A single base mutation in the 5 ' splice site of intron 7 of the lck gene is responsible for the deletion of exon 7 in lck mRNA of the JCaM1 cell line

The failure of signal transduction in the JCaM1 cell line was associated wi th the presence of an abnormal lck mRNA deleted of the exon 7 encoding for an inactive p56(lck) kinase. Our study of the Cck mRNA from various T cell lines and from peripheral blood lymphocytes of healthy donors has revealed the presence of both complete and exon 7-deleted lck transcripts. Thus the exon 7-deleted lck transcript initially described in the JCaM1 mutant cell line, arises from an alternative splicing event occurring in each cells exp ressing the lck gene. Genomic DNA sequencing of the lck exons 6-8 portion f rom both the mutant JCaM1 and its parental Jurkat cell lines revealed as th e only difference, the presence of a A to G mutation within the 5' splice s ite of intron 7 in the JCaM1 cell line DNA. To demonstrate the role of this point mutation in the lck pre-mRNA maturation, COS cells were transfected by lck minigenes from the Jurkat and JCaM1 cell lines. In COS cells transfe cted with minigene from the Jurkat cell line both lck transcripts (with and without exon 7) were observed whereas only the exon 7-spliced lck transcri pt was observed in COS cells transfected with minigene from the JCaM1 cell line. Thus the mutation is per se responsible for the deletion of exon 7 an d the absence of complete lck mRNA in the JCaM1 cell line. Presence of a re striction site (HphI) in the 5' splice site of lck intron 7 from Jurkat DNA allowed to confirm the presence of the mutation on both alleles in the JCa M1 cell line.