On the biological relevance of alternative splicing in plants: dual function of an Arabidopsis membrane transporter

Eukaryotic genes typically contain introns that are removed post-transcriptionally from the precursor mRNA (pre-mRNA) through splicing. The presence of numerous exons per gene enables the splicing machinery to process the same pre-mRNA differently by selectively joining different exons, generating different transcripts from a single gene via a process named alternative splicing. In contrast to transcriptional control, alternative splicing can influence almost all aspects of protein function and has emerged as a key mechanism for generating proteome diversity and functional complexity. Its prevalence in many genomes, including those of higher plants, suggests that alternative splicing plays crucial roles in biological processes, as is emphasized by the fact that its misregulation can lead to many human diseases. However, information on the functional significance of this posttranscriptional regulation mechanism in plant systems is surprisingly scarce. We have identified an Arabidopsis thaliana gene, ZIFL1 , encoding a membrane transporter from the Major Facilitator Superfamily that plays important roles in both root auxin transport and drought stress tolerance. Selection of an alternative 3N splice site in the ZIFL1 pre-mRNA generates two splice variants that differ in only two nucleotides. While the longer transcript encodes the full-length transporter, the shorter contains a premature stop codon and codes for a truncated protein lacking the 67 C-terminal amino acids. Sequencing, promoter-reporter gene and fluorescent protein fusion experiments indicate that the full-length protein localizes specifically at the tonoplast of root cells, whereas the C-terminal truncation targets the transporter to the plasma membrane of stomatal guard cells. Using reverse genetics, we show that the root tonoplast-localized transporter regulates various auxin-related processes, while the truncated protein mediates drought tolerance by regulating stomatal closure. Heterologous expression in yeast revealed that the two splice forms share proton-coupled potassium transport activity. Thus, by determining the subcellular and tissue localization of two isoforms, alternative splicing allows the same gene to fulfill two very different but equally relevant roles in the plant.