Quartz Crystal Microbalance (QCM-D) Confirms Specific GDI-GINT3 Interaction
Connie Yu Heng Cheng ('09), Bob Rawle ('08), Clarissa Cheney, Malkiat Johal
Interdisciplinary Departments: Biology, Chemistry
Rab GTPases direct vesicles to the correct membranes and organelles. After membrane fusion and release of vesicular contents, rabs are left embedded in the target membranes. GDI (GDP Dissociation Inhibitor) is a protein that extracts rabs from target membranes and returns rabs to donor membranes. The GDI-Rab interaction is hypothesized to be mediated by receptors, namely GDI Displacement Factors (GDF) and Rab Release Factors (RRF). GDFs and RRFs help GDI extract and release rabs to the correct membranes. One potential GDF or RRF called Gint3 has been identified by the Cheney Lab using the Yeast Two Hybrid System. The Quartz Crystal Microbalance with Dissipation (QCM-D) was used to examine the interaction between the two fusion proteins GST-GDI and GST-Gint3, purified from E.coli. The QCM-D passes an electric current through the quartz crystal, which oscillates at a particular frequency. Changes in frequency of oscillation are then measured as molecules deposit onto the crystal’s surface. The machine also measures the dissipation of substances deposited on the crystal. Dissipation, a measure of the film’s softness, can help characterize the nature of inter-molecular interactions. After investigating the interactions between GST-GDI, GST-Gint3, and several control proteins, it was determined that GST-GDI and GST-Gint3 bind specifically.
Funding provided by: Rose Hills Foundation
Identifying Drosophila RABGDI Interactors by Tandem Affinity Purification - Mass Spectrometry
Palak Amin ('08), Clarissa Cheney
Interdisciplinary Departments: Biology, Molecular Biology
RabGTPases help regulate vesicle transport and direct vesicles to the correct cellular location. GDI (GDP disassociation inhibitor) then extracts inactive GDP-bound Rabs from target membranes and recycles them back to donor membranes. The specificity with which a variety of different Rabs are returned to their correct donor membrane by a single GDI in Drosophila suggests that other proteins are involved in the process, as does the discovery of a GDF (GDI disassociation factor), which catalyzes the disassociation of the stable Rab-GDI complex. To identify novel potential GDI interactors, transgenic Drosophila lines with affinity-tagged GDI genes under a UAS Gal4 promoter have been established. The lines are pUASp Tetra- GDI, pUASp GDI-Tetra, pUASp Tetra, pUASp GST- GDI, pUASp GST, and pUASp GDI. Crossing these flies with Gal4 drivers results in transcription of tagged GDI in specific tissues at specific stages of development. Western Blotting shows that GST-GDI is being expressed in one of these lines, as expected. Tandem affinity purification will be used to isolate tagged GDI – GDI interactor complexes, and the GDI interactors can then be identified by mass spectrometry. The effects of over-expressing GDI can also be determined.
Funding provided by: HHMI, SURP (PA); Hirsch, NSF-RUI (CC)
Exploring Burgeoning Civil Society in West Africa: A Look at the Challenges Local Women's NGOs Face in Senegal and Burkina Faso
Rebecca Troeger ('08), Pierre Englebert
Interdisciplinary Departments: Gender Studies, International Relations, African Studies
My project’s aim was to examine the management and resources of local West African women-run NGOs that serve women in development. My method of study involved a combination of observation and interviewing. I interned with two women who direct such NGOs in Dakar, Senegal and Ouagadougou, Burkina Faso, respectively. I also conducted interviews with employees and directors of local and international NGOs and donor organizations such as foundations, embassies, and other government agencies. During this process, I discovered some important issues that I was not aware of before the start of my project. These include the complex relationship that exists between civil society and government in Senegal and Burkina Faso and the effect of the 2005 Paris Declaration on Aid Effectiveness on the current reality of local women’s NGOs and relations between civil society and government. My true hope with this project is to provide useful information and constructive criticism to both NGO directors and donors. One observation I offer along these lines is the need for less competition and more partnerships and communication between local NGOs; I believe this change would increase the efficiency of both individual local NGOs and civil society. I also propose that donor organizations must practice increased transparency in their requirements for grant applicants and in the grant application process that local organizations must go through.
Funding provided by: SURP (Richter), Oldenborg International Center
Induction of Apoptosis Is Inversely Related to DNA Damage in Yeast with Phenolic Compounds
Matthew Louis Goldman ('08), Steven Oh (’06), David Dorsey (’07), Tina Negritto, Cynthia Selassie
Interdisciplinary Departments: Molecular Biology, Chemistry
Frequently used food preservatives such as BHA and BHT, which inhibit autooxidation of food by forming stable radicals that would typically reduce oxidative stress in the body, have been shown to promote tumor growth, possibly due to DNA damage or due to inhibition of a cell’s ability to undergo apoptosis. Yeast cells were recently shown to undergo apoptosis in response to cell stresses such as treatment with hydrogen peroxide, which makes yeast a suitable model system for studying induction of apoptosis in eukaryotic cells. The DEL recombination assay can be used to determine the extent of DNA damage, and D2R and TUNEL assays detect apoptotic phenotypes in yeast cells treated with a range of phenolic compounds. Therefore, it is possible to compare DNA damage with the induction of apoptosis in order to better understand the mechanism by which these phenols damage yeast cells. Previous research by Steven Oh ’06 and David Dorsey ’07 has shown that BHA and BHT increase recombination frequency in the X40 yeast strain. Preliminary results of the current project show that BHA and BHT quench apoptosis in the RS112 yeast strain, which suggests that an increase in recombination frequency correlates to a decrease in induction of apoptosis.
Funding provided by: HHMI
The N400 as a Measure of Cultural Differences in Contexual Information Processing
Alicia Yee ('08), Yumi Ando ('08), Carol Huang ('08), Richard Lewis, Sharon Goto
Interdisciplinary Departments: Psychology, Neuroscience
Culture can significantly influence the way information is processed. Individuals from collectivist cultures have been shown to be more sensitive to contextual features of the environment than are those from individualist cultures. The purpose of our study is to investigate neural mechanisms of context that are influenced by culture. Electrophysiology can provide certain benefits to the study of social, cognitive, and cultural processes. These benefits include providing an independent, biological measure of psychological experience that may be sensitive to subtle changes in cognitive processes and may be less susceptible to bias than are self-report measures. They can also provide an on-line measure of activity that can differentiate stages of processing information. We are presenting subjects with complex pictures and using the N400 as an index of semantic expectancy to test for cultural differences in processing contextual information. We will present our results at the annual meeting of the Cognitive Neuroscience Society in April.
Funding provided by: SURP (Richter)