Introduction and Objectives Cell adhesion molecule 1 (CADM1) is implicated in several diseases and a prognostic factor for lung cancer. In human lung mast cells (HLMC) it contributes to cell-cell adhesion and proliferation. Functional CADM1 isoforms arise from alternative splicing between exons 7 and 11. Exon 9 may be functionally important because it encodes a specific cleavage site for TNF-alpha-cutting enzyme, TACE, in the longest SP1 and SP6 isoforms. Our aim was to characterise the isoforms of CADM1 expressed in HLMC.
Methods CADM1 expression in isolated HLMC and human mast cell lines (HMC-1 and LAD2), was investigated using RT-PCR, cloning, and transfection.
Results Multiple highly glycosylated CADM1 isoforms were found in HLMCs and cell lines. The SP4 isoform containing exon 8 represented ∼80% of clones in both HLMCs and ‘differentiated’ LAD2 cells, and ∼96% in non-differentiated HMC-1 cells. The SP1 isoform with exons 8+9 represented ≤20% of clones in HLMCs and LAD2 cells. A novel SP6 isoform with exons 8+9+10 was found only in HLMCs (<5% of clones). More sensitive PCR analysis detected all these isoforms and an additional isoform SP3, lacking exons 8–10, in HLMCs and in cell lines. In contrast to these functional isoforms, non-functional isoforms were also found with two cryptic exons between exons 1 and 2, which cause a translational frame shift and premature termination of the protein. The cryptic exon B is located within the hot spot for SNPs. When the isoforms SP4, SP1 and SP6 were expressed in the HMC-1 cell line and the epithelial cell line HEK293 as fusion CADM1-GFP proteins, they were found at the cell periphery as well as in the cytoplasm. They were concentrated in cell boundaries in clumps of HMC1 cells.
Conclusions Multiple and novel CADM1 isoforms are found in HLMCs. The longest SP1 and SP6 isoforms might be involved in the negative regulation of HLMC adhesion following secretion of TACE after activation. Some cryptic exons are likely to be a result of specific SNPs, creating new branching points for splicing. Non-functional cryptic CADM1 isoforms may reduce mature protein expression and affect the function of CADM1 expressing cells.