Alternative splicing of the RBP1 gene clusters in an internal exon that encodes potential phosphorylation sites.
We have isolated cDNA and genomic clones for the human retinoblastoma binding protein 1 (RBP1) gene, and have identified alternative splicing of RBP1 clustered within a 207-nucleotide internal exon. Three of the predicted RPB1 peptides share amino-terminal and carboxy-terminal domains, while a fourth species encodes a distinct carboxy-terminal domain. Functional analysis of these peptides demonstrated that they are capable of precipitating retinoblastoma (RB) protein in vitro from K562 cell lysates, but cannot bind to mutant RB protein. However, each of the RBP1 peptides differed within an internal exon that contains potential casein kinase II and p34cdc2 phosphorylation sites. Immunoblot analysis using polyclonal alpha-RBP1 antiserum revealed that the RBP1 protein is expressed in a wide range of cell lines of differing histologic type and migrates on sodium dodecyl sulfate-polyacrylamide gel electrophoresis predominantly as a 200-kDa protein. Immunohistochemical analysis using the alpha-RBP1 antiserum demonstrated a distinct nuclear staining pattern that was eliminated when the antiserum was preabsorbed with RBP1 peptide. The RBP1 gene encodes a widely expressed 200-kDa nuclear protein and undergoes alternative splicing that predicts a family of RB-binding peptides.
Otterson
G A
GA
NCI-Navy Oncology Branch, Bethesda, Maryland 20889.
Kratzke
R A
RA
Lin
A Y
AY
Johnston
P G
PG
Kaye
F J
FJ
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D005091:Exons; D015870:Gene Expression; D005796:Genes; D006801:Humans; D008969:Molecular Sequence Data; D005810:Multigene Family; D009687:Nuclear Proteins / Q000235:genetics*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010750:Phosphoproteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism*
D002352:Carrier Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D010750:Phosphoproteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein