In vivo and in vitro characterization of the B1 and B2 zinc-binding domains from the acute promyelocytic leukemia protooncoprotein PML.
Acute promyelocytic leukemia (APL) has been ascribed to a chromosomal translocation event which results in a fusion protein comprising the PML protein and retinoic acid receptor alpha. PML is normally a component of a nuclear multiprotein complex which is disrupted in the APL disease state. Here, two newly defined cysteine/histidine-rich protein motifs called the B-box (B1 and B2) from PML have been characterized in terms of their effect on PML nuclear body formation, their dimerization, and their biophysical properties. We have shown that both peptides bind Zn2+, which induces changes in the peptides' structures. We demonstrate that mutants in both B1 and B2 do not form PML nuclear bodies in vivo and have a phenotype that is different from that observed in the APL disease state. Interestingly, these mutations do not affect the ability of wild-type PML to dimerize with mutant proteins in vitro, suggesting that the B1 and B2 domains are involved in an additional interaction central to PML nuclear body formation. This report in conjunction with our previous work demonstrates that the PML RING-Bl/B2 motif plays a fundamental role in formation of a large multiprotein complex, a function that may be common to those unrelated proteins which contain the motif.
Borden
K L
KL
Protein Structure Laboratory, Imperial Cancer Research Fund, London, United Kingdom.
Lally
J M
JM
Martin
S R
SR
O'Reilly
N J
NJ
Solomon
E
E
Freemont
P S
PS
D000495:Allosteric Site; D000595:Amino Acid Sequence; D001665:Binding Sites; D003035:Cobalt / Q000378:metabolism; D006639:Histidine; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D000071156:Promyelocytic Leukemia Protein; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism; D014162:Transfection; D025521:Tumor Suppressor Proteins; D015032:Zinc / Q000378:metabolism*
D008024:Ligands; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion; D010446:Peptide Fragments; D000071156:Promyelocytic Leukemia Protein; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C093476:promyelocytic leukemia-retinoic acid receptor alpha fusion oncoprotein; C070576:PML protein, human; D003035:Cobalt; D006639:Histidine; D015032:Zinc