Impact of the SARS-CoV-2 nucleocapsid 203K/204R mutations on the inflammatory immune response in COVID-19 severity

Muhammad Shuaib*, Sabir Adroub, Tobias Mourier, Sara Mfarrej, Huoming Zhang, Luke Esau, Afrah Alsomali, Fadwa S. Alofi, Adeel Nazir Ahmad, Abbas Shamsan, Asim Khogeer, Anwar M. Hashem, Naif A.M. Almontashiri, Sharif Hala, Arnab Pain*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

2 Scopus citations


Background: The excessive inflammatory responses provoked by SARS-CoV-2 infection are critical factors affecting the severity and mortality of COVID-19. Previous work found that two adjacent co-occurring mutations R203K and G204R (KR) on the nucleocapsid (N) protein correlate with increased disease severity in COVID-19 patients. However, links with the host immune response remain unclear. Methods: Here, we grouped nasopharyngeal swab samples of COVID-19 patients into two cohorts based on the presence and absence of SARS-CoV-2 nucleocapsid KR mutations. We performed nasopharyngeal transcriptome analysis of age, gender, and ethnicity-matched COVID-19 patients infected with either SARS-CoV-2 with KR mutations in the N protein (KR patients n = 39) or with the wild-type N protein (RG patients n = 39) and compared to healthy controls (n = 34). The impact of KR mutation on immune response was further characterized experimentally by transcriptomic and proteomic profiling of virus-like-particle (VLP) incubated cells. Results: We observed markedly elevated expression of proinflammatory cytokines, chemokines, and interferon-stimulated (ISGs) genes in the KR patients compared to RG patients. Using nasopharyngeal transcriptome data, we found significantly higher levels of neutrophils and neutrophil-to-lymphocyte (NLR) ratio in KR patients than in the RG patients. Furthermore, transcriptomic and proteomic profiling of VLP incubated cells confirmed a similar hyper-inflammatory response mediated by the KR variant. Conclusions: Our data demonstrate an unforeseen connection between nucleocapsid KR mutations and augmented inflammatory immune response in severe COVID-19 patients. These findings provide insights into how mutations in SARS-CoV-2 modulate host immune output and pathogenesis and may contribute to more efficient therapeutics and vaccine development.

Original languageEnglish (US)
Article number54
JournalGenome medicine
Issue number1
StatePublished - Dec 2023


  • COVID-19
  • Cytokine storm
  • Host immune response
  • Interferon-stimulated genes (ISGs)
  • Nucleocapsid (N) R203K/G204R (KR) mutations
  • Proteomics
  • SARS-CoV-2
  • Transcriptomics
  • Variant of concern
  • Virus-like particle (VLP)

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Genetics(clinical)


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