Problem Set 5

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Problem Set 5 Biology 311 Fall 2022 Nicholas D’Souza 1. Lysosomes and proteasomes play critical roles in cell function. a. What is the importance of acidity to lysosomal function? A pH of around 4.5 to 5 is required for lysosomes to function properly. This is because lysosomal enzymes lose their phosphate group at this pH in the lysosomes which increases their activity. b. What is the difference between proteasomes and lysosomes? The main difference between lysosome and peroxisome is the enzymes they contain and their functions. This is realized by lysosomes being able to break down a large variety of molecules while proteasomes can only break down proteins. c. How does ubiquitination play a role in proteasome function? Ubiquitination is the conjugation of ubiquitin to proteins that need to be degraded by proteasomes. In this way, Ubiquitination allows proteasomes to target proteins that they will break down. d. Enzyme Replacement Therapy (ERT) is one common approach to treating children and adults with select lysosomal storage diseases. What is the molecular/cellular basis underlying how ERT works in this regard? By receiving enzyme replacements, the body of people with lysosomal storage diseases are able to successfully perform the functions inhibited by the deficiency. This means that their lysosomes are now able to digest and break down the molecule which was previously undegradable. e. Why was the co-culture/conditioned medium experiment described in class showing that the co- culture of normal cells can “phenotypically correct” Hurler’s cells important to both our understanding of lysosomal enzyme trafficking as well as developing ERT? This experiment showed that lysosomal enzymes can both be secreted from cells and be absorbed and used by other cells to cover their deficiency in those enzymes. This means that lysosomal storage diseases can be treated by allowing cells to absorb the enzymes they are missing allowing them to return to a normal phenotype. f. Challenge : Bone marrow transplant is an alternative to ERT for treating lysosomal storage diseases. Why does this make sense based on how ERT works? The transplanted bone marrow would produce cells that are able to correctly produce lysosomes with all the required enzymes which would then be secreted and absorbed by diseased cells allowing them to return to a normal phenotype. g. There are many organelles that appear similar to lysosomes yet they are endocytotic vesicles like late and early endosomes. What is the accepted histochemical method to unequivocally identify a lysosome from other similar appearing organelles? A lysosome is an organelle that is marked by acid phosphate. h. Mitophagy is a form of autophagy that involves the fusion of lysosomes to targeted mitochondria. How may mitophagy be related to aging, health and longevity? (Hint: Use Pubmed and put in “longevity”, “aging” and “mitophagy” as key words) Mitochondria regulate organismal fitness and longevity via multiple pathways, including cellular senescence, stem cell function, inflammation, mitochondrial unfolded protein response (mtUPR), and bioenergetics. Thus, mitophagy, which is the proper removal of damaged or unwanted mitochondria, is thought to be pivotal for maintaining organismal health span and lifespan and the protection against aged-related degeneration.

2. Protein trafficking of lysosomal enzymes is critical to ensure that newly synthesized lysosomal enzymes are packaged inside lysosomes. a. Where are lysosomal enzymes synthesized in cells? Lysosomal enzymes are synthesized in the Rough endoplasmic reticulum. b. What is the importance of the cis-Golgi and Trans Golgi Network (TGN) in the tagging and sorting of lysosomal proteins? The cis and Trans golgi Network make sure that the enzymes that work in the lysosomes are able to be delivered to the lysosomes. Furthermore, the Trans Golgi Network also delivers some macromolecules to the lysosomes to be degraded. c. How does I cell disease differ from Hurler’s Syndrome, Pompe’s Disease and Gaucher’s disease? Unlike in other diseases in I cell disease lysosomal enzymes are still being produced normally however they are unable to be sent to the lysosomes and rather end up in the extracellular medium. 3. Mitochondria generate ATP but are also known to trigger cell death. Furthermore they are involved in other disease states as well. a. Explain the dual role of cytochrome C in mitochondrial function. Cytochrome C normally works as an electron receptor and shuttle in the electron transport chain but also plays a major role in mitochondrial mediated cell apoptosis. When Cytochrome C leaves the mitochondria and enters the cytoplasm cell death follows. b. What is the function of tunneling nanotubes and why is it important to mitochondria? Tunneling nanotubes allow materials to easily move between cells. Mitochondria are able to travel between cells using tunneling nanotubes. c. What is the function of Endoplasmic Reticulum Mitochondrial Associated Membranes (MAMs)? Mitochondria-associated membranes are a region of the endoplasmic reticulum which is tethered to mitochondria. These membranes are involved in import of certain lipids from the ER to mitochondria and in regulation of calcium homeostasis, mitochondrial function, autophagy and apoptosis. d. Why is Caspase 3 often nick named the "executioner" enzyme? Caspase 3 directly causes apoptosis by coordinating the destruction of cellular structures such as DNA fragmentation or degradation of cytoskeletal proteins. e. Why is bcl-2 important to consider in select cancer treatments and what steps have been taken recently to address this issue? Bcl-2 is an anti-apoptotic protein because it prevents mitochondrial mediated apoptosis from occurring. Overproduction of Bcl-2 is common in many different cancers and prevents cells from being killed by chemotherapy. This problem is being addressed by adding Bcl-2 antisense oligonucleotides like Genasense to chemotherapy. f. Both Velcade and Obatoclax are currently being used to treat a variety of cancers. How do they differ in function? Velcade is a proteasome inhibitor while Obatoclax is an inhibitor of Bcl-2 g. What is the Warburg effect and why is this especially important to cancer cells? The Warburg effect is that in cancerous cells there is an increase in glycolysis so that more ATP can be produced which allows cancerous cells to divide rapidly. h. Cyanide, carbon monoxide and rotenone all work differently on mitochondrial function compared to Cyclosporin A. Explain this difference.

Cyanide, CO, and rotenone are electron chain inhibitors while Cyclosporin A blocks MPTP from opening preventing mitochondrial mediated apoptosis from occurring. i. Where is thermogenin located and what is its purpose? Thermogenin is found on the inner membrane of mitochondria and generates heat through mitochondrial uncoupling. 4. Photosynthesis is one of the most important biological reactions on the earth with RUBISCO being the most plentiful enzyme on earth. a. RUBISCO, while in high abundance, is a very sluggish (by enzymatic standards) enzyme. It makes up that difference by increasing the amount present in plants. RUBISCO can catalyze both carbon fixation or if oxygen is high it can carry on photorespiration. Why is the latter considered “wasteful”? Unlike in carbon fixation during photorespiration there is very little production of ATP b. Photoinhibition is distinctly different from photorespiration. What is photoinhibition and Photoinhibition is the light induced reduction in photosynthetic activity. c. While we are talking about words that begin with the letter “P”, what is plastocyanin and what characteristics does it have in common with Cytochrome C? Plastocyanin is a copper-containing protein that plays a role in the electron transport process associated with photosynthesis. Just like Cytochrome C it moves electrons through the ETC. d. Another “P” word is phycoerythrin. Why isn’t phycoerythrin, a pigment associated with photosynthesis, found in land plants? Phycoerythrin absorbs and harvests energy from blue light and reflects red, which gives algae that possess it an advantage in living in deeper water. However, on land red light isn’t any more available than other colors so this adaptation gives no benefit. e. Finally, the last “P” word for this question. Phosphoenolpyruvate plays a critical role in C4 plants. What does it do? Phosphoenolpyruvate acts as the second source of ATP in glycolysis in C4 plants. f. Opinion : As discussed in class, in an article in GEN published in October 2022, Neoplants, a Paris based biotech company, announced that its first product, Neo P1, will be available for sale. This plant was developed through DNA sequencing a common houseplant, using CRISPR-Cas9 for gene editing, and using directed evolution so that this plant can consume 30 times more VOCs (Volatile Organic Compounds) than most house plants. Neo P1 converts formaldehyde into fructose that it can used as a food source for the plant. It converts benzene, toluene, ethyl-benzene and xylene into an amino acid. It comes with a special, microbiome bacteria cocktail that also does the same. The cost is $179 for the plant, the self-watering stand and the first dose of microbiome. Of interest is the fact that the well-known and revered magazine, Consumer Reports, just printed a lead article at the same time on the presence and danger and VOCs in homes (November 2022, “ How to Stay Safe from Hidden Home Hazards”). But if you are the consumer you would never be able to determine if Neo P1 is really working because you simply have no easy way to measure the levels of VOCs in your home or apartment. So here is the question: Would you merely take the word of Neoplants that it works and really reduces the ambient VOC levels in a typical household space and thus purchase a plant? Or would you require that a well-known third-party testing laboratory such as Consumer Reports verify the authenticity of the function of Neo P1?

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I would wait for their claims to be validated before buying a plant. $179 is super expensive for a plant that doesn’t do anything. The results would have to be impactful and impactful very quickly for the plant to be worth it. Also, I doubt the price of these plants would increase all too much even if their claims were validated so there’s no harm in waiting.

Problem Set 5

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