MOLECULAR-CLONING AND CHARACTERIZATION OF HUMAN FGF8 ALTERNATIVE MESSENGER-RNA FORMS

Three alternatively spliced mRNA isoforms of the human fibroblast grow th factor-8 (FGF8) gene, expressed in a prostatic carcinoma cell line, have been isolated as cDNA clones and characterized by DNA sequencing . The clones, designated FGF8a, FGF8b, and FGF8e, differ from each oth er at the NH2-terminal region of the mature proteins and share extensi ve nucleotide sequence homology in the protein coding region to the co rresponding mouse cDNA isoforms that were previously reported. FGF8a a nd FGF8b exhibit identical amino acid sequences to those of their muri ne counterparts, FGF8e displays partial sequence variation from the co rresponding mouse clone only in the extra exon sequence found in this isoform in both species, There is extensive sequence diversity between FGF8 (human) and Fgf8 (murine) genes in the 3'-untranslated region of the mRNAs. Northern blot analyses revealed FGF8 mRNA expression only in fetal kidney tissue among the various fetal and adult human tissues tested. The reverse transcription-PCR amplification method, however, detected FGF8 mRNA expression in adult prostate, kidney, and testes (t he tissues that were tested) and in all normal and tumor prostatic epi thelial cell lines examined; although expression of both FGF8a and FGF 8b was seen in kidney and testes, FGF8b appeared to be the predominant ly expressed species in the prostatic tissue and cell lines analyzed b y reverse transcription-PCR. To address the biological effect of speci fic isoform expression, NIH3T3 cells were transfected with a eukaryoti c expression vector containing cDNA for FGF8a, FGF8b, or FGF8e. Consis tent with previous reports on differences in the transforming potentia l of mouse FGF8 isoforms, human FGF8b was found to induce marked morph ological transformation to NIH3T3 cells and strong tumorigenicity of t he transfected cells in nude mice, Human FGF8a and FGF8e were moderate ly transforming in NIH3T3 cells, and the transfected cells were modera tely tumorigenic in vivo. These results document the production of thr ee alternatively spliced FGF8 mRNAs in human tissues and the transform ing and tumorigenic potential of their protein products, Moreover, the se data, combined with the tissue-specific expression of these isoform s, suggest that they may have different biological functions.