Using Cellular Proteins to Reveal Mechanisms of HIV Infection
A vital step in HIV infection is the insertion of viral DNA into the genome of the host cell. In order for the insertion to occur, viral nucleic acid must be transported through the membrane that separates the main cellular compartment (the cytoplasm) from the nucleus, where the host DNA is located. Scientists are actively studying the mechanism used to transport viral DNA into the nucleus in the hopes of targeting this step with future anti-HIV treatments. Up to this point, researchers have identified some of the viral components that play a role in nuclear transport, but they have not determined how viral interactions with other molecules in the cell contribute to the process.
To increase understanding of the HIV lifecycle, KyeongEun Lee, Ph.D., a research fellow in the CCR HIV Drug Resistance Program, has developed a method that uses proteins from immune cells to disrupt the infectious process of HIV. This facilitates detection of HIV interactions with cellular proteins and identification of potential targets for antiviral therapies. A report of Dr. Lee’s findings was recently published in Cell Host & Microbe.
Dr. Lee’s method involves screening all of the proteins expressed in mouse thymus cells for the ability to prevent HIV infection of cultured cells. This screen identified a protein called CPSF6-358, which rendered cultured human T cells resistant to HIV infection by preventing transport of HIV DNA into the nucleus of the cell. To identify how CPSF6-358 functions, the researchers exposed the virus to the protein and looked for mutations that allow the virus to evade inhibition by CPSF6-358. A single mutation was found in a viral protein called capsid protein.
Before HIV invades a cell, multiple copies of the capsid protein form a shell that encapsulates the genetic material of the virus. However, once inside the cell, the capsid performs other functions, including escorting the viral DNA into the nucleus. CPSF6-358 is able to prevent the transport of HIV DNA by binding to the capsid protein. While this interaction between CPSF6-358 and capsid protein does not occur in human T cells, the experiments provided useful insight into the mechanisms of HIV DNA transport.
The researchers then looked for human cellular proteins that interact with capsid protein. To accomplish this feat, cultured human cells were systematically altered to remove specific cellular proteins crucial for transporting molecules to the nucleus. From these experiments, several human proteins that interact with the capsid protein were identified, allowing for future studies to investigate the specific requirements for HIV DNA transport in human cells.
This method developed by Dr. Lee and her colleagues was able to identify viral proteins important in the transport of viral DNA to the nucleus and link the capsid protein to cellular components involved in the HIV lifecycle. Further study of the requirements for DNA transport are now possible and may lead to the development of new HIV therapies that prevent the integration of viral DNA into the cellular genome.Summary Posted: 04/2010
Lee K, Ambrose Z, Martin TD, Oztop I, Mulky A, Julias JG, Vandegraaff N, Baumann JG, Wang R, Yuen W, Takemura T, Shelton K, Taniuchi I, Li Y, Sodroski J, Littman DR, Coffin JM, Hughes SH, Unutmaz D, Engelman A, KewalRamani VN. Flexible use of nuclear import pathways by HIV-1. Cell Host Microbe. 2010 Mar 18;7(3):221-33. PubMed Link