Immunological and biochemical characterisation of 7ap, a short protein translated from an alternative frame of ORF7 of PRRSV

Sequence analysis revealed a short alternative open reading frame (ORF) named ORF7a within the nucleocapsid gene of genetically divergent porcine reproductive and respiratory syndrome virus (PRRSV) genomes. Alignment of the corresponding protein sequences (named 7ap) revealed substantial heterogeneity among 7aps of different genotypes, though all of them are predicted to be positively charged. Green fluorescent protein and FLAG fusion constructs of ORF7a of the HU-14432/2011 PRRSV demonstrated that 7ap is expressed. 7ap of HU- 14432/2011 (Hu7ap) was synthesised chemically, and ELISA experiments revealed that Hu7ap binds strongly to mammalian IgGs. Protein-protein gel retardation assays and complement fixation inhibition suggest that 7aps bind to the CH2 domain of the IgG(Fc) fragment. Cellular localisation and immunological characteristics of PRRSV 7ap may indicate multiple functions including nuclear and cytoplasmic over-tuning of normal cellular processes and immunosuppression.