pp120/ecto-ATPase, an endogenous substrate of the insulin receptor tyrosine kinase, is expressed as two variably spliced isoforms.
The insulin receptor possesses tyrosine kinase activity which is thought to mediate the biological effects of insulin upon target cells. pp120 is a liver-specific glycoprotein of apparent molecular size of 120 kDa that is phosphorylated on tyrosine residues by the receptors for insulin, insulin-like growth factor-I, and epidermal growth factor. Previously, we demonstrated that pp120 is identical to a liver-specific ecto-ATPase. In the present study, we have cloned the rat gene encoding pp120/ecto-ATPase. The gene is contained within approximately 15 kilobases of genomic DNA, and consists of nine exons interrupted by eight introns. Using the reverse transcriptase/polymerase chain reaction, we isolated cDNA clones complementary to rat liver mRNA encoding pp120/ecto-ATPase. Sequence analysis indicated the presence of two populations of cDNA's that differ by the presence or absence of a 53-base pair (bp) fragment encoding the juxta-membrane region of the cytoplasmic domain. By cloning the corresponding region of the ecto-ATPase gene, we demonstrated that the 53-bp represents exon 7 of the gene. This 53-bp exon undergoes alternative splicing, thereby giving rise to two mRNA variants. Deletion of this 53-bp cassette exon introduces a frameshift, and results in a premature chain termination codon that truncates the cytoplasmic domain. The truncated cytoplasmic domain contains 10 rather than 71 amino acid residues. Because the short isoform of ecto-ATPase lacks the putative sites for tyrosine- and serine-specific phosphorylation, this alternative splicing may have a major effect upon the physiological function of the enzyme.
Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
S M
SM
Najjar
D
D
Accili
N
N
Philippe
J
J
Jernberg
R
R
Margolis
S I
SI
Taylor
D000251:Adenosine Triphosphatases / Q000235:genetics* / Q000378:metabolism*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D005091:Exons; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D016133:Polymerase Chain Reaction; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D011972:Receptor, Insulin / Q000378:metabolism*; D017384:Sequence Deletion; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity
D007527:Isoenzymes; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D000251:Adenosine Triphosphatases; C027201:ectoATPase