Phosphorylation of the nucleocapsid protein of Hantaan virus by casein kinase II

Abstract

Hantaanvirus (HTNV) is the prototype of the genus Hantavirus, which belongs to the family Bunyaviridae. Hantaviruses are carried and transmitted by rodents and are known to cause two serious disease syndromes in humans i.e., hemorrhagic fever with renal syndrome (HFRS) and the hantavirus pulmonary syndrome (HPS). HTNV is an enveloped virus that contains a tripartite genome consisting of three negative-sense RNA segments (L, M, S), and the S and M segment of HTNV, respectively, encode the viral nucleocapsid protein (NP) and envelope glycoproteins. Possible phosphorylation motifs of casein kinase II (CKII) and protein kinase C (PKC) were identified in HTNV NP through bioinformatics searches. Sucrose gradient SDS-PAGE analysis indicated that dephosphorylated HTNV NP migrated faster than non-dephosphorylated NP, suggesting that HTNV NP is phosphorylated in infected Vero E6 cells. Immunoblot anaylsis of HTNV particles with anti-phosphoserine antibody and anti-phosphothreonine antibody after immunoprecipitation showed that viral particles are readily phosphorylated at threonine residues. In vitro kinase assay further showed that HTNV NP is phosphorylated by CK II, but not by PKC. Full length or truncated HTNV NPs expressed in E. coli were phosphorylated in vitro by CKII suggesting that phosphorylation may occur in vivo at multiple sites. Site specific mutagenesis studies suggest that HTNV NP phosphorylation might occur at unknown sites excluding the site-directly mutagenized locations. Taken together, HTNV NP can be phosphorylated mainly at threonine residues in vivo by CK II treatment.

Unique Interferon Pathway Regulation by the Andes Virus Nucleocapsid Protein Is Conferred by Phosphorylation of Serine 386

“…The role of phosphorylation in Bunyaviridae family viruses is poorly understood. One study suggests that the N protein from Hantaan virus (HTNV) is serine/threonine phosphorylated; however, neither the phosphorylation functions nor the residues involved were identified (72). Although not a hantavirus, the NSs protein of the Phlebovirus Rift Valley Fever virus is suggested to be serine/threonine phosphorylated by casein kinase II (CKII) (73,74), which has a consensus target sequence of (S/T)XX(D/E) (74).…”

Protein kinase CK2: a potential therapeutic target for diverse human diseases

“…Gradually, other substrates (as HIV-1 Rev protein, 187 hepatitis C and B virus proteins 188 , 189 , and many others 5 ) were added to the list of viral CK2 targets, which in 2003 already included ~40 proteins. 2 New members are continuously added; many recently identified viral targets of CK2 are relevant for human diseases, as NSP1 of rotavirus (that causes severe diarrhea in young children), 190 the nucleocapsid protein of the Hantaan virus (that causes serious disease syndromes in humans), 191 the matrix M protein of human respiratory syncytial virus (that causes bronchiolitis and pneumonia in infants and the elderly), 192 the leader L protein of the encephalomyocarditis virus, 193 the Kaposi’s sarcoma-associated herpesvirus ORF57. 194 Interestingly, this latter, when phosphorylated by CK2 in the proximity of a caspase-7 cleavage site, is no more cleaved, indicating that this classical antiapoptotic mechanism exerted by CK2 (see Fig.…”

Tensin Phosphatase-Like System of Hantavirus Facilitates Membrane Fusion to Disrupt Vascular permeability

“…HTNV is a virus with an genome composed of three negative-sense RNA segments (L, M, S) (8). The segments L, S, and M of HTNV, respectively, encode RNA polymerase L, viral nucleocapsid protein N, and membrane glycoprotein M (8). CK II can phosphorylate HTNV N at threonine residues (8).…”

“…The segments L, S, and M of HTNV, respectively, encode RNA polymerase L, viral nucleocapsid protein N, and membrane glycoprotein M (8). CK II can phosphorylate HTNV N at threonine residues (8). N protects RNA from cellular nucleases and collaborates with L to guarantee that the viral genome replicates effectively (9).…”