Designing a novel panel of transcriptome assays to study the the viral mechanisms of of HIV-2

Scientist's Name: Michael Lu

Marin Academy Research Collaborative Program

Corias, Nature 2009

Past

What is HIV

Human Immunodeficiency Syndrome (HIV) was discovered as the infectious agent of acquired immune deficiency syndrome (AIDS) during 1981 (1, 2). Human Immunodeficiency Syndrome infects the CD4+ T-cells of the human immune system, destroying them, and crippling our body’s natural defense from diseases (3). There are two subtypes of HIV, HIV-1, and HIV-2, the latter of which is characterized as being much less pathogenic than HIV-1. For example, HIV-2 transmission rates are lower and infections are less likely to progress to AIDS (4). However, since HIV-2 has been concentrated in West Africa, affecting fewer people, the virus has not been as well-studied (5).

Importance of HIV latency

HIV has yet to be cured as the virus can persist in resting CD4+ T-cells (also known as the latent HIV reservoir) despite combination Antiretroviral Therapy (6, 7). The HIV reservoir that exists in these CD4+ T-cells has a long half-life and persists through a patient’s life (8, 9). Latency is caused by infected cells that are unable to complete their life cycle due to mutations and transcriptional blocks (10).

Past Research

Dr. Yukl and other members of his lab at the San Francisco Veterans Association Medical Center published a paper in 2018 in which they designed a panel of transcriptome assays for HIV-1 and used them to study the expression of specific HIV-1 RNA species in infected patient cells. Their results suggested the existence of blocks to HIV-1 elongation, completion, and splicing which prevents the virus from finishing its life cycle, causing the formation of latent reservoirs.

Barré-Sinoussi, F. et al. Isolation of a T‑lymphotropic retrovirus from a patient at risk for acquired immune deficiency syndrome (AIDS). Science 220, 868–871 (1983).
Centers for Disease Control and Prevention. Pneumocystis pneumonia — Los Angeles. MMWR Morb. Mortal Wkly Rep. 30, 250–252 (1981).
Loetscher, P., Moser, B. & Baggiolini, M. Chemokines and their receptors in lymphocyte traffic and HIV infection. Adv. Immunol. 74, 127–180 (2000).
Visseaux, Benoit et al. Hiv-2 molecular epidemiology. Infections, Genetics and Evolution Volume 46, 233-240 (2016).
Nyamweya, Samuel, et al. “Comparing HIV-1 and HIV-2 Infection: Lessons for Viral Immunopathogenesis.” Reviews in Medical Virology, 26 Feb. 2013, doi:10.1002/rmv.1739.
Pomerantz, R. J. Reservoirs of human immunodeficiency virus type 1: the main obstacles to viral eradication. Clin. Inf. Dis. 34, 91–97 (2002).
Shen, L. & Siliciano, R. F. Viral reservoirs, residual viremia, and the potential of highly active antiretroviral therapy to eradicate HIV infection.
Cohn, Lillian et al. HIV-1 integration landscape during latent and active infection. Cell. 2015 January 29; 160(3): 420–432.
Maldarelli, F. Specific HIV integration sites are linked to clonal expansion and persistence of infected cells. Science. 2014 July 11; 345(6193): 179–183.
Corias, Mayte et al. Understanding HIV‑1 latency provides clues for the eradication of long-term Reservoirs. Nature Reviews Volume 7, 798-812 (2009).

PRESENT

For my project, I am studying and comparing the transcriptional characteristics and HIV-2. I am working with Dr. Yukl and Dr. Kumar at the San Francisco Veterans Association Medical Center. We hypothesize that studying the transcriptome of HIV-2 will allow us to better understand the viral mechanisms which drive viral latency. We can then compare the similarities and differences between HIV-1 and HIV-2 to gain a better understanding of what makes HIV-2 less virulent. Currently, I am working on optimizing and validating a panel of new HIV-2 assays which will allow us to conduct future experiments with patient cells that are being provided by Dr. Athe Tsibris.

Dr. Steven Yukl and Dr. Nitasha Kumar

Mentors

Dr. Steve Yukl is an associate professor of medicine at UCFS and is the principal investigator HIV research lab at the SFVA. Dr. Nitasha Kumar is a post-doctorate fellow at the Yukl lab who developed an interest in HIV persistence in vivo during her graduate studies.

Droplet Digital PCR

ddPCR

One of the key instruments which I use in the majority of my experiments is Bio-Rad's Droplet Digital PCR platform. ddPCR is a form of quantitative PCR in which samples are fractioned into 20,000 droplets which are then amplified individually. This allows for increased PCR precision and absolute quantification.

Lab Work Spaces

Fume hoods and Bio-Safety Cabinets

I usually complete my lab work in fume hoods or biosafety cabinets to prevent any possible contamination. Workspaces are constantly cleaned with water, ethynol, and RNAse away which protects both the samples and ourselves.

Future

I hope to identify the presence or lack of transcriptional blocks to HIV-2. This data will tell us about the mechanisms which drive HIV-2 latency. We will also compare our data with other data gathered about HIV0-1 transcriptional blocks to identify similarities or differences between the two viruses.

My mentor and I hope to have my project done before I graduate or during the summer before college. However, I plan on continuing research in a lab setting in the future.