COVID-19 Immunology: Study Shows That SARS-CoV-2 Viral Protein ORF3a Activates NLRP3 Inflammasome Causing Severe Inflammatory Responses

COVID-19 Immunology: Scientist from the University of Florida have found that the SARS-CoV-2 viroporin triggers the NLR Family Pyrin Domain Containing 3 (NLRP3) inflammasome, which is responsible for the inflammatory pathology in severely ill COVID-19 patients. Viroporins are small and usually hydrophobic multifunctional viral proteins that modify cellular membranes, thereby facilitating virus release from infected cells.


 
Cytokine storm resulting from a heightened inflammatory response is a prominent feature of severe COVID-19 disease. This inflammatory response results from assembly/activation of a cell-intrinsic defense platform known as the inflammasome. The study team report s that the SARS-CoV-2 viroporin encoded by ORF3a activates the NLRP3 inflammasome, the most promiscuous of known inflammasomes. ORF3a triggers IL-1β expression via NFκB, thus priming the inflammasome while also activating it via ASC-dependent and -independent modes.
 
The research findings were published on a preprint server and is pending peer review https://www.biorxiv.org/content/10.1101/2020.10.27.357731v1
 
Typically a cytokine storm syndrome occurs when several medical conditions emerge, and the immune system produces too many inflammatory signals. This syndrome can lead to organ failure and death.
 
Cytokines are small glycoproteins produced by many types of cells in the body and are important in cell signaling.. When these molecules are released, they can trigger body functions, including the control of cell proliferation, endocrine activity, and regulating immune and inflammatory responses.
 
Normally when there is an infection, various inflammatory cytokines are produced at a much higher rate. These molecules also recruit other immune cells to the site of injury, potentially causing organ damage.
 
Of late, the cytokine storm has received more attention due to the COVID-19 pandemic. Though health experts and clinicians are learning more each day, cytokine storm seems to be a part of the reason why people develop life-threatening symptoms from COVID-19.

The researchers said that the heightened inflammatory response results from the assembly and activation of a cell-intrinsic defense platform known as the inflammasome.
 
The study team assessed the influence of the SARS-CoV-2 ORF3a, which is thought to be a protein with ion channel activity (viroporin) that activates the NLRP3 inflammasome. ORF3a also plays a role in virus replication and the development of the disease.
 
Basically, viroporins are a group of proteins that participate in many viral functions, such as promoting the release of viral particles from cells. These proteins also affect cellular functions, including membrane permeability, glycoprotein trafficking, and the cell vesicle system. Though not essential for virus replication, some of these small proteins can generate pores that facilitate ion transport across cell membranes. As a result, they ensure virus release that can activate inflammasomes.
 
These inflammasomes gather and respond to invading organisms as a component of the innate immune system. Hence, they form the first line of defense against invading path ogens and infection.
 
As a result of the inflammatory response being a contributor to severe illness in COVID-19 patients, it is important to develop therapies that can help block infection and halt virus replication. The team investigated the influence of ORF3a on the inflammatory machinery. They also defined key amino acid residues on ORF3a required to activate the inflammatory response.
 
The study team found that the SARS-CoV-2 ORF3a protein primes and activates the inflammasome through the efflux of potassium ions and the kinase NEK7.
 
The researchers also found that though the coronavirus ORF3a protein has separated from its homologs in other coronaviruses, some of the newly divergent residues are crucial for the activation of the NLRP3 inflammasome.
 
These specific viporins are conserved in virus isolates across continents.
 
The  study team also noted that an essential viroporin needed for the release of SARS-CoV-2 from infected cells could also activate the NLRP3 inflammasome.
 
Corresponding author Dr Sumita Bhaduri-McIntosh from the Department of Molecular Genetics and Microbiology, University of Florida told Thailand Medical News, “ORF3a’s indispensability to the virus’s life cycle makes it an important therapeutic candidate. Moreover, while different from its homologs in other coronaviruses, the high conservation of the newly divergent SARS-CoV-2 ORF3a across isolates from several continents combined with our observation that multiple single point mutations reduce its ability to activate the inflammasome, argues against rapid emergence of resistance phenotypes.”
 
The study team also explained that targeting ORF3a may help block virus spread and inflammation. Hence, the study can help in formulating new therapies that can reduce the severe and fatal outcomes of COVID-19 in people who are high risk. These include those who have underlying medical conditions.
 
Dr Sumita Bhaduri-McIntosh added, “These findings reveal ORF3a and NLRP3 to be attractive targets for therapy.”
 
Besides ORF3a, other viroporins such as ORF-E and ORF8 may also contribute to the inflammatory response by similar or related mechanisms. Activation of the NLRP3 inflammasome also bears mention in broader contexts. In particular, two recent reports have found that a fraction of severely ill COVID-19 patients display defective type I interferon immunity. https://pubmed.ncbi.nlm.nih.gov/32972996/
https://pubmed.ncbi.nlm.nih.gov/32972995/
 
It is likely that severe disease in these individuals also stemmed from unchecked pro-inflammatory responses since type I interferon can counteract the NLRP3 inflammasome in a number of ways . Similarly, for those who have metabolic disturbances such as hypokalemia that often results from antihypertensive medications, ORF3a may have a lower threshold for activating the inflammasome due to a higher K+  gradient across the infected cell. https://www.jacionline.org/article/S0091-6749(16)30359-1/pdf
 
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