Coexpression of normal and mutated CD40 ligand with deletion of a putativeRNA lariat branchpoint sequence in X-linked hyper-IgM syndrome

We describe a novel CD40 ligand (CD40L) splicing mutation in a patient with X-linked hyper-IgM syndrome (X-HIM) associated with alternate splicing of exon 3, resulting in the expression of both full-length and exon-3-skipped CD40L mRNA. The mutation is an 8-bp deletion 25 bp upstream of the intron 2 /exon 3 junction which overlaps a putative RNA branchpoint, suggesting that it may impair RNA lariat formation. The exon-3-skipped CD40L transcript en codes a truncated protein (CD40L Delta E3) encompassing the cytoplasmic, tr ansmembrane, and extracellular stalk domains, but lacking the CD40L recepto r binding domain. CD40L Delta E3 protein expression was readily detectable in transfected Cos cells by immunofluorescence. In cells cotransfected with CD40L Delta E3 and wild-type CD40L, expression of CD40L Delta E3 did not i nhibit the expression of wild-type CD40L monomers, but strongly inhibited s taining by the conformationally sensitive anti-CD40L mAb 5c8, suggesting th at CD40L Delta E3 acts in a dominant negative manner to inhibit the assembl y of functional CD40L trimers. This mechanism may contribute to the pathoph ysiology of CD40L deficiency in X-HIM patients with leaky splice site mutat ions. (C) 2001 Academic Press.