Identification of peptide domains involved in the subcellular localization of the feline coronavirus 3b protein

Abstract

Feline coronavirus (FCoV) has been identified as the aetiological agent of feline infectious peritonitis (FIP), a highly fatal systemic disease in cats. FCoV open reading frame 3 (ORF3) encodes accessory proteins 3a, 3b and 3 c. The FCoV 3b accessory protein consists of 72 amino acid residues and localizes to nucleoli and mitochondria. The present work focused on peptide domains within FCoV 3b that drive its intracellular trafficking. Transfection of different cell types with FCoV 3b fused to enhanced green fluorescent protein (EGFP) or 3×FLAG confirmed localization of FCoV 3b in the mitochondria and nucleoli. Using serial truncated mutants, we showed that nucleolar accumulation is controlled by a joint nucleolar and nuclear localization signal (NoLS/NLS) in which the identified overlapping pat4 motifs (residues 53–57) play a critical role. Mutational analysis also revealed that mitochondrial translocation is mediated by N-terminal residues 10–35, in which a Tom20 recognition motif (residues 13–17) and two other overlapping hexamers (residues 24–30) associated with mitochondrial targeting were identified. In addition, a second Tom20 recognition motif was identified further downstream (residues 61–65), although the mitochondrial translocation evoked by these residues seemed less efficient as a diffuse cytoplasmic distribution was also observed. Assessing the spatiotemporal distribution of FCoV 3b did not provide convincing evidence of dynamic shuttling behaviour between the nucleoli and the mitochondria.

Proteins Nsp12 and 13 of SARS-CoV-2 Have Mitochondrial Recognition Signal: A Connection with Cellular Mitochondrial Dysfunction and Disease Manifestation

“…MPP cleavage occurs in the mitochondrial matrix and is required to cleave the N terminal mitochondrial signal sequence before a protein can enter inside. There are reports on other corona viruses where experiments have shown that the viral proteins showing mitochondrial localization possess TOM20 recognition motifs in the N terminal region [6]. Hence, these factors are important to consider while assessing the localization of any viral protein in mitochondria.…”

Genomic evolution of the human and animal coronavirus diseases

“…These infections often serve as useful models of human and animal disease. MHV represents a spectrum of coronaviruses with tropisms for a wide variety of mouse cells or tissues, such as the enteric epithelium, respiratory epithelium, vascular endothelium, lymphoid tissue, hemopoietic tissue, liver and central nervous system [ 4 , 5 , 8 ].…”