Cellular proteins form complex networks of interactions with other proteins. Although it is clear that these networks are dynamic structures that change in response to environmental signals and that are altered in disease states, our ability to assess the temporal changes in the networks is still at an early stage. Malaria-infected RBCs are a simple model in which to study changes in protein interaction networks over time. RBCs are highly differentiated cells that lack organelles and that contain a limited set of proteins. During infection, malaria parasites export proteins into the RBC that dramatically alter the properties of the host cell. Because RBCs lack nuclei, and thus cannot respond to parasite infection by altering their protein content, the changes that occur in the RBC phenotype must be due to the approximately 300 malaria proteins that are exported into the RBC. The exported malaria proteins are thought to bind to RBC proteins and, in this way, act as lesions that cause perturbations in the cellular protein interaction network. Since detailed information about the timing of expression of most malaria genes is already available, we can model changes in the cellular protein interaction network over time and correlate those temporal changes in the malaria-RBC interactome with changes in cellular properties. However, functional information about these malaria proteins is scarce and only a very limited number of interactions between exported malaria proteins and RBC proteins have been reported. The overall goal of this research is to elucidate, validate, and temporally model the network of protein-protein interactions between the malaria parasite Plasmodium falciparum and RBC proteins. A combination of new yeast strains, improved yeast two-hybrid screening methodology, and two complementary yeast two-hybrid screening approaches will be used to create a high-quality malaria-RBC protein interaction network. Interactions will be validated by GST pull downs and the split-luciferase assay, which we develop in this project for high-throughput analysis of protein-protein interactions in yeast. From this data we will develop a confidence score for each interaction. The malaria-RBC protein interactions will then be integrated with existing gene expression data to develop a time-dependent view of the malaria-RBC interactome. Using the results of our model as a guide, we will investigate the function of temporally distinct malaria-RBC protein interactions using the resealed RBC ghosts and ektacytometry. This project is innovative in its application and development of technologies to study the malaria-RBC protein interaction network. The data generated from this project is relevant not only to our understanding of interactome dynamics but also for understanding how malaria parasites cause disease.

Public Health Relevance

Proteins are linked to each other by a complex network of interactions. To better understand how these networks change over time and in disease states, we will develop a temporal model of protein- protein interactions in malaria-infected red blood cells. This data is generally relevant to our understanding of cellular processes and specifically relevant to how malaria parasites cause disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM092829-04
Application #
8477209
Study Section
Genomics, Computational Biology and Technology Study Section (GCAT)
Program Officer
Sakalian, Michael
Project Start
2010-06-15
Project End
2014-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
4
Fiscal Year
2013
Total Cost
$273,324
Indirect Cost
$87,031
Name
Purdue University
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907
Dolan, Patrick T; Zhang, Chaoying; Khadka, Sudip et al. (2013) Identification and comparative analysis of hepatitis C virus-host cell protein interactions. Mol Biosyst 9:3199-209
Lee, Shaoying; Salwinski, Lukasz; Zhang, Chaoying et al. (2011) An integrated approach to elucidate the intra-viral and viral-cellular protein interaction networks of a gamma-herpesvirus. PLoS Pathog 7:e1002297
Brown, Hakeenah F; Wang, Ling; Khadka, Sudip et al. (2011) A densely overlapping gene fragmentation approach improves yeast two-hybrid screens for Plasmodium falciparum proteins. Mol Biochem Parasitol 178:56-9
Kilili, Geoffrey K; LaCount, Douglas J (2011) An erythrocyte cytoskeleton-binding motif in exported Plasmodium falciparum proteins. Eukaryot Cell 10:1439-47
Khadka, Sudip; Vangeloff, Abbey D; Zhang, Chaoying et al. (2011) A physical interaction network of dengue virus and human proteins. Mol Cell Proteomics 10:M111.012187
Das, Sujaan; Shevade, Saudamini; LaCount, Douglas J et al. (2011) Plasmodium falciparum enolase complements yeast enolase functions and associates with the parasite food vacuole. Mol Biochem Parasitol 179:8-17