Functional characterization of allelic variants of pulmonary adenoma susceptibility 1 (Pas1) genes for their cancer modifier activity

Galbiati, Federica (2007). Functional characterization of allelic variants of pulmonary adenoma susceptibility 1 (Pas1) genes for their cancer modifier activity. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000fa74

Abstract

The pulmonary adenoma susceptibility 1 (Pas1) locus affects inherited predisposition and resistance to chemically induced lung tumorigenesis in mice. A/J and C57BL6/J mouse strains carry the susceptibility and resistance allele, respectively. In the present study we identified and genotyped 64 polymorphisms in the Pas1 locus in 29 mouse inbred strains in order to reduce the region containing the Pas1 locus, and we succeeded in delimiting the Pas1 locus to a minimal region of 468kb containing six genes (Bcat1, Lrmp, Cascl, Ghiso, Kras2 and Lmn-rs1). That region defined a core Pas1 haplotype (A/J- and C57BL/6J-type) with 42 tightly linked markers, including intragenic polymorphisms in five genes (Bcat1, Lrmpf Casc1, Ghiso and Kras2) and amino-acid changes in three genes (Lrmp, Casc1 and Lmn-rs1). mRNA expression study of these candidate genes revealed that, in (A/JxC57BL/6J)F1 mouse lung tumors, the Lmna-rs1 gene was completely downregulated, whereas allele-specific downregulation of the C57BL/6J-derived allele was observed at the Casc1 gene, suggesting the potential role of these genes in tumor suppression. These first results indicate a complex multigenic nature of the Pas1 locus, and point to a functional role for both intronic and exonic polymorphisms of the six genes of the Pas1 haplotype in lung tumor susceptibility. Then we continued by testing functional activity of candidate genes in vitro. In vitro colony formation assay of human lung cancer cell lines A549 and NCI-H520 transfected with the allelic variants of the four genes revealed allele-specific modulations of colony numbers by Lmna-rs1 and Casc1, but not by Lrmp and Ghiso. We found that inhibition of clonogenicity by allelic forms of Pas1 candidate genes was not mediated by induction of apoptosis. These findings provide evidence that allelic variants of mouse Pas1 candidate genes differentially modulate growth of human cancer cells.