The regulation of gene expression is being studied in three systems involving the immune system. We previously showed that all laboratory rabbits in our study harbored two sequences hybridizing to kappa immunoglobulin gene probes in Southern blots: the gene for the """"""""nominal"""""""" expressed kappa1 chain and an unexpressed gene encoding a kappa2 isotype light chain. We have established that the J region cluster of the kappa1b5 gene resembles the kappa1b4 cluster in containing only a single apparently functional J gene segment within a cluster of five J-like sequences. Experiments in progress are aimed at analyzing the potential relationship between transcriptional regulatory sequences in the J-C intron and the relative in vivo expression of different rabbit kappa genes. In a second project we are attempting to examine the chromosomal state of a known regulatory region (enhancer) of the human kappa immunoglobulin gene using genomic gene blotting technology, including genomic sequencing methods which have achieved the degree of sensitivity to allow comparison of the effects of different chromosomal proteins (e.g., in B versus T cells) on the accessibility of the kappa enhancer region to DNA-modifying reagents (footprint analysis). This region is, in a B cell, unusually sensitive to certain restriction endonucleases. In a third project we have cloned the complete J chain gene and determined the nucleotide sequence of all the exons and some flanking regions. Using probes from the cloned gene we have examined J chain gene expression and gene methylation in several pre-B and B cell lines. Currently we are attempting to assess the potential regulatory function of a sequence 5' of the gene using a chloramphenicol acetyl transferase (CAT) transient expression system.
