September 12, 2018
Suppressing HIV in infected cells
Transcription gene silencing for HIV in macrophages (photo)
prevent the virus from spreading.
For the past two decades, better education has consistently dropped annual HIV infection rates, while better treatment has consistently extended the live span of infected patients. However, current treatment does not clear the virus, and patients must undergo regular antiviral therapy to keep the virus dormant. Using iPS cell technology, scientists at CiRA report a new treatment strategy based on transcription gene silencing that can halt HIV replication in infected cells without regular treatment.
As a retrovirus, HIV is made of RNA, but it tricks an infected cell to convert the RNA into DNA, which then incorporate into the cell's own DNA. Thus, as the cell expresses its DNA, it also expresses HIV DNA. CiRA Associate Professor Shin Kaneko, who led the study, has been investigating molecules that can be used to stop this expression.
"Transcription gene silencing uses short hairpin RNA to bind to a DNA promoter to prevent gene expression," he explains.
Short hairpin RNA (shRNA) are found in all cells. Their function is to change the structure of DNA or RNA, thus preventing expression of the gene. By transfecting shRNA that bind to HIV DNA, Kaneko and his research team could prevent HIV expression in infected cells without affecting the expression of other genes.
"We transfected iPS cells with shRNA for the HIV promoter and differentiated them to macrophages. HIV DNA was present in the macrophages, but HIV did not replicate," he says.
Macrophages are one of two types of immune cells that HIV infects. By transfecting iPS cells and not macrophages directly, the scientists could take advantage of the high proliferation rate of iPS cells to make a large number of transfected macrophages for observation.
Unlike those macrophages, macrophages made from infected iPS cells that did not receive the shRNA transfection acted like HIV factories, allowing the virus to replicate and thrive.
Further study revealed how the HIV DNA was modified.
"There was an enrichment of H3K27me3 and failure of enrichment of H3K9Ac," says Kaneko, describing changes in the methylation and acetylation of the DNA, two common modifications that a cell naturally uses to regulate whether a gene is expressed.
Importantly, the shRNA did not bind to other DNA, assuring that only the HIV DNA was not expressed, suggesting macrophage function was unaffected.
"Many gene therapies aim to prevent HIV infection. Our method suggests a therapy for immune cells already infected by HIV," says Kaneko.
- Journal: Molecular Therapy Nucleic Acids
- Title: Generation of HIV-resistant macrophages from iPSCs by using transcriptional gene silencing and promoter-targeted RNA
- Authors: Kei Higaki1, Masako Hirao1, Ai Kawana-Tachikawa2, Shoichi Iriguchi1, Ayako Kumagai1, Norihiro Ueda1, Wang Bo1, Sanae Kamibayashi1, Akira Watanabe1, Hiromitsu Nakauchi3,4, Kazuo Suzuki6, and Shin Kaneko1
- Author Affiliations:
- Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
- AID Research Center, National Institute of Infectious Diseases, Tokyo, Japan
- Institute of Medical Science, University of Tokyo, Tokyo, Japan
- Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA, USA
- St. Vincent's Centre for Applied Medical Research, St. Vincent's Hospital, Darlinghurst, Australia