Molecular cloning, expression and characterization of the ovine IL-2R alpha chain.
Interleukin-2 (IL-2) stimulates the proliferation of activated antigen-specific T cells through its interaction with high affinity receptors. This event is largely regulated by the inducible expression of the alpha-chain (CD25) which, in combination with the beta-chain and possibly additional chains, forms the high affinity IL-2 receptor (IL-2R) complex. From a concanavalin A (Con A)-activated ovine T-cell complementary DNA (cDNA) library we have isolated two cDNA clones which together constitute a 2650 base pair (bp) messenger RNA (mRNA) species encoding the ovine IL-2R alpha chain. The nucleotide sequence has high homology with analogous cDNA from other species and predicts a mature protein of 254 amino acids. In addition to the predominate 2.6 kilobase (kb) ovine IL-2R alpha chain mRNA species. Northern blot analysis of activated T-cell RNA revealed two larger mRNA species. The ovine IL-2R alpha chain cDNA was transfected into CHO cells and low affinity binding of human recombinant IL-2 demonstrated. Polyclonal antisera generated against the transfected cells cross-reacted with Con A-activated ovine lymphocytes. In addition these antisera were used to immunoprecipitate a unique 50,000 MW protein from the transfected cells. It is likely that this protein represents the expressed ovine IL-2R alpha chain cDNA which is heavily glycosylated as distinct from the 30,869 MW primary translation product. Southern blot analysis of ovine genomic DNA suggests that the ovine IL-2R alpha chain is encoded by a single copy gene.
Verhagen
A M
AM
Centre for Animal Biotechnology, School of Veterinary Science, University of Melbourne, Parkville, Australia.
Andrews
A E
AE
Brandon
M R
MR
Nash
A D
AD
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003208:Concanavalin A / Q000276:immunology; D004247:DNA / Q000032:analysis; D005260:Female; D008969:Molecular Sequence Data; D015375:Receptors, Interleukin-2 / Q000235:genetics*; D012756:Sheep / Q000235:genetics*; D013601:T-Lymphocytes / Q000276:immunology
D015375:Receptors, Interleukin-2; D003208:Concanavalin A; D004247:DNA