Traitor-virus-guided discovery of antiviral factors

Introduction

Viruses may seem smart since they rapidly develop new skills to spread in humans. However, they are actually just masters of trial and error. Their short generation time, enormous reproduction, and high variability allow viruses to try countless variations. A few of these will enhance viral spread and, in the worst case, enable viral pandemics.

Evolution of Viruses

It is conceivable that viruses evolve to counteract those defense mechanisms that would otherwise be most effective against them. Usually, it is hard to assess why specific changes are advantageous. This is especially true for adaptations enabling viral pathogens to counteract innate antiviral factors because these cellular proteins and their viral antagonists are numerous and highly versatile.

Proposed Approach

Here, I propose to combine the advantages of the revolutionary CRISPR/Cas9 technology with the enormous adaptive power of viruses to develop a novel approach allowing robust, specific, and genome-wide unmasking of antiviral mechanisms.

Methodology

In principle, we will equip HIV-1 with genetic scissors to convert them into traitor viruses revealing their cellular opponents. To achieve this, we will:

1. Generate libraries of replication-competent HIV-1 constructs expressing guide RNAs as genetic tools with the potential to inactivate every human gene in Cas9 expressing cells.
2. Select virus variants expressing guide RNAs eliminating antiviral genes by in vitro passaging.
3. Identify these variants by next generation sequencing.

Utilization of different viral backbones will allow the discovery of key defense factors against viral zoonoses and spread in humans.

Goals

Finally, we will determine the antiviral spectrum and inducibility of innate antiviral factors to identify vulnerabilities of viral pathogens that can be exploited in preventive and therapeutic approaches. The project will establish and apply an innovative, highly versatile approach to uncover our antiviral defense mechanisms with the ultimate goal of achieving better control of viral pathogens.