER microsomes, and the signal sequence is cleaved off by signal peptidase present in the ER lumen, resulting in a smaller protein.