The Epstein-Barr virus latent membrane protein 2A PY motif recruits WW domain-containing ubiquitin-protein ligases.
Latent membrane protein 2A (LMP2A) is expressed in latent Epstein-Barr virus (EBV) infection. LMP2A functions to downregulate B-cell signal transduction and viral reactivation from latency in EBV-immortalized B cells in vitro, and acts to provide B cells with both a survival and developmental signal in vivo. Identification of proteins associated with LMP2A is important for elucidation of the mechanism that LMP2A employs to regulate B-cell signal transduction and EBV latency. LMP2A is constitutively tyrosine phosphorylated and is associated with protein tyrosine kinases such as Lyn and Syk when specific LMP2A tyrosines are phosphorylated. The amino-terminal domain of LMP2A includes multiple proline-rich regions, which may provide binding sites for proteins containing SH3 or WW domains. In this study, we demonstrate that four cellular proteins bind specifically to two PPPPY (PY) motifs present within the LMP2A amino-terminal domain. Protein microsequence analysis determined that three of these proteins were AIP4, WWP2/AIP2, and Nedd4. All of these proteins are members of the Nedd4-like ubiquitin-protein ligases family and have conserved domains including the C2, WW, and ubiquitin-protein ligase domain. The mutation of both PY motifs completely abolished binding activity of these proteins to LMP2A and the interaction of AIP4 and WWP2 with LMP2A was confirmed in cell lines expressing LMP2A, WWP2, and AIP4. Furthermore, a reduction in the level of Lyn and the rapid turnover of LMP2A and Lyn were observed in LMP2A-expressing cells. These findings suggest that LMP2A recruits Nedd4-like ubiquitin-protein ligases and B-cell signal transduction molecules, resulting in the degradation of LMP2A and Lyn by a ubiquitin-dependent mechanism. This provides a new means by which LMP2A may modulate B-cell signal transduction.
Ikeda
M
M
Department of Microbiology-Immunology, Northwestern University Medical School, Chicago, Illinois, 60611, USA.
Ikeda
A
A
Longan
L C
LC
Longnecker
R
R
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D018076:DNA, Complementary; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000378:metabolism*; D008025:Ligases / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014452:Ubiquitins / Q000378:metabolism*; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism
D002135:Calcium-Binding Proteins; D018076:DNA, Complementary; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D014763:Viral Matrix Proteins; D005982:Glutathione Transferase; D019061:src-Family Kinases; D008025:Ligases