Functional significance of globotriaosyl ceramide in interferon-alpha(2)/type 1 interferon receptor-mediated antiviral activity.
The N-terminus of the type 1 interferon receptor subunit, IFNAR1, has high amino acid sequence similarity to the receptor binding B subunit of the Escherichia coli-derived verotoxin 1, VT1. The glycolipid, globotriaosyl ceramide (Gb(3): Gal alpha(1) --> 4 Gal beta 1 --> 4 Glu beta 1 --> 1 Cer) is the specific cell receptor for VT1. Gb(3)-deficient variant cells selected for VT resistance are cross-resistant to interferon-alpha (IFN-alpha)-mediated antiproliferative activity. The association of eIFNAR1 with Gal alpha 1 --> 4 Gal containing glycolipids has been previously shown to be important for the receptor-mediated IFN-alpha signal transduction for growth inhibition. The crucial role of Gb(3) for the signal transduction of IFN-alpha-mediated antiviral activity is now reported. IFN-alpha-mediated antiviral activity, nuclear translocation of activated Stat1, and increased expression of PKR were defective in Gb(3)-deficient vero mutant cells, although the surface expression of IFNAR1 was unaltered. The VT1B subunit was found to inhibit IFN-alpha-mediated antiviral activity, Stat1 nuclear translocation and PKR upregulation. Unlike VT1 cytotoxicity, IFN-alpha-induced Stat1 nuclear translocation was not inhibited when RME was prevented, suggesting that the accessory function of Gb(3) occurs at the plasma membrane. IFN-alpha antiviral activity was also studied in Gb(3)-positive MRC-5 cells, which are resistant to IFN-alpha growth inhibition, partially resistant to VT1 but still remain fully sensitive to IFN-alpha antiviral activity, and two astrocytoma cell lines expressing different Gb(3) fatty acid isoforms. In both systems, long chain fatty acid-containing Gb(3) isoforms, which are less effective to mediate VT1 cytotoxicity, were found to correlate with higher IFN-alpha-mediated antiviral activity. Inhibition of Gb(3) synthesis in toto prevented IFN-alpha antiviral activity in all cells. We propose that the long chain Gb(3) fatty isoforms preferentially remain in the plasma membrane, and by associating with IFNAR1, mediate IFN-alpha antiviral signaling, whereas short chain Gb(3) fatty acid isoforms are preferentially internalized to mediate VT1 cytotoxicity and IFNAR1-dependent IFN-alpha growth inhibition.
Khine
A A
AA
Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada.
Lingwood
C A
CA
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000037:antagonists & inhibitors / Q000633:toxicity; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D029822:Enterovirus B, Human / Q000187:drug effects / Q000502:physiology; D005227:Fatty Acids / Q000032:analysis / Q000378:metabolism; D050832:Interferon-Stimulated Gene Factor 3; D016898:Interferon-alpha / Q000494:pharmacology*; D008565:Membrane Proteins; D008970:Molecular Weight; D009154:Mutation; D053633:Receptor, Interferon alpha-beta; D017471:Receptors, Interferon / Q000378:metabolism*; D050794:STAT1 Transcription Factor; D022622:Shiga Toxin 1; D015398:Signal Transduction / Q000187:drug effects; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014281:Trihexosylceramides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014709:Vero Cells; D014721:Vesicular stomatitis Indiana virus / Q000187:drug effects / Q000502:physiology; D019892:eIF-2 Kinase / Q000737:chemistry / Q000378:metabolism
D000998:Antiviral Agents; D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D005227:Fatty Acids; C507964:IFNAR1 protein, human; D050832:Interferon-Stimulated Gene Factor 3; D016898:Interferon-alpha; D008565:Membrane Proteins; D017471:Receptors, Interferon; D050794:STAT1 Transcription Factor; D022622:Shiga Toxin 1; D015534:Trans-Activators; D014157:Transcription Factors; D014281:Trihexosylceramides; D053633:Receptor, Interferon alpha-beta; C018549:globotriaosylceramide; D019892:eIF-2 Kinase