TMCnet Feature
January 27, 2022

How Researchers Can Use CRISPR Techniques in Gain-of-Function Studies to Prevent Pandemics

Biotechniques explains how researchers can employ CRISPR techniques in metagenomic studies to pre-empt and prepare for health crises.

The COVID-19 crisis has left scientific communities around the world questioning how we can trace and manage virus outbreaks in the event of future pandemics. These communities are using data and case studies from the coronavirus pandemic to prepare and strategize for future crises. Many have concluded that CRISPR (clustered regularly interspaced short palindromic repeats), a family of DNA sequences, offers huge potential in gain-of-function studies that can help researchers prepare for the emergence of a virus.

Researchers can use gain-of-function studies to induce new functions or screen for these functions. However, these studies can be controversial, and, although researchers called for gain-of-function studies to examine coronaviruses as early as 2016, the U.S. government blocked these calls. Here, the open-access journal BioTechniques examines the metagenomic studies that researchers would have undertaken had the U.S. government not issued a moratorium against these studies and how CRISPR can pave the way for the future of safer gain-of-function experiments.

Proposing Metagenomic Studies to Prevent Pandemics

Researchers often use metagenomics — the study of genetic material from environmental samples — to identify new viruses. This way, researchers can stunt the spread of a virus early and reduce the likelihood of a pandemic. For example, researchers had identified SARS-like viruses through metagenomic studies years before COVID-19 first emerged in Wuhan, China: In 2016, a paper published in PNAS warned of a SARS-like virus that was “poised for human emergence”. The paper noted that WIV1-CoV (which had been isolated from Chinese horseshoe bat populations in another metagenomic study) was at a high risk of emerging in humans. Previous research had demonstrated that chimeric and full-length zoonotic versions of the virus could replicate in vivo in humans.

The paper explained that while metagenomic screens had identified the circulation of SARS-like viruses among bat populations, previous studies had identified species that had the potential to evolve into human infectious strains. At the time, Vincent Racaniello, a professor at the Department of Microbiology and Immunology at Columbia University, concluded that “gazing at viral sequences has its limits; experiments need to be done”. So, the authors of the paper prepared an approach to examine metagenomic data. They planned to use this data to determine:

·       The probability of a virus becoming infectious to humans.

·       The probable severity of the virus.

·       Preparation steps to manage the emergent species.

To achieve this, the researchers conducted a series of mouse model tests that involved transgenic mice. Researchers manipulated the mice to express the human ACE2 receptor, which is the receptor bound by COVID-19 spike proteins.

The researchers performed these mouse model tests to examine our preparedness for the emergence of WIV1-CoV in human populations. They tested the effectiveness of antibodies generated to protect against SARS-CoV by blocking infection, first in cell culture and then in the mouse model. This analysis led to the identification of an antibody that prevented the replication of the virus. That said, initial efforts to identify whether vaccines containing inactivated SARS-CoV would be effective against WIV1-CoV proved negative.

Controversy Over Gain-of-Function Studies

As a result of this study, Racaniello made various recommendations to advance this research. He suggested that researchers should perform additional studies to:

·       Determine a panel of antibodies that can prevent the invasive action of the spike protein that is common with COVID-19.

·       Identify the genome alterations that WIV1-CoV would need to become infectious to humans.

·       Examine the mechanism by which the pathogenicity of coronaviruses could increase.

Investigating this metagenomic research further would have required researchers to complete gain-of-function studies on the viruses. But when PNAS published the paper in 2016, the publication triggered heated discussion over the risks associated with gain-of-function studies. These risks arise from the fact that gain-of-function experiments involve manipulating pathogens to make them more infectious and harmful so that researchers can design defenses against these.

Despite little evidence against the use of gain-of-function studies to test for coronaviruses, the U.S. government issued a moratorium, which restricted any experiments on the viruses. Although the government lifted this restriction in December 2017, critical time had passed. The impact of the paper had dampened, which may have prevented researchers from pursuing Racaniello’s recommendations.

The Use of CRISPR in Metagenomic Studies

The debate surrounding gain-of-function studies is still ongoing: While many researchers have been put off undertaking these studies, other researchers are keen to perform gain-of-function experiments with protocols in place to ensure they conduct studies in a safe, open, and accountable manner. Such protocols could take the heat out of the debate and enable invaluable progress in research.

As a rapidly evolving technology, CRISPR may prove essential to the future of safer gain-of-function studies. Researchers can use non-cutting CRISPR versions to modulate gene expression. They can fuse deactivated Cas9 to transcriptional activators and use the guide RNA to direct the Cas9 in a sequence-specific manner to promoter regions. This can lead to target gene activation of CRISPRa, which is a successful component of many gain-of-function studies in pooled screening formats. That said, it’s difficult to sort many biologically relevant phenotypes quickly, so arrayed screens are often a better approach.

Learn more about CRISPR gene editing from BioTechniques.

About BioTechniques

Established in 1983, the peer-reviewed journal BioTechniques publishes developments in lab methodologies, technologies, and tools. Over the past four decades, BioTechniques has provided the knowledge hub for life scientists and other research professionals all over the world to keep up with the reproducibility and efficacy of these techniques and systems, each of which is essential to scientific progression. Users can also find key information on BioTechniques’ multimedia website, which offers informative videos, webinars, infographics, podcasts, and eBooks.

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