cDNA clones for human platelet GPIIb corresponding to mRNA from megakaryocytes and HEL cells

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Abstract
Platelet glycoprotein (GP) IIb is one of the two subunits of the common platelet adhesion receptor, GPIIb-IIIa. The isolation, characterization and sequencing of cDNA clones encoding for the two polypeptide chains of GPIIb are described. A number of clones were isolated from .lambda.gt11 libraries constructed with mRNA from an erythroleukemic cell line, HEL, and human megakaryocytes. Two of these clones, .lambda.IIb1, from HEL cells, and .lambda.IIb2, from megakaryocytes, cross-hybridized and were selected for detailed analysis. The identification of these as authentic GPIIb clones was based on immunological criteia and confirmed by the presence ofnucleotide sequences in each insert encoding for known protein sequences of platelet GPIIb. These clones contained inserts of 1.54 kb and 1.39 kb, respectively,with overlapping sequence of 801 bp. The nucleotide sequence of the overlapping region was identical indicating that HEL cells produce a protein closely related, if not identical, to platelet GPIIb. The determined nucleotide sequence of two inserts included a coding sequence for 648 amino acid residues, a TAG stop codon and 185 nucleotides of 3'' non-coding sequence followed by a poly(A) tail. The coding sequence contained a portion of the heavy chain, the junction between the heavy and light chains and the entire light chain including a potential transmembrane-spanning domain and a short cytoplasmic tail. When these cDNA were used to probe for GPIIb mRNA, a single mRNA species of 3.9 kb was identified in both HEL cells and human megakaryocytes. A comparison of the deduced amino acid sequence for sequence for GPIIb with those of the .alpha. subunit of the vitronectin and the fibronectin receptors revealed extensive homologies. These homologies further establish the GPIIb-IIIa from platelets, together with the vitronectin and the fibronectin receptors, are members of a supergene family of adhesion receptors with a recognition specificity for Arg-Cly-Asp amino acid sequences.