Adenovirus ADP protein (E3-11.6K), which is required for efficient cell lysis and virus release, interacts with human MAD2B.
The human subgroup C adenovirus (Ad) protein named adenovirus death protein (ADP) (previously named E3-11.6K) is synthesized at very late stages of infection when it mediates efficient lysis of cells and release of adenovirus to infect other cells. ADP is an integral membrane N-linked, O-linked palmitoylated glycoprotein of 101 amino acids (aa) that localizes to the nuclear membrane, endoplasmic reticulum (ER), and Golgi. It has a single membrane spanning region (roughly aa 40-60) and is oriented with aa 1-40 in the lumen and aa 61-101 in the nucleoplasm and cytoplasm. Using aa 61-101 of Ad2 ADP as bait in a yeast two-hybrid screen, we isolated a cDNA for a 211-aa protein that initially was not in the database but has now been published by others with the names human MAD2B, MAD2L2, and REV7. ADP binds strongly to human MAD2B not only in yeast but also in GST pull-down experiments and in coimmunoprecipitations of ADP and MAD2B synthesized in vitro or in vivo. ADP mutants with deletions throughout the bait region do not interact with human MAD2B, whereas a Pro69Pro70 to Ala69Ala70 mutant in the "basic-proline" domain of ADP does interact. Northern blot analyses indicate that human MAD2B is expressed ubiquitously. Human MAD2B is about 25% identical to human MAD2, a spindle assembly checkpoint protein. Two human A549 cell lines were made that constitutively overexpress MAD2B. Wild-type adenovirus lyses these cells significantly more slowly than it lyses parental A549 cells, raising the possibility that ADP and MAD2B act in opposition and suggesting that the ADP-MAD2B interaction is biologically relevant.
Ying
Baoling
B
Department of Molecular Microbiology and Immunology, St. Louis University School of Medicine, 1402 South Grand Boulevard, St. Louis, MO 63104, USA.
Wold
William S M
WS
D017503:Adenovirus E3 Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D006801:Humans; D064246:Mad2 Proteins; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding; D011506:Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014779:Virus Replication
D017503:Adenovirus E3 Proteins; C120552:MAD2L2 protein, human; D064246:Mad2 Proteins; D008562:Membrane Glycoproteins; D011506:Proteins; C099609:adenovirus death protein, Adenovirus