PhD PROPOSAL FOR THE PASTEUR-PARIS UNIVERSITY INTERNATIONAL PROGRAM

 

Deadline for full application: December 15th, 2013

Interviews: March, 2014

Start of the Ph.D.: October 1st, 2014

 

 

 

Department: Department of Parasitology and Mycology

Title of the PhD project: Signaling pathways required for erythrocyte invasion by malaria parasites

Name of the lab: Malaria Parasite Biology and Vaccines

Head of the lab: Dr. Chetan E. Chitnis

PhD advisor: Dr. Chetan E. Chitnis

Email address: cchitnis@gmail.com

Web site address of the lab:

Doctoral school affiliation and University: Pasteur-Paris University International Program

 

Presentation of the laboratory and its research topics:

 

The Malaria Parasite Biology and Vaccines Unit will be established in the Department of Parasitology and Mycology at Institut Pasteur, Paris in 2014. The main focus of the laboratory will be to understand the molecular mechanisms that mediate the invasion of erythrocytes by malaria parasites. Erythrocyte invasion is a multi-step process that requires multiple, specific molecular interactions between the target host cell and invading parasite. We study the receptor-ligand interactions involved in the process of erythrocyte invasion by Plasmodium merozoites. We also study the signaling mechanisms that regulate and co-ordinate key steps such as microneme and rhoptry discharge during the complex process of invasion. These basic studies have the potential to lead to identification of new drug targets as well as vaccine candidates. We undertake translational research to develop such potential leads into novel therapeutic and prophylactic strategies to protect against malaria.

 

Description of the project:

 

Our laboratory is interested in understanding the molecular mechanisms involved in invasion of red blood cells by malaria parasites. The invasion of red cells is a complex process that is mediated by a cascade of molecular interactions between host receptors and parasite protein ligands (Gaur and Chitnis, 2011). Interestingly, many of the parasite proteins that mediate these interactions are not found on the surface of Plasmodium merozoites but in apical organelles referred to as micronemes and rhoptries. For example, the erythrocyte binding antigen EBA175 and apical merozoite antigen-1 (AMA1) is found in micronemes whereas the PfRH proteins, PfRH2 and PfRH5, are found in rhoptries. Over the past few years, we have identified the external signals that trigger the timely secretion of microneme and rhoptry proteins to the merozoite surface during invasion (Sharma and Chitnis, 2013). We have demonstrated that exposure of merozoites to a low K+ environment as found in blood plasma leads to a rise in cytosolic Ca2, which triggers secretion of microneme proteins such as EBA175 and AMA1 (Singh et al., 2010). Subsequently, the binding of EBA175 with its red cell receptor, glycophorin A (gly A), triggers the release of rhoptry proteins (Singh et al., 2010). While we have now identified the external signals that trigger microneme and rhoptry release, the signaling pathways involved in regulating these processes are not yet understood. Some of the questions that we are currently addressing are as follows: How does exposure of merozoites to a low K+ environment lead to a rise in cytosolic Ca2+ in Plasmodium merozoites? How does the rise in cytosolic Ca2+ trigger microneme release? How does the binding of EBA175 with its receptor gly A trigger release of rhoptry proteins?

 

The project offered for Ph. D. dissertation research will focus on understanding the downstream signaling pathways that lead to microneme discharge in response to a rise in cytosolic Ca2+ in P. falciparum merozoites. What are the key Ca2+ responsive effectors that regulate microneme secretion? We have demonstrated that P. falciparum calcium-dependent protein kinase 1, PfCDPK1 (Bansal et al., 2012), and calcium-dependent phosphatase, calcineurin (Singh et al., 2013), mediate regulation of microneme secretion in merozoites. Inhibition of PfCDPK1 and calcineurin with pharmacological agents leads to a block in microneme discharge (Bansal et al., 2012 and Singh et al., 2013). In this project we will use genetic, pharmacological and proteomic approaches to study the functional roles of such downstream Ca2+ responsive effectors in regulation of microneme discharge. For example, to evaluate the functional role of calcineurin in microneme secretion, we will transfect P. falciparum blood stage parasites to express a mutant calcineurin that is resistant to cyclosporin A (CsA). Treatment of such transgenic parasites with CsA should not inhibit microneme discharge in contrast to wild type P. falciparum confirming role of calcineurin in this process. We will also overexpress the regulatory subunit of calcineurin, CnB, under control of a regulatable promoter to inhibit calcineurin activity. We will examine if overexpression of CnB in merozoites inhibits microneme discharge. We will use a phosphoproteomic approach to identify the substrates of calcineurin in P. falciparum merozoites that mediate microneme discharge. P. falciparum merozoites will be transferred from high K+ to low K+ buffers in presence or absence of calcineurin inhibitors. Changes in phosphorylation of merozoite proteins will be identified by determining changes in the global phosphoproteome. Analysis of the changes in global phosphoproteome should identify both the signaling pathways involved in microneme discharge as well as the protein substrates of calcineurin. These studies will provide information on the downstream signaling mechanisms involved in microneme secretion by P. falciparum merozoites during erythrocyte invasion. 

 

References:

 

Bansal A, Singh S, More KR, Hans D, Nangalia K, Yogavel M, Sharma A, Chitnis CE. 2012. Characterization of Plasmodium falciparum calcium-dependent protein kinase 1 (PfCDPK1) and its role in microneme secretion during erythrocyte invasion. J Biol Chem. 288(3):1590-1602.

 

Gaur D and Chitnis CE. 2011. Molecular interactions and signaling mechanisms during erythrocyte invasion by malaria parasites. Curr Opin Microbiol. 14:422-428.

 

Sharma P and Chitnis CE. 2013. Key molecular events during host cell invasion by Apicomplexan pathogens. Curr Opin Microbiol. 16:432-427.

 

Singh S, Alam MM, Pal-Bhowmick I, Brzostowski JA, Chitnis CE. 2010. Distinct external signals trigger sequential release of apical organelles during erythrocyte invasion by malaria parasites. PLoS Pathog. 6(2):e1000746.

 

Singh S, More KR, Chitnis CE. 2013. Role of calcineurin and actin dynamics in regulated secretion of microneme proteins in Plasmodium falciparum merozoites during erythrocyte invasion. Cell. Microbiol. doi: 10.1111/cmi.12177.

 

Keywords:

 

Malaria, Host-parasite interactions, Host cell invasion, Receptor-ligand interactions, Signaling, Regulated vesicle secretion

 

Expected profile of the candidate (optional):

 

The candidate should have an undergraduate degree in the life sciences with a strong background in biochemistry, genetics, cell biology or molecular biology. Experience in laboratory research and familiarity with wide range of basic biochemistry, cell culture, cell biology and molecular biology techniques is preferred.

 

Contact: Dr. Chetan E. Chitnis at cchitnis@gmail.com

 

Mis à jour le 16/09/2013