RESUMO
We have previously prepared two B7-1 transfectant clones (AC29 B7-6 and AC29 B7-7) from the AC29 murine mesothelioma (MM) cell line which displayed markedly different in vivo growth rates and susceptibility to cytotoxic T cell killing. Using suppression subtractive hybridisation (SSH), we searched for factors which may determine the biological distinction seen in these clones. We isolated 19 cDNA clones from two SSH generated libraries by screening using subtracted cDNA probes and characterised them using Northern hybridisation, sequencing, RT-PCR and real-time RT-PCR. The 19 cDNA clones comprised 16 different transcripts of which 15 were identified by homology to known genes and one was novel. Expression of a murine endogenous retroviral (mERV) transcript mERV-AC29 was found in the immunogenic AC29 B7-6 clone and parental AC29 but absent in AC29 B7-7. Real-time RT-PCR was used to confirm that galectin-1, the disintegrin/metalloproteinase MDC9 and ribonucleotide reductase M1 were overexpressed in AC29 B7-7. Our results show that SSH is a powerful method for the identification of genes expressed differentially between phenotypically different tumour cell lines or clones. Characterisation of the role of those identified here will provide useful information in understanding genes responsible for differential tumorigenicity.
