Defining a common mechanism of SAMHD1 function in cancer, HIV, and AGS

Project Summary As of 2019 there were 38.0 million individuals living with HIV globally. This number continues to rise yearly despite advances in HIV treatment. A significant challenge to developing a cure for HIV continues to be the reservoir of latently infected cells.

Current antiretrovirals cannot target this subset of non-dividing cells that harbor transcriptionally inactive provirus.

It is well established that HIV infection in non-dividing cells has different properties than HIV infection in dividing cells.

For example, non-dividing cells harbor dNTP concentrations below the Km of reverse transcriptase which significantly slows the kinetics of viral replication. SAM and HD domain containing protein 1 (SAMHD1) is the host dNTPase responsible for this restriction.

A thorough understanding of the contribution of SAMHD1 to the unique environment of HIV-1 infection in non-dividing reservoir cells has the potential to inform novel therapeutics to target this reservoir.

To this effect, I propose to characterize a novel series of SAMHD1 mutations that have been implicated in several cancers and use these mutants as tools to further probe the role of SAMHD1 in HIV-1 restriction.

I have identified a leukemia and colon cancer associated SAMHD1 mutant, R366C/H, that retains protein stability but loses dNTPase activity.

Interactions with cell cycle proteins and involvement in double strand DNA break repair, dNTPase independent functions of SAMHD1, are preserved in this mutant.

Interestingly, SAMHD1 mutations that result in impaired dNTPase activity were first reported in Aicardi Goutières Syndrome (AGS), a rare inherited encephalopathy characterized by aberrant type 1 interferon production.

Many of these AGS mutants are unable to restrict HIV-1 infection in macrophages due to their inability to deplete cellular dNTPs.

Given that the dNTP concentration in cancer cells is 6-11 fold higher than in normal cells and that SAMHD1 knock-out results in cancer cell phenotypes such as increased proliferation and reduced apoptosis, it is likely that the role of SAMHD1 as a dNTPase also involves the enzyme in cancer.

Therefore, I hypothesize that the dNTPase deficiency of the R366C/H SAMHD1 cancer mutant contributes to cancer cell phenotypes and abrogates both HIV-1 restriction in non-dividing cells and innate immune response suppression.

In Aim 1, I will complete my functional characterization of the R366C/H cancer mutant and determine the impact of these functional deficits on cancer cell phenotypes using biochemical and cell biology techniques.

Subsequently in Aim 2, I will use virology, cell biology, and immunology approaches to analyze the effect of the R366C/H cancer mutation on HIV-1 restriction and innate immune suppression, thus using this cancer mutation to probe more traditionally studied SAMHD1 disease phenotypes .

Collectively, these complimentary aims will highlight the role of SAMHD1 dNTPase activity in cancer and mechanistically link the involvement of SAMHD1 in cancer, HIV-1 restriction, and AGS.