Silencing HIV
Viral suppression via DNA methylation
Background
Human Immunodeficiency Virus (HIV) is a devastating pathogen that infects people for life and slowly destroys their immune system. Fortunately, there have been treatments developed in recent years that do a pretty good job of counteracting HIV’s spread through the body. However, they are not perfect, and the virus can still develop resistance to these standard therapies. In this article, a research group in California describes their work to develop a new kind of therapy for HIV, one that attacks it at its root. But before we dive into the paper, we need to talk about how HIV invades your cells, and why it is so hard to treat.
Bad DNA
As many of you may know, HIV belongs to a class of pathogens known as retroviruses. The “retro” part of the word means “reverse”, making these literally the “reverse-viruses.” This name comes from their ability to engage in reverse transcription, a biological activity that is not very common but extraordinarily powerful. As you also probably remember, your cells operate in the following manner:
DNA → RNA → Protein
But, in reverse transcription, the flow is reversed, and instead you have:
RNA → DNA
But what does it mean that retroviruses like HIV can do this “in reverse”, and how does it affect disease?
When HIV invades a cell, it copies its own genetic instructions using a special reverse transcriptase protein, converting RNA into DNA. This is the step that makes HIV a retrovirus. Then, using another protein known as an integrase, it inserts its own DNA into the genome of the cell it is attacking. If this written description isn’t working for you, here’s an excellent diagram for you to reference.
That integration step is critical, because it is the reason why HIV is so difficult to treat. Once the HIV DNA is incorporated, it becomes a permanent part of the host DNA, and there’s no way for the cell to remove it. On top of that, the HIV DNA can be transcribed after it has integrated into the genome, a process which generates new viruses to infect new cells. Because of the virus’s ability to “hide” in the host’s DNA, individuals who contract HIV are infected for life. While they can take medication to fight off the virus circulating in their body, they cannot attack the virus that has already integrated into their DNA. That is…until now.
Summary: Taking Out Bad DNA (but not removing it)
In order to tackle the problem of HIV infection, this research team took advantage of a form of genetic modification that already exists in human cells: DNA methylation.
DNA methylation falls into the category of epigenetics — genetic changes that alter the function of DNA without irreversibly changing its code. The process is very simple. A protein known very creatively as “DNA methylase” adds methyl groups (carbon bonded to three hydrogens) to stretches of DNA to which it is targeted. If you want to see the mechanism for the methylation of cytosine, click here
When a section of DNA is methylated, it is silenced, meaning it is no longer transcribed. The team of researchers in this study took advantage of this mechanism by creating proteins specifically designed to target and methylate HIV DNA, effectively shutting it down. They started with a group of slightly different fusion proteins and then narrowed their focus to the most promising candidates. For simplicity, we’re going to focus on just one of these candidate proteins: ZPAMt.
You can think of each domain of this protein as a specialized targeting system. First there is the nuclear localization signal (NLS) that targets the protein to the nucleus. Then, there is the zinc finger domain (ZFP), which was specifically designed to bind to any HIV DNA in the genome. Finally, there is the methylase domain, which signals the cell to methylate the strand of HIV DNA that the fusion protein is bound to.
Nuclear localization signal, zinc finger domain, methylase
To test their design, they treated an HIV-infected human cell line with the mRNA for their fusion protein. As you can imagine, they couldn’t just dose a cell with plain mRNA, so they had to first package it into exosomes, which are then able to fuse with the target cells membrane and deliver the mRNA therapy inside the cell. This strategy is common in genetic therapies, and is similar to the strategy used for the COVID-19 mRNA vaccines.
When the team measured the expression of HIV RNA markers over a period of two months, they found that the cells treated with ZPAMt had a steep drop in their RNA measurements as compared to the control. This data is presented in figure 2g which you can find here.
In my opinion, their most impressive data is in figure 4. They infected mice with HIV, then treated them several days later with both the traditional therapy regimen + ZPAMt. They sampled viral RNA every day after beginning the treatment, and found that, by day 10, the combined regimen with ZPAMt decreased viral load by over a thousand fold! That data is reported in figure 4c which I’ve linked here.
As always with these summaries, the paper describes many more experiments that further explore and validate the fusion proteins, and I would encourage you to look through more of their data to test your ability to read primary research. This paper represents an impressive new breakthrough in the field of HIV research that will hopefully help in the development of similar therapies that work to repress HIV DNA transcription and permanently cure those chronically infected.
