Week 1:
Carver College of Medicine

May 27, 2014

It is always a challenge to get around on a new campus, especially that of the University of Iowa. This 10-week research experience is predetermined to be rather intensive and requires myself to have a good time management. Staying at Cornell this summer, I need to commute to Iowa City every day. Moreover, Highway 1 is currently under construction and the traffic moves extremely slowly in addition to the rough road. The Cornell gym only opens every morning from 6 to 8, meaning that I can only exercise in the morning before going to work. What bothered me the most, however, was the parking problem in Iowa City. Apparently most people working in UI take the bus because the public parking on campus is either nonexistent or extremely expensive. On my first day of work, I parked at the Newton Road parking ramp next to the Bowen Science Building (BSB), where I work at before I met my PI, Dr. Stefan Strack, who told me that the ramp charges $1.50 for every hour of parking. It took us and Dr. Ron Merrill, the research scientist at the lab, quite some time and my extensive walking, getting lost, and exploring in 90 F weather to finally figure out a feasible place for me to park without extensive walking. Thanks to the Cambus, the school bus free to the public provided by UI, I can park my car in the City Park and then take the Blue Route to BSB. When I get off work, I can take the Red Route back to the City Park. Both the Blue and Red Routes run every 15 minutes during the summer.

On the first few days, Stefan and Ron also showed me around the lab, the pharmacology floor of BSB (the second floor), and all the buildings of Carver College of Medicine, and the UI hospitals, which are all connected to each other through footbridges. They and the other two undergraduates from UI who are also working in the lab during the summer, Nandita and Bethany, showed me several places where I can have lunch. There are actually a few Indian buffets that are within walking distance from the medical complexes and a few Cambus routes, such as UI-Research Park and Hawkeye-Hospital, can take me directly to Coralville which has several really good Chinese lunch buffets. I feel like all this information should be helpful for any other Cornell students who will also be doing research at UI in the future but still live at Cornell.

Ron, Nandita, and I went to an Indian buffet in downtown Iowa City on Friday. Nandita is originally from Bangladesh so she recommended several dishes which were all really good. I especially enjoyed the mango latte and the chicken masala. Ron and I also went to the UI hospital’s cafeteria on Thursday. We met two of his former students who are now pursuing MD-PhD dual degrees right now in UI and had a thorough discussion about the medical school application process and the different stages one has to go through to become a doctor. Now when I see people with different uniforms on Newton Road or in the UI hospital I can at least know how close they are to become doctors. This conversation would also be really helpful if I eventually decide to go to medical school in the future.

Now it is time to talk about what actually happened in the lab. It was nice to first walk into the lab and realize that you have all this space to yourself and fancy instruments to work with which are not readily available at Cornell. I also enjoy talking to Stefan and Ron and ask questions whenever I get confused. I also like to visit the lunch room of the department, not mainly for lunch but just to meet and talk to people, mostly graduates and postdocs, from different backgrounds and working on different projects.

For the first week, I did some extensive reading of the journal articles related to the current project that were already published by Stefan and Ron. Below is what I submitted for the “Learning and Assessment Plan,” which would provide really good background information regarding to the outline of the project that I will be a part of for the next ten weeks.

Mitochondrial fragmentation (fission) is responsible for neuronal death in stroke and other neurodegenerative diseases. Thus, blocking mitochondrial fission can protect neurons from a variety of threats. Targeting the fission enzyme, dynamin-related protein 1 (Drp1), is problematic because it is an essential and ubiquitous enzyme in the body. Instead, we hypothesize that inhibiting a neuron-specific activator of Drp1 may provide therapeutic benefit in multiple neurological disorders characterized by mitochondrial dysfunction, including stroke and other neurodegenerative diseases.

We identified Bbeta2, a neuron-specific regulatory subunit of protein phosphatase 2A (PP2A) as a promising drug target. Via an N-terminus targeting sequence, Bbeta2 recruits PP2A to mitochondria, where it triggers mitochondrial fission by dephosphorylating the fission enzyme Drp1. Therefore, silencing of Bbeta2 protects neurons from various insults, whereas Bbeta2 overexpression damages neurons.

We generated mice that lack PP2A/Bbeta2 and found that they are healthy and fertile. Surprisingly, even mice lacking a single Bbeta2 allele display a 50% reduction in infract volume in an ischemic stroke model. Furthermore, Bbeta2 KO mice are completely resistant to peripheral diabetic neuropathy. Therefore, inhibition of Bbeta2 is neuroprotective in a variety of neuronal diseases.

PP2A/Bbeta2 expression is confined to neurons and removing both Bbeta2 alleles has no adverse effects in mice. Moreover, Bbeta2 KO mice are free of stroke and diabetic neuropathy, this PP2A regulatory subunit is a promising target for the safe and effective neuroprotective drugs. Our HTS (High-Throughput) screen is designed to identify cell-permanent compounds that block the association of PP2A/Bbeta2 with mitochondria, which is responsible for mitochondrial fragmentation and neuronal death.

We developed an innovative cell assay based on luminescence induction. The Bbeta2 N-terminus, Beta2(1-24), is sufficient to translocate firefly luciferase (Luc) into mitochondria. Bbeta2(1-24)-Luc has ~5-fold lower activity than a point mutant that stays in the cytosol (Bbeta2(1-24)-Luc K2A). Luc inactivation in the mitochondrial matrix may occur by either covalent modification or inefficient refolding. Molecules that inhibit mitochondrial import of Bbeta2(1-24) would promote a gradual increase in luminescence as newly synthesized fusion proteins accumulate in the cytosol. Inhibitors could interact with either Bbeta2(1-24) or the mitochondrial importing complex. Compounds are applied to neuronal PC12 cell lines that stably express Luc fusions under doxycycline control. We are currently adapting this assay to the 384-well plate format and doxycycline-inducible PC12 cell lines are under selection.