Alternative splicing and differential subcellular localization of the rat FGF antisense gene product

RT-PCR and immunohistochemistry revealed tissue-specific GFG mRNA isoform expression and subcellular distribution of GFG immunoreactivity in cytoplasm and nuclei of a wide range of normal rat tissues. FGF-2 and GFG immunoreactivity were co-localized in some, but not all, tissues examined. Computational analysis identified a mitochondrial targeting sequence (MTS) in the N-terminus of three previously described rGFG isoforms. Confocal laser scanning microscopy and subcellular fractionation analysis revealed that all rGFG isoforms bearing the MTS were specifically targeted to mitochondria whereas isoforms and deletion mutants lacking the MTS were localized in the cytoplasm and nucleus. Mutation and deletion analysis confirmed that the predicted MTS was necessary and sufficient for mitochondrial compartmentalization.Previous findings strongly support a role for the FGF antisense RNA as a regulator of FGF2 expression. The present study demonstrates that the antisense RNA itself is translated, and that protein isoforms resulting form alternative RNA splicing are sorted to different subcellular compartments. FGF-2 and its antisense protein are co-expressed in many tissues and in some cases in the same cells. The strong conservation of sequence and genomic organization across animal species suggests important functional significance to the physical association of these transcript pairs.The FGF-2 gene is post-transcriptionally regulated by an endogenous complementary (antisense) mRNA transcribed from the GFG/NUDT6 gene on the opposite DNA strand (Fig. 1). The FGF antisense (FGF-AS) transcript was first identified in Xenopus laevis [1] and has since been identified in a variety of other vertebrate species including chicken [2], rat [3] and human [4,5]. The highly conserved organization and sequence of the FGF-2 and GFG genes across many vertebrate species suggest that this structural relationship has an important function. The sense and antisense RNAs form stable dsRNA comple