The Expression and Biologic Effects of Ovine Interleukin-4 on T and B Cell Proliferation

Abstract

Using the reverse-transcriptase polymerase chain reaction (RT-PCR), cDNA encoding ovine (Ov) interleukin4 (OvIL-4) was generated from mitogen-stimulated peripheral blood mononuclear cells (PBMC). Two identical clones generated from separate RT-PCR reactions differed from a published OvIL-4 sequence, although they had a high degree of identity with the bovine and human homologs. We show by sequence analysis that the OvIL-4 cDNA retained the four alpha-helix structure and disulfide bonds identified in human IL-4 (HuIL-4). Moreover, the cDNA encoding OvIL-4 was expressed in insect cells using the baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV) as a vector. Supernatants from insect cells infected with the recombinant virus secreted an additional protein with a relative molecular mass of 17,000. This protein was recognized by an anticervine IL-4 monoclonal antibody (mAb) in a Western blot and did not react with any proteins in supernatants from uninfected insect cells or cells infected with the wild-type AcMNPV. Supernatants from insect cells infected with the recombinant virus induced the proliferation of activated B cells in a dose-dependent manner and typically demonstrated 5 X 10⁵ dilution U/ml of activity. However, OvIL-4 had no effect on the proliferation of resting T cells isolated from efferent lymph and actually inhibited the ability of a mitogen to stimulate these resting lymphocytes. In contrast, OvIL-4 induced the proliferation of mitogen-activated lymphoblast, demonstrating the complex role(s) OvIL-4 plays in the regulation of B and T cells.