Changing transcriptional initiation sites and alternative 5′- and 3′-splice site selection of the first intron deploys Arabidopsis PROTEIN ISOASPARTYL METHYLTRANSFERASE2 variants to different subcellular compartments

Arabidopsis thaliana (L.) Heynh. possesses two PROTEIN-L-ISOASPARTATE METHYLTRANSFERASE (PIMT) genes encoding enzymes (EC 2.1.1.77) capable of converting uncoded l-isoaspartyl residues, arising spontaneously at l-asparaginyl and l-aspartyl sites in proteins, to l-aspartate. PIMT2 produces at least eight transcripts by using four transcriptional initiation sites (TIS; resulting in three different initiating methionines) and both 5′- and 3′-alternative splice site selection of the first intron. The transcripts produce mature proteins capable of converting l-isoaspartate to l-aspartate in small peptide substrates. PIMT:GFP fusion proteins generated a detectable signal in the nucleus. However, whether the protein was also detectable in the cytoplasm, endo-membrane system, chloroplasts, and/or mitochondria, depended on the transcript from which it was produced. On-blot-methylation of proteins, prior to the completion of germination, indicated that cruciferin subunits contain isoaspartate. The implications of using transcriptional mechanisms to expand a single gene's repertoire to protein variants capable of entry into the cell's various compartments are discussed in light of PIMT's presumed role in repairing the proteome.