REDUCTION OF CLASS-I EXPRESSION IN A BOVINE B-LYMPHOBLASTOID CELL-LINE IS LOCUS-SPECIFIC

A bovine B-lymphoblastoid cell line, BL3.1.2, defective in MHC class I expression was isolated following gamma irradiation and immunoselecti on of a previously mutated cell, BL3.1. Flow microfluorimetry demonstr ated that BL3.1.2 cells expressed reduced amount of class I molecules on the cell surface when compared with the parent cells. 2D gel electr ophoresis showed that two less class I gene products were synthesized by BL3.1.2 cells than the parent cells. However, no detectable RFLP di fference was revealed between this mutant and the parent cell lines as analyzed by Southern blots using bovine class I cDNA and locus-specif ic oligonucleotide probes, suggesting no major deletions, insertions, or translocations within class I genes. Northern blot analysis, using class I cDNA probes, revealed a marked reduction of class I mRNA in BL 3.1.2 cells. RNA unblots, probed with loci-specific oligonucleotides, and slot blot analysis demonstrated that absence of class I gene trans cription was locus specific. This finding indicates there is separate regulation of each class I locus in bovine cells. Furthermore, recombi nant IFN-gamma upregulated class I gene expression on the BL3.1.2 cell s, as indicated by flow cytometry and slot blot analysis. These findin gs indicate that the locus-specific MHC class I gene was functionally downregulated in its expression. The silent class I gene could be acti vated by cytokines known to activate the cis-interferon-responsive ele ment for subsequent transcription and translation of the newly synthes ized class I RNA. These studies provide evidence that absence of class I gene transcription in BL3.1.2 cells resulted from regulation of a l ocus-specific gene and suggest that a deficiency in locus-specific tra ns-acting factor(s) is instrumental in silencing the class I gene.