PT  - JOURNAL ARTICLE
AU  - Kristina V. Tugaeva
AU  - Dorothy E. D. P. Hawkins
AU  - Jake L. R. Smith
AU  - Oliver W. Bayfield
AU  - De-Sheng Ker
AU  - Andrey A. Sysoev
AU  - Oleg I. Klychnikov
AU  - Alfred A. Antson
AU  - Nikolai N. Sluchanko
TI  - The mechanism of SARS-CoV-2 nucleocapsid protein recognition by the human 14-3-3 proteins
AID  - 10.1101/2020.12.26.424450
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.12.26.424450
4099  - http://biorxiv.org/content/early/2020/12/27/2020.12.26.424450.short
4100  - http://biorxiv.org/content/early/2020/12/27/2020.12.26.424450.full
AB  - The coronavirus nucleocapsid protein (N) controls viral genome packaging and contains numerous phosphorylation sites located within unstructured regions. Phosphorylated SARS-CoV N was shown to bind to the host 14-3-3 protein in the cytoplasm. Proteomic data indicate that seven human 14-3-3 proteins are highly abundant in human tissues vulnerable to SARS-CoV-2 infection, collectively reaching ~1.8% of all proteins in the lungs, ~1.4% in the gastrointestinal system, ~2.3% in the nervous system. Although the association between 14-3-3 and SARS-CoV-2 N proteins can represent one of the key host-pathogen interactions, its mechanism and the specific critical phosphosites were unknown. Here, we show that phosphorylated SARS-CoV-2 N protein (pN) dimers, reconstituted via bacterial co-expression with protein kinase A, directly associate, in a phosphorylation-dependent manner, with the dimeric 14-3-3 protein hub, but not with its monomeric mutant. We demonstrate that pN is recognized by all seven human 14-3-3 isoforms with various efficiencies and determine the apparent KD to selected isoforms in a low micromolar range. Serial truncations pinpointed a critical phosphorylation site to Ser197, located within the SR-rich region of N. The tight 14-3-3/pN association suggests it could regulate nucleocytoplasmic shuttling of N, while hijacking cellular pathways by 14-3-3 sequestration. As such, the assembly may represent a valuable target for therapeutic intervention.Competing Interest StatementThe authors have declared no competing interest.