Acetylation of the adenovirus-transforming protein E1A determines nuclear localization by disrupting association with importin-alpha.
Posttranslational modifications may alter the biochemical functions of a protein by modifying associations with other macromolecules, allosterically altering intrinsic catalytic activities, or determining subcellular localization. The adenovirus-transforming protein E1A is acetylated by its cellular targets, the co-activators CREB-binding protein, p300, and p300/CREB-binding protein-associated factor in vitro and also in vivo at a single lysine residue (Lys(239)) within a multifunctional carboxyl-terminal domain necessary for both nuclear localization and interaction with the transcriptional co-repressor carboxyl-terminal binding protein (CtBP). In contrast to a previous report, we demonstrate that acetylation of Lys(239) does not disrupt CtBP binding and that 12 S E1A-mediated repression of CREB-binding protein-dependent transcription does not require recruitment of CtBP. Instead we find that the cytoplasmic fraction of E1-transformed 293 cells is enriched for acetylated E1A with relative exclusion from the nuclear compartment. Whereas wild type 12 S E1A binds importin-alpha 3, binding affinity was markedly reduced both by single amino acid substitution mutations and acetylation at Lys(239). This is the first demonstration that acetylation may alter nuclear partitioning by direct interference with nuclear import receptor recognition. The finding that the cytoplasmic fraction of E1A is acetylated indicates that E1A may exert its pleiotropic effects on cellular transformation in part by affecting cytoplasmic processes.
Madison
Dana L
DL
Division of Molecular Medicine, Department of Medicine, Oregon Health and Science University, Portland, Oregon 97201, USA.
Yaciuk
Peter
P
Kwok
Roland P S
RP
Lundblad
James R
JR
D000107:Acetylation; D021581:Active Transport, Cell Nucleus; D017500:Adenovirus E1A Proteins / Q000235:genetics / Q000378:metabolism*; D000429:Alcohol Oxidoreductases; D000818:Animals; D050882:CREB-Binding Protein; D002458:Cell Fractionation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D050881:E1A-Associated p300 Protein; D017930:Genes, Reporter; D006801:Humans; D008239:Lysine / Q000378:metabolism; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D028901:alpha Karyopherins / Q000378:metabolism*
D017500:Adenovirus E1A Proteins; D004268:DNA-Binding Proteins; C106259:KPNA4 protein, human; D009687:Nuclear Proteins; D010750:Phosphoproteins; D020033:Protein Isoforms; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; D028901:alpha Karyopherins; D000429:Alcohol Oxidoreductases; C096794:C-terminal binding protein; D050882:CREB-Binding Protein; C494363:CREBBP protein, human; C494364:Crebbp protein, mouse; D050881:E1A-Associated p300 Protein; C494361:Ep300 protein, mouse; D008239:Lysine