Because human immunodeficiency virus 1 (HIV-1) has an RNA genome, it is no surprise that protein-RNA interactions play key roles both in the replication cycle of HIV-1 and in the host defense against HIV-1 infection. A better understanding of these processes will lay the foundation for next-generation therapies against HIV-1, a pressing need due to the emergence of drug-resistant HIV variants. In addition to its direct applications in sequencing of whole genomes, rapid advancements in next-generation sequencing have recently led to the development of novel assays that can identify the precise RNA sequences bound by a protein of interest in biological settings. My short-term goal is to apply these novel approaches to identify the targets of several RNA-binding proteins that are critical for HIV-1 replication. This will be the first time such techniques will have been applied n the study of viral protein-RNA interactions in a biologically relevant system.
The first aim of this project is to identify the exact viral and cellular RNA sequences bound by the HIV-1 structural protein, Gag, which selectively recruits viral genomic RNA from a large pool of cellular RNA. Neither the precise viral/cellular RNA sequences bound by Gag during the course of infection nor the contribution of cellular RNAs to virus assembly and infectivity are known. Therapeutic targeting of these specific Gag-RNA interactions could lead to substantial defects in virion assembly, given the importance of viral RNA in the structural integrity of the HIV particle.
The second aim i s to identify viral and cellular RNA sequences bound by a class of cellular proteins (APOBEC3F, 3G and 3H) that restrict HIV infection by leading to the hypermutation of the viral genomic DNA during reverse transcription. It is thought that viral or cellular RNA mediates the incorporation of APOBEC proteins into virus particles, a necessary step for their restriction activity. The RNA molecules identified herein can be utilized to target these extremely potent restriction factors into HIV-1 particles and therefore be the basis of new anti-retroviral therapies.
The third aim i s to employ the powerful techniques developed in this study to understand how two classes of host proteins regulate the splicing of HIV- 1 genes, a very poorly understood process. Once the molecular basis of this process is revealed, one can envision developing novel drugs that specifically target viral splicing. The initial phase of this research will be performed under the supervision of Dr. Paul D. Bieniasz, a leader in the field of retrovirology with outstanding track record of publications and mentorship. I have already gained a unique and broad set of intellectual as well as bench and bioinformatics skills, which will make me a highly competitive candidate for faculty positions. I do, however, plan to enhance my research skills and focus on career development activities as I am transitioning to independence. I will have the opportunity to do so, as the Aaron Diamond AIDS Research Center and Rockefeller University provide cutting-edge facilities and support for the execution of this project as well as career development. In addition, I will be able to interac with world- class scientists at Rockefeller University and collaborate with others who are part of the Center for RNA Studies, an NIH-funded collaboration. My long-term goal as an independent scientist is to apply the unique skill set that I will gain during the execution of this project to poorly understood areas of retrovirus molecular biology and host responses against retroviral infections. I expect that the knowledge gained by doing so will be translated into development of new therapies to cure or prevent retroviral and other pathogenic viral infections.

Public Health Relevance

Due to the emergence of drug resistance mutations during the course of anti-retroviral therapy against human immunodeficiency virus-1 (HIV-1), development of new therapies is a pressing need. This requires a better understanding of the molecular details of HIV-1 infection and host responses against it. I propose to employ novel techniques to unveil the details of protein-RNA interactions that are critical for HIV-1 replicatio, which can form the basis of next-generation therapies against HIV-1.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Career Transition Award (K22)
Project #
1K22AI116258-01
Application #
8847019
Study Section
Acquired Immunodeficiency Syndrome Research Review Committee (AIDS)
Program Officer
Church, Elizabeth S
Project Start
2015-03-15
Project End
2017-02-28
Budget Start
2015-03-15
Budget End
2016-02-29
Support Year
1
Fiscal Year
2015
Total Cost
Indirect Cost
Name
Washington University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Madison, Michaela K; Lawson, Dana Q; Elliott, Jennifer et al. (2017) Allosteric HIV-1 Integrase Inhibitors Lead to Premature Degradation of the Viral RNA Genome and Integrase in Target Cells. J Virol 91:
York, Ashley; Kutluay, Sebla B; Errando, Manel et al. (2016) The RNA Binding Specificity of Human APOBEC3 Proteins Resembles That of HIV-1 Nucleocapsid. PLoS Pathog 12:e1005833
Kessl, Jacques J; Kutluay, Sebla B; Townsend, Dana et al. (2016) HIV-1 Integrase Binds the Viral RNA Genome and Is Essential during Virion Morphogenesis. Cell 166:1257-1268.e12
Kutluay, Sebla B; Zang, Trinity; Blanco-Melo, Daniel et al. (2014) Global changes in the RNA binding specificity of HIV-1 gag regulate virion genesis. Cell 159:1096-1109