In response to antigenic or mitogenic stimulation, lymphocytes secrete an array of meditors, known as lymphokines which exert profound effects on cells of the immune system. Macrophages play important roles in the initiation of immune responses and in host defense and these functions are under the control of a variety of lymphokines. Migration inhibitory factor (MIF), a lymphokine that inhibits the migration of macrophages is one of the lymphokines that correlates especially well with cellular immune responses. Until now, the lack of pure MIF precluded the determination of its physical and biological properties. We have recently isolated and sequenced a cDNA encoding a human MIF, MIF-1 (additional independent clones have been isolated but not yet examined in detail). It will allow the characterization of its gene and protein structure as well as facilitate the elucidation of its biological activities. The proposed work aims to investigate the structure of MIF-1 gene using the cloned MIF-cDNA as a probe to screen a human genomic library. Positive clones will be selected for analysis of intron/exon junctions and 5' and 3' regions of the gene and for mapping of the exons. The proposed work plans to purify recombinant MIF-1 using gel filtration, Phenyl-Sepharose chromatography, isoelectric focusing, and reverse phase high pressure liquid chromatography and to produce monoclonal antibodies to recombinant MIF-1 using established methods. This study further aims to investigate the effects of recombinant MIF-1 on cultured macrophages and the interactions of MIF-1 with other lymphokines that affect macrophages. The role of MIF in host defense and in cellular immunity will be delineated by monitoring antileishmanial activity and tumoricidal activity of macrophages treated with MIF alone or in combination with other lymphokines., This study also includes the analysis of the additional cDNA clones encoding different MIF species in order to determine their DNA sequences, to characterize the corresponding MIF proteins, to examine their bioactivities, and to correlate the production of specific MIF species in response to different antigenic stimulation at the level of mRNA.
