Hughes Mentor:  Teresa Gunn

Department: Vet Biomedical Sciences

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A Day in the Life of an Undergraduate in the Gunn Lab

Working as an undergraduate researcher in the Gunn lab came quite late, and unexpected, in my academic career as a Cornell student. I had originally come to Cornell in order to pursue some type of undergraduate research, but had been largely unsuccessful in finding a lab for the first 2_ years. However, by establishing a network of Cornell faculty during that search, I was able to find a lab opportunity in the Gunn lab in the middle of junior year. With that in mind, it took a lot of patience, perseverance, and networking to find such an opportunity, and I encourage anyone wishing to conduct research to forge relationships with their professors and other faculty.

Gunn lab, which is a molecular genetics lab in the Department of Biomedical Sciences, focuses on elucidating the molecular mechanisms behind Mahogunin (Mgrn1), a gene in Mus musculus (mouse) that encodes a protein (Mahogunin Ring Finger 1 – MGRN1) that participates in the pathway responsible for the degradation of cellular proteins. There are a number of phenotypes that appear in the absence of this protein, such as darker fur pigmentation, and neurodegeneration. Given the scope of this research, it was beneficial that I had previously taken the introductory biochemistry and molecular biology courses; in addition to the basics of that field, the techniques that are covered in these classes were ones that I do use in the lab, such as PCR, gel electrophoresis, DNA cloning, Northern and Western Blots, etc. To this day, my biochemistry textbook has served as a decent reference guide for some of the techniques or concepts that I am using. And because I focus more on the neurodegenerative aspect of Mgrn1, it helped to have also taken Introduction to Neurobiology 222, which focuses on more molecular, biochemical, and anatomical aspects of neurobiology. Other courses that may be of benefit include genetics, cell biology, and physiology.

A day in the lab varies depending on how well your experiments go and how efficient you are with managing your time. There are days when I’m constantly on my feet running from one part of the lab or building to another, running experiments simultaneously, and there are days things are going pretty slowly. To avoid such days, I try to plan my experiments for the day ahead of time. On those days, it’s good to keep oneself busy rather than sit around—I prefer to do literature searches and read publications in order to learn more background about my own research. Any questions that I have about the literature or my own project can be directed to any of the graduate students who’re not as busy in the lab, or my PI, Teresa Gunn. In addition, there are a number of meetings throughout the week, whether they be lab member meetings (where everyone presents at some point), specialty group meetings, or other talks that are happening in the Vet School that serves as an opportunity to learn about someone else’s research (such as journal club).

For my research, I am ultimately testing for potential interactions between candidate proteins and MGRN1 protein because these candidate proteins are overexpressed in mice without MGRN1 protein, which suggests that they accumulating (and not degrading) inside the cell. But before I can do this, I must test to make sure that RNA expression levels are equal in the presence and absence of MGRN1. This requires working with RNA extracted from mouse brains, and using RT-PCR techniques to generate DNA that can be diluted down into specific concentrations that serve as my standards. Then, using quantitative RT-PCR, I can correlate the rate at which gene-specific RNA is transcribed into DNA and amplified to a specific concentration using these DNA standards, which determines the initial concentration of RNA for a specific gene. This type of procedure is very sensitive to contaminants and other sources of error; this error is compounded by the fragility of RNA, which is easily degradable. Thus my project requires that I work as quickly and efficiently as possible to reduce the amount of error, and that everything be done on ice in order to prevent RNA degradation. I follow this procedure with agarose gel electrophoresis to make sure that only one product was amplified in my qRT-PCR reactions. Despite the apparent complexity of the procedure, it is comprised of basic techniques which are taught in introductory biochemistry courses, and even introductory biology.

This research takes up a lot of time, which is something that I did not take into consideration during my first semester of research. On top of research, I was also taking 20 credits worth of science courses, and a part-time job, as well as various on-campus activities. I found that semester very stressful as a result, so I urge others to take around 16 credits maximum if you plan on doing research, with a decent amount of time still left for studying. Because of the work that is required of research, I think it is easy for one to lose sight of the fact that academic courses are the first priority. I spent three hours a day, four days a week, in a lab, but I feel that it would have been better if it were more like six hours a day, two days a week. You end up spreading out your workload to a more manageable level, and you get more experiments done that way. Most experiments take more than three hours to perform.

I’ve learned a lot from this lab experience over the past few months, not only in factual knowledge, but about myself and about science in general. One thing you constantly here is that research is very demanding, and doesn’t always work the first time. After experiencing it firsthand, I say that that is an understatement. One literally has to develop a strong reserve in order to keep going with a research project that keeps giving unsuccessful results. As I said at the beginning, it takes a lot of patience and perseverance; there’s no instant gratification involved. Students who seek otherwise would probably not find research to be any rewarding. It’s also taught me to think in a broader, more efficient context, and not worry myself about the little details as I’m personally prone to doing. I’ve also gotten a lot of experience talking to people about my research, both in a one-on-one basis and in addressing an audience. It’s helped to build up some confidence in a very nerve-wracking experience. Anyone who’s looking to learning the same things as I have should apply to the Gunn lab, because these are the things that Teresa Gunn tries to teach you.

A Hughes Scholar Guide to Doing Research in the Gunn Lab

As a freshman, I had the good fortune to be selected as a Cornell Presidential Research Scholar, which made the process of finding a lab to work in particularly easy (it helps when you come with your own funding). To obtain a research position, a good way to start is to look through faculty websites and email professors whose research interests you. This is how I eventually found my way into the Gunn lab, where our research efforts mainly concern two genes (Attractin and Mahogunin) and the signaling pathways in which they are involved. These genes have pigmentation effects in mice as well as neurodegeneration and other interesting phenotypes.

A typical day in the lab might start off with a trip to the mouse facility to check if mice are pregnant, to set up breeding pairs, and/or to wean mice. Handling mice takes a bit of practice, but it becomes quite easy once you get accustomed to it. Upon returning to the lab, the rest of your day could be filled with a variety of activities. There may be some organs to dissect out from mice (the most popular are brains, livers, kidneys, hearts, and lungs). These organs would then go on to be embedded in either paraffin or tissue freezing media and then sectioned onto glass slides. The slides could then be stained to visualize certain cell types or organelles, or where a specific gene is being expressed. Dissected organs could also be used to obtain DNA or RNA to use in other experiments.

Less “mousy” activities are also very common in the Gunn lab. PCRs (polymerase chain reactions) are often set up to amplify specific segments of DNA. The results of a PCR could then be checked by running out some of the PCR product on a gel. Gel electrophoresis separates DNA (or RNA or protein) based on size, so a long segment of DNA could be distinguished from a short one. This process involves heating and pouring the gel (which has the consistency of Jell-O), letting it cool, and then adding the sample(s) into small wells in the gel and applying an electric field. Afterwards, UV light can be used to visualize the results. Another common lab procedure is cloning – not making identical copies of entire organisms, but rather making identical copies of a certain gene by inserting it into a vector (a circular piece of DNA) and then transforming bacteria with the vector. By letting the bacteria grow and divide, the vector will also be replicated, so there are many copies of your gene. Aside from these few basic procedures, there are numerous other protocols going on in the Gunn lab as well. The main goal of these methods is to characterize Attractin and Mahogunin (and a few other genes too) by identifying the biochemical pathways in which they function. In addition, a few people in the Gunn lab are working on a completely different project regarding the genetics of cardiac arrhythmias in dogs.

Aside from learning a lot about doing research in biology, working in a lab also provides great insight about oneself. I have learned that I would definitely like to continue researching and make it a major part of my future career. Additionally, I have discovered that I have an extraordinarily high tolerance for grossness (i.e. mouse dissections and such), and that I just have bad luck on Fridays. I’ve yet to learn on which day of the week I have good luck.

Finally, doing research full-time over the summer is very different than working part-time during the school year, when research has to compete with classes, homework, and tests. The summer is a more relaxed atmosphere, and taking trips down to the Biotechnology building to drop off samples is enjoyable due to the warm weather, whereas in the winter…not so fun. And although it is more relaxed, the summer is a great time to get things done since you can be at the lab every day without having to worry at all about schoolwork.

An Interview with Dr. Teresa Gunn

Dr. Teresa Gunn, assistant professor at the Department of Biomedical Sciences in the College of Veterinary Medicine focuses her research work to understand neurodegenerative diseases in humans using mutant mice as models. Ever since she was an undergraduate at the McGill University in Canada, her interest in science was in mammalian development and she majored in mammalian genetics. She further pursued her love for science by attending University of British Columbia for her doctoral work. During this time, working in the lab of Dr. Diana Juriloff, she discovered the defective genetic basis of exencephaly or neural tube closer defect. The defect occurs during embryonic development, when the skull fails to close exposing the brain to the outside environment leading to death of the embryo. Much of her graduate work dealt with identifying and characterizing the many genes associated with exencephaly using mice model.

In the year 1995, she came to the United States for her post-doctoral training at the laboratory of Gregory Barsh at Stanford University. Here, she identified the gene mutated in mahogany mutant mice as Attractin and studied its role in pigmentation and body weight regulation. When she came to Cornell University as an assistant professor in 2001, she sought to continue her work with the mahogany mutant mice. Since then, her research at Cornell has been to characterize the mutated genes in these mice, and answering the question, how does the mutation cause the various phenotypical abnormalities seen in these animals.

Mice with mutations in the Attracti1n (Atrn1; formerly named mahogany) and Mahogunin ring finger 1 (Mgrn1; formerly named mahoganoid) genes have dark coats, due to a role in the Agouti-melanocortin signaling pathway that regulates whether melanocytes produce black or yellow pigment. Other than coat color and slightly smaller body size then normal mice, mutants are not much different than wild type in appearance. In her later works, Dr. Gunn and the members of her lab have found that, besides having pigmentation and possible body weight regulation roles, the mutation also leads to adult-onset neourodegeneration in the mouse. The degenerations of the brain cells in these animals are clearly visible in light microscopic view, as the animal gets older. In the Atrn1 mutant mice, the neurodegeneration is accompanied by hyperactivity and weight loss.

The gene that is mutated in mahogany mice, Atrn1, encodes a type I transmembrane protein that functions as an accessory receptor for Agouti protein. Agouti proteins play a crucial role in switching from eumelanin (brown/black) to pheomelanin (red/yellow) synthesis during hair growth. The coat color mutations mahogany and mahoganoid prevent hair follicle melanocytes from responding to Agouti protein, because of the mutation in Atrn1. However, the gene mutated in mahoganoid mice, Mgrn1 encodes a protein with ubiquitin ligase activity, more specifically E3 ubiquitin ligase, which is important in protein turnover, degradation and trafficking. It is believed that the Atrn1 and Mgrn1 signaling pathways are conserved and are closely related. In order to understand the mechanisms of the mutations, the members of the Gunn’s lab use genetic, molecular, biochemical and proteomics approaches to identify new proteins and components involved in the Attractin and Mahogunin signaling pathway. Answering the question of how these mutations contribute to brown/black coating of the mice at the same time causing in neurodegeneration in the brain, could lend clues to the underlining basis for some of the human neurodegenerative diseases.