BIO 121 Final Exam
A peptide is being translated in the cytosol and immediately begins to misfold (while it is being translated) because its hydrophobic residues start to aggregate with other hydrophobic residues. Aside from this unfortunate peptide, what protein is likely not functioning properly in this situation?
Hsp70 (a chaperone machinery which is required for degradation of misfolded protein located in the cytoplasm)
A critical mitochondrial transmembrane protein is completely translated on a free ribosome, but not inserted post-translationally into the mitochondrial membrane. What could have prevented insertion into the membrane?
Insertion enzymes are not present
A multi-pass transmembrane protein is found in the ER membrane. Based on that information, what can you tell us about the processing of this protein?
It has multiple start and stop sequences
A scientist changed the sequence of a nuclear-encoded mitochondrial protein—she replaced the mitochondrial signal sequence with an ER signal sequence. How will this change affect the process of translation for this protein?
It will now be translocated on a bound protein instead of a free protein. SRP will now bind to this peptide as well.
You are studying a gene that encodes a secreted protein. What would happen if its signal sequence was removed or missing prior to translation?
It would be translated on a free ribosome in the cytosol
The mRNA transcript encoded by a gene called TLR4 has 3 EJCs (Exon Junction Complexes) bound to it, while the mRNA transcript encoded by a gene called RBBP5 has 8 EJCs bound to it. What does this tell you about these two mRNAs?
RBBP5 mRNA must have more splicing enhancers than TLR4
What levels of protein structure would be disrupted if hydrogen bonding was prevented in a protein composed of a single peptide?
Secondary and Tertiary structures
What does the stability of the transcription pre-initiation complex determine?
Stable PIC = More RNA Pol II recruited = More mRNA transcripts produced = higher transcription efficiency
How are TFII transcription factors involved with the transcription pre-initiation complex?
TFII's are expressed in every cell type. They form a complex at the promoter and work together to recruit RNA Pol II. RNA Pol II and TFII's are critical for transcription.
You are studying a disease that occurs in individuals when an ER transmembrane protein called D2R is not inserted into the ER membrane of neurons. You find that the D2R gene is transcribed, and the mRNA is exported out of the nucleus. You also notice that all other ER transmembrane proteins are inserted into the ER membrane properly. Based on what you have learned about RNA and protein processing in Lectures 9-13, what could be a cause of this disease?
The D2R gene has a mutation in start/stop transfer sequences that prevents recognition by the ER protein translocator
What would happen to developing cardiomyocytes in the sinoatrial node (SAN) of an embryonic heart if you replaced the SAN ECM with ECM from the left ventricle (LV). (In other words, you removed the SAN ECM and replaced it with LV ECM in the sinoatrial node.)
The developing cardiomyocytes in the SAN would differentiate into contractile cardiomyocytes instead of pacemaker cells.
What is the evolutionary history of non-nuclear DNA in mitochondria?
The endosymbiotic theory posits that mitochondria and chloroplasts evolved from prokaryotic cells that were engulfed by ancestral anaerobic eukaryotic cells. Mitochondria also have their own genomes made of plasmid DNA, reminiscent of most prokaryotic DNA.
Defensin is a defense-related gene produced in macrophages when they are activated against pathogens (for example, upon sensing LPS). You are treating a patient with a severe bacterial infection and notice that although their macrophages are transcribing defensin mRNA, the mRNA is not being efficiently translated. What change should occur to the defensin mRNA upon infection? What could be happening in this patient?
LPS typically the macrophages to produce polyadenylation factors which causes a switch in poly-A site usage for inflammation related genes such as defensin
Describe the process of "leaky scanning". What does this process produce?
Leaky scanning occurs when a scanning ribosome misses the first AUG and uses the second as the START codon instead. This process produces an "open reading frame". This results in using the kozak sequence to initiate translation.
A scientist has engineered a novel glutamate receptor for monitoring excitatory neurotransmission at synapses in neurons. The domain that binds glutamate is in the extracellular space of the synaptic cleft. Which compartments has it traveled through on its way to the plasma membrane?
Lumen of the ER, Golgi, and intracellular vesicles
If a cell was missing the M6P receptor, how would that affect the way lysosomal hydrolases are processed?
Lysosomal hydrolases would not be packaged into vesicles at the TGN (trans golgi network)
Describe the process of vesicle formation during outward flow. Include the names of proteins required for each step, as well as important details related to how phospholipid bilayers fuse. (Your answer should prepare you for a short answer question OR a multiple choice question, like "Which of the following statements about vesicular formation and fusion are true?")
1. Vesicles bud off of a membrane with the help of coat proteins 2. Vesicles are transported to their next membrane with the help of motor-binding proteins and molecular motors 3. Vesicles are specifically targeted to their next membrane by vesicle membrane proteins called Rabs 4. Vesicles fuse to their destination membrane due to their lipid constituents and membrane proteins called SNARES
A peptide is produced in two different cells. The primary structure is the same in both cells, but the amount of peptide is increased in one cell (i.e. more molecules of the peptide are translated in one cell). What could be happening to the mRNA transcript within the cell that has the higher amount of this peptide?
1. an alternative poly-A site is being selected 2. Leaky scanning occured
What does an mRNA need to have in order to be exported from the nucleus?
1. at least 1 exon-junction complex (EJC) 2. Properly processed poly-A tail 3. 5' cap
Name and describe the 3 types of inward flow (Hint: you should be prepared for both short answer and multiple choice versions of this question, for example: "Which of the following are the 3 types of inward flow?" or "Which of the following is NOT one of the types of inward flow?")
1. phagocytosis: import of solid material 2. pinocytosis: import of fluid 3. receptor-mediated endocytosis: import of specific material recognized by receptor protiens
When a protein is subject to proteolysis, what are the potential outcomes?
1. protein is degraded 2. protein is functionally activated 3. protein is irreversibly modified
TGF is a gene that is normally expressed in retinal cells. How could you inhibit expression of TGF in retinal cells?
Mutate the enhancer or TSTF so it can't bind
Neuroligin is a gene that is normally expressed in neurons. A patient has a neurological disorder, and you discover that this gene is not expressed in the brain of this patient. How might this happen?
Mutations - ?
Describe the structural details of phospholipids. Which parts can differ between phospholipids and how do these changes to the molecular alter how the phospholipid behaves? (Note: a multiple choice version of this question could be: "Which of the following is NOT a true statement about phospholipids?")
- Fatty acid tails: length varies; saturated vs unsaturated (double bond = more fluid) - Polar head contains (bottom to top): - Glycerol (DOES NOT VARY) - Phosphate - Modification on top: 4 major classes (if it has a charge or is neutral will change what it is attracted to)
What is a snRNP? What process does it participate in. What does it do and how does it recognize the location it needs to bind to?
- snRNP = small nuclear ribonucleoproteins - several units make up the spliceosome that binds to an intron sequence and recognizes these intron-exon boundaries - recognizes the sequence and nucleotides that make up an amino acid bordering the introns to cut it out - Holds the pre mRNA in the correct configuration - Catalyzes the chemical reactions that remove introns and covalently link exons (participates in pre mRNA splicing)
In another cell, TBK2 is transcribed, but the mRNA for TBK2 is never exported from the nucleus. What are the possible explanations for this?
1. A premature termination (stop) codon triggered nonsense mediated decay (NMD) (EARLY DEGENERATION) 2. Poly-A binding proteins (PABP's) did not bind to the Poly-A tail (lack of mRNA transport which is associated with transporting molecule mechanisms)
Genetic mutations can quickly lead to antibiotic resistant bacteria. Why can antibiotic resistance develop so quickly in bacteria?
1. Bacteria reproduce quickly --> overuse of antibiotics --> resistent bacteria 2. Bacteria pass ALL their DNA to their progeny
What are the common functions of transmembrane proteins?
1. Cell-adhesion to the ECM 2. Signal transduction 3. Transport of charge molecules 4. Cell-to-cell recognition
Compare and contrast the four major types of intracellular signaling based on the types of ligands involved, how far the ligand travels between the ligand-producing cell and the responding cell, and whether each is considered to be a type of local or systemic signaling and what that means. (NOTE: A multiple choice version of this question could be significantly simpler than the open ended wording here, asking you simply to identify the mode of intracellular signaling based on certain unique features...)
1. Contact-dependent/Juxtacrine: Membrane-bound ligand, highly local 2. Paracrine Signaling: local, receptor and specific ligand, short range, trigger response in same tissue (NO) 3. Synaptic Signaling: Contact and diffusion, quick and precise 4. Endocrine Signaling: long range diffusion, ligand = hormone
List and describe the different post-translational modifications that we talked about in lecture
1. Covalent modifications: any chemical change to an amino acid - it can be permanent or reversible - often changes conformation of protein that results in a functional change a. Glycosylation: Addition of carbohydrate group (usually occurs in ER) - facilitates protein folding by making intermediate folding steps more water soluble and recruiting from proteolytic enzymes - protecting proteins from proteolytic enzymes - forming mucus - identifying protein to other cellular machinery b. Phosphorylation: Addition of a phosphate group - kinase and phosphatase are molecular switches c. Lipid Modification/ Palmitoylation: Addition of a lipid group so a protein can have a tight association with the membrane (NOT TRANSMEMBRANE PROTEINS!) Can leave whenever they wants since controlled by enzymes. 2. Proteolytic modifications: some polypeptides must be processed by proteases before they are fully functional (insulin) 3. Degradation of Proteins by the Ubiquitin-Proteosome System
A protein is produced in two different cells. The primary structure is the same in both cells, but the amount of peptide is greater in one cell. What could be happening to the mRNA transcript in the cell in which the amount of peptide is greater?
1. Cytosolic Poly-A polymerases could be continuously lengthening the Poly-A tail 2. The recognition of an alternative poly-A site during transcription could have led the mRNA for this protein to have a shorter 3' UTR which does not have a binding site for a translational inhibitor
What are the different ways in which new genes can be created?
1. DNA shuffling 2. Mutation 3. Gene duplication and divergence
You are characterizing cells in a tissue section. You have identified a group of cells that perform the primary functional role associated with the tissue, are terminally differentiated, and are tightly connected with sparse ECM. What terms would apply to this cell type?
1. Differentiated cells 2. Epithelial cells 3. Parenchymal cells
NCAM is a gene that is normally only transcribed in neurons. How could you induce (force) expression of NCAM in muscle cells?
1. In the muscle cell, express the TSTF from neurons that binds the NCAM enhancer 2. In the muscle cell mutate the NCAM enhancer sequence so another TSTF already present in the muscle can bind
Which processes contribute to genetic variability of offspring during vertical transfer?
1. Independent assortment 2. Crossing over 3. Random fertilization
What are the different connective tissues in the body? What are the names of the cells that make up each tissue?
1. Loose connective tissue: Adipocytes and fibroblasts 2. Dense connective tissue: fibroblasts and chondrocyte 3. Bone: osteoblasts, osteoclasts, osteocytes 4. Cartilage: Chondrocytes 5. Blood: WBC, RBC, platelets
What are the ECM functions in multicellular organisms?
1. Mechanical support and defense 2. Adhesion for cells and tissues 3. Substrate for cell movement 4. Regulation of cell function 5. Role in cellular differentiation
What would happen within a cell if it had a premature termination codon in the gene encoding the ER protein translocator?
1. NMD could interfere with translation of the mRNA encoding in the ER protein translocator 2. N-link glycosylation on proteins usually translocated into the ER would not occur
List all the ways a protein may be associated with the membrane (Hint: be prepared for both short answer and multiple choice versions of this question, for example "which of the following are ways a protein may/may not be associated with the membrane?"
1. Protein contains a single-pass transmembrane protein 2. contains an alpha helix embedded in the cytosolic leaflet of a membrane, made possible by hydrophilic amino acids facing one side of the helix and hydrophobic amino acids facing the opposite 3. Protein has a protein-protein interaction domain which binds to a transmembrane protein
What are the functions of the 5' cap?
1. Required for export of mRNA from the nucleus 2. Required for splicing the first intron 3. Promotes recruitment of ribosomes for translation
A gene was successfully transcribed into mRNA, and then exported into the cytosol. But the peptide it encodes is not successfully translated. What could prevent or disrupt translation in the cytosol?
1. The activity of deadenylases 2. The 3' UTR has a miRNA recognition site 3. The 3' UTR has a binding site for a translational inhibitor 4. There is a premature stop codon in the first exon of the transcript, causing the ribosome to stop before clearing all of the EJC's
What is required for Nonsense Mediated Decay to occur?
1. The mRNA must be exported from the nucleus 2. A test round of translation must be performed 3. The ribosome must stop before clearing all the EJC's
The CFTR gene is very well conserved from mice to humans. What conclusions can you make about CFTR based on this information?
1. The protein encoded by the CFTR gene has a critical function in those organisms 2. CFTR is just as susceptible to mutation as other genes, but many CFTR genes are lethal, thereby preventing them from being passed on.
You are trying to determine the molecular cause of a bone disorder. In the diseased osteoblasts, a gene called Cav1—which normally encodes a calcium channel membrane protein—is not being transcribed (no Cav1 pre-mRNA is being produced at all). All other genes in the osteoblast are appropriately transcribed. What is an epigenetic reason that Cav1 might not be transcribed? What is a genetic reason that Cav1 might not be transcribed?
?
How would loss of SRP affect a cell
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What are the main functions of the Endoplasmic Reticulum and the Golgi Apparatus?
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You are diagnosing a person with a neuromuscular disorder that prevents their muscles from contracting (they are in a permanently relaxed state). You realize the problem: the specific step involving an allosteric regulator is not occurring. What could be the direct cause of this?
A critical binding site is blocked
Why is it important that highly expressed genes have long poly-A tails?
A long poly-A tail increase stability of mRNA and slows down degredation
Actin filaments and collagen fibers are examples of polymers. A protein polymer is defined as a _______________________
A polymer means more than one thing, hence more than one protein. Polymer formation is a source of diversity for the final functional molecule
When the calcium channel is not expressed, calcium cannot be released into the bone ECM. How would this impact the bone?
Bone will weaken
How does DNA methylation inhibit transcription?
By preventing transcription factors from recognizing their sequence-specific binding sites.
According to the fundamental cell theory, where do cells arise from?
Cells arise from pre-existing cells
Define what a scaffold complex is and how its function could be disrupted
Cells can start and stop a protein's activity by regulating interactions of proteins within a scaffold complex. - If any single protein is prevented from associating with the scaffold complex, IT CANNOT FUNCTION PROPERLY, and the target protein is not ubiquitylated.
An intracellular vesicle leaving the TGN is targeted for the plasma membrane. How could its destination be changed so that it would target an endosome instead?
Change the Rab proteins on the cytosolic side of the vesicle so they match Rab effector proteins on the endosome
A scientist looking at a group of cells in an epithelium under the microscope noted that they likely had a role in moving mucus over the cells, were rectangular-shaped, and had nuclei at different heights, but all shared a common basal lamina. What epithelial cell type is the scientist observing?
Ciliated pseudostratified columnar epithelial tissue
Why are CpG islands important for epigenetic memory?
DNMT 1 to daughter DNA strands after replication ?
According to the histone-code hypothesis, what is one way that cell specialization occurs?
Differential expression of histone modifying enzymes
What is the main function of oligosaccharides on glycosylated proteins in the plasma membrane?
Direct proper folding of proteins in the ER
You are looking at an image of a cell that has a defect in dynamin. It is obvious to you that the dynamin in this cell is either not functioning or not expressed. Why is it obvious? (i.e., What would you expect to see in this image?)
Dynamin assembles into a ring around the neck of the forming bud. ** there are vesicular buds at the plasma membrane, but no vesicles are pinched from the membrane
A gene called TBK2 has different splicing isoforms found in different cell types, including intestinal epithelial cells, white blood cells, and skeletal muscle cells. What mechanism allows different splicing isoforms to be expressed in the different cell types?
Each cell type expresses different splicing recognition factors
What is meant by "combinatorial regulatory code" when discussing covalent modification?
Each proteins set of covalent modifications creates a combinatorial regulatory code --> different combinations of modifications on the same protein = different functions
What is an epithelial layer that could perform osmosis and diffusion well? (Be sure to understand why.)
Endothelium (blood vessels). performs gas and fluid exchange
How do enhancers regulate gene expression?
Enhancers bind TSTF's. This drives transcription
Two peptides have almost the exact same primary structure, except that one has about 10 fewer amino acids at the amino-terminus (N-terminus) of the protein. What is a possible explanation for the protein that is missing those 10 amino acids?
Experienced leaky scanning, skipping the first start codon
You are trying to characterize a cell as either eukaryotic or prokaryotic. It has DNA, RNA, and proteins; it is metabolically active; it participates in intracellular homeostasis; and it does not have a membrane-bound nucleus. What is this cell?
Prokaryote
What types of proteins are translated on free ribosomes?
Proteins that are directed to their final destination not the ER after they are translated. Proteins destined for: - cytosol - nucleus - mitochondria - chloroplasts - peroxisomes
What types of proteins are translated on ribosomes that become bound to the ER?
Proteins that must go to the ER first before their normal functions. (endomembrane proteins) Proteins destined for: - The extracellilar space - Plasma membrane - lysosome - ER -Golgi - Secreted proteins
You are studying a disease caused when the gene BMP5 is not transcribed in osteoblasts. All other genes are transcribed normally. To investigate why BMP5 is not transcribed, you create a reporter transgene by putting GFP under the control of the BMP5 enhancer found in diseased osteoblasts. You put this reporter transgene into a normal osteoblast. The normal osteoblast with your reporter transgene does NOT turn green (GFP is not expressed). What does this result indicate?
The enhancer gene BMP5 is mutated in disease osteoblasts. Enhancer genes are DNA sequences that bind TSTF's. Since the enhancer gene BMP5 is mutated it cannot bind to its TSTF. Therefore, the GFP cannot be expressed.
Define and describe the process of endocytosis
The inward flow of material into the cell by budding vesicles at the plasma membrane
You isolate a group of newly formed vesicles from a cell. All of the vesicles have clathrin associated with them. What does that information tell you about where this vesicle could or could not have originated from?
The vesicles could have formed from the Golgi, plasma membrane, or through receptor-mediated endocytosis
What do a transcriptional enhancer, a splicing enhancer, and a poly-A site all have in common?
They are all nucleotide sequences that recruit specific protein factors and regulate the expression of a gene.
What do transmembrane proteins in the peroxisome, nucleus, and mitochondria all have in common?
They are all post-translationally inserted in the membrane
You are analyzing the activity of the protein p53 in two different cell types. You notice that p53 has the same primary structure in both cells but has a different function in each cell. What could be the reason for the functional difference in this case?
They have different covalent modifications
The gut and tracheal epithelium both have tight junctions. Why?
Tight junctions form a virtually impermeable barrier that is critical to prevent infection
What specific epithelium is being described in the previous question? (i.e. Where in the body would you find this epithelium?)
Tracheal Epithelium
If a person with Type A blood receives a blood transfusion of Type B blood it can have lethal consequences. Why?
Type A and Type B blood cells have different oligosaccharide structures on their surface that are recognize differently by our immune system
What does it mean to say that proteins have modularity of structure?
Various functions of a protein can be performed by independent domains connected by short, unconstructed stretches of amino acids
Is an individual enhancer able to bind more than one transcription factor? Why?
Yes ?
Can phosphorylation—a post-translational covalent modification—cause a protein to have different functions in different cells? Why or why not?
Yes. It leads to conformational changes which essentially changes the function of the protein.
In the question above, does it matter if the BMP5 enhancer-GFP reporter gene is tested in a normal/healthy osteoblast or in a diseased osteoblast?
Yes. Normal cells can transcribe BMP5. Diseased cells expression is not at a detectable level
The same gene is transcribed in two different cells, but the mRNA sequences produced from this gene are different in each cell. What are possible explanations?
alternative splicing & alternative poly-A site selection
Lactose intolerant individuals do not produce the enzyme lactase. Studies show that the gene that encodes lactase is not transcribed. Describe the conformation of the nucleosomes (condensed or uncondensed) surrounding this gene and the potential histone modifications that could cause this conformation.
condensed, deacetylation
What does it mean to be amphipathic?
having both hydrophilic and hydrophobic parts
You are examining a cell that has a very rigid plasma membrane at room temperature. What is a likely explanation?
high levels of cholesterol
Cystic fibrosis is caused by mutations in the CFTR gene, which prevent the function of the chloride channel (a plasma membrane protein) it encodes. Which phenotype is being described?
molecular phenotype
If a cell was specifically having trouble transporting vesicles from a source membrane (like the ER or Golgi), what proteins might be defective?
motor proteins
An mRNA is exported from the nucleus, but not circularized. What could prevent circularization? What would happen to this mRNA?
poly-A tail and 5' cap????
What is a mechanism that allows the same protein (with the same amino acid sequence) to have different functions within cells?
post-translational covalent modification
How can you change the structure and function of a protein without changing the DNA sequence of the gene?
post-translational modification
After attempting to fold, the protein described in the previous question was degraded by a ___________________ because too many __________________ residues were located on the exterior of the protein (facing the cytosol) after folding.
proteasome, hydrophobic
CASK (Calcium/calmodulin-dependent serine kinase) is a membrane-associated guanylate kinase (MAGUK) found in metazoans. It is in the same gene family as PSD-95 and has PDZ, SH3, and non-catalytic guanylate kinase domains used for binding to other proteins at cell junctions, but it also has a Calcium/Calmodulin-dependent kinase (CaMK) domain which it uses to phosphorylate targets. How might the PSD-95-like ancestor of CASK have obtained the CaMK domain? Where did the CaMK domain come from and how did it find itself fused to the MAGUK protein?
A transposon near a gene encoding protein which contained a CaMK domain "jumped" into the 5' UTR of the MAGUK gene, taking with it a chunk of neighboring DNA including the coding sequence for the CAMk domain.
What are MicroRNAs (miRNAs) known for?
A very short single stranded rna that interferes with gene expression by base pairing with their targets on the mRNA and recruiting various targets. - They usually bind in the 3' UTR - originally evolved as a way to combat RNA viruses - helps tune localized gene expression - base pairs to itself - control of gene expression
A gene is transcribed in a fibroblast and a chondrocyte, but the primary structure of the encoded peptide is different in the two cells. In the fibroblast, there appears to be missing amino acids at the C-terminus. What could be the cause of this difference in the fibroblast?
An alternative poly-A site was used, causing the last exon to be removed from the primary transcript
You are examining a patient with a degenerative condition caused by a build-up of aging cellular machinery. Usually aging cellular machinery is trafficked to the lysosome so that it can be degraded. What form of vesicular trafficking is not occurring properly in this patient?
Autophagy: forming vesicles around cellular components for the purpose of transporting to the lysosome for digestion (self eating inside the cell)
What would happen if a lysine residue on a histone tail that was normally acetylated was mutated to an arginine? (Note: arginine is a positively charged amino acid that cannot be acetylated; for the purpose of this question, assume this arginine is not subject to methylation)
Genes within or near the nucleosome affected by this mutation would be more likely to be condensed
Does dense connective tissue have relatively low or high amounts of ECM compared to cells?
HIGH. more ecm=more dense
Compare and contrast how organisms were grouped into taxonomic rankings historically versus currently.
Historically (before DNA sequencing), organisms were grouped into specific taxonomic rankings based on observable similarities and differences in morphology. Now we sequence DNA and compare how related organisms are based on how similar their DNA is.
What phenotype would occur if there was a decrease in myelin-producing cells?
Nerve dysfunction. Nerves do not conduct electrical impulses normally (damaged information networks)
What would happen if the balance between osteoblasts and osteoclasts were disrupted and too many osteoblasts became active? What if the reverse were true, and there were too many osteoclasts active?
Osteoblasts secrete bone ECM. If osteoblasts become too active, too much bone tissue is produced which leads to bone enlargement. Osteoclasts help maintain bones, develop osteoblasts, and reabsorb tissue. If too many are acting, there is more bone resorption than production and the bones will weaken.
A protein called Ras is tethered (linked) to the plasma membranes of stem cells where it participates in signal transduction pathways that affect stem cell proliferation. However, Ras does not have any transmembrane domains in its amino acid sequence. What covalent modification would be responsible for tethering (linking) Ras to the membrane?
Palmitoylation (lipid mod)
