title: PROTEIN L-ISOASPARTYL METHYLTRANSFERASE2 is differentially expressed in chickpea and enhances seed vigor and longevity by reducing abnormal isoaspartyl accumulation predominantly in seed nuclear proteins.
abstract: PROTEIN l-ISOASPARTYL METHYLTRANSFERASE (PIMT) is a widely distributed protein-repairing enzyme that catalyzes the conversion of abnormal l-isoaspartyl residues in spontaneously damaged proteins to normal aspartyl residues. This enzyme is encoded by two divergent genes (PIMT1 and PIMT2) in plants, unlike many other organisms. While the biological role of PIMT1 has been elucidated, the role and significance of the PIMT2 gene in plants is not well defined. Here, we isolated the PIMT2 gene (CaPIMT2) from chickpea (Cicer arietinum), which exhibits a significant increase in isoaspartyl residues in seed proteins coupled with reduced germination vigor under artificial aging conditions. The CaPIMT2 gene is found to be highly divergent and encodes two possible isoforms (CaPIMT2 and CaPIMT2') differing by two amino acids in the region I catalytic domain through alternative splicing. Unlike CaPIMT1, both isoforms possess a unique 56-amino acid amino terminus and exhibit similar yet distinct enzymatic properties. Expression analysis revealed that CaPIMT2 is differentially regulated by stresses and abscisic acid. Confocal visualization of stably expressed green fluorescent protein-fused PIMT proteins and cell fractionation-immunoblot analysis revealed that apart from the plasma membrane, both CaPIMT2 isoforms localize predominantly in the nucleus, while CaPIMT1 localizes in the cytosol. Remarkably, CaPIMT2 enhances seed vigor and longevity by repairing abnormal isoaspartyl residues predominantly in nuclear proteins upon seed-specific expression in Arabidopsis (Arabidopsis thaliana), while CaPIMT1 enhances seed vigor and longevity by repairing such abnormal proteins mainly in the cytosolic fraction. Together, our data suggest that CaPIMT2 has most likely evolved through gene duplication, followed by subfunctionalization to specialize in repairing the nuclear proteome.
authors.affiliation: National Institute of Plant Genome Research, Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Pooja
authors.initials: P
authors.lastname: Verma
authors.forename: Harmeet
authors.initials: H
authors.lastname: Kaur
authors.forename: Bhanu Prakash
authors.initials: BP
authors.lastname: Petla
authors.forename: Venkateswara
authors.initials: V
authors.lastname: Rao
authors.forename: Saurabh C
authors.initials: SC
authors.lastname: Saxena
authors.forename: Manoj
authors.initials: M
authors.lastname: Majee
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D000595:Amino Acid Sequence; D015153:Blotting, Western; D029863:Cicer / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D003600:Cytosol / Q000378:metabolism; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics; D018525:Germination / Q000235:genetics; D026581:Isoaspartic Acid / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D009928:Organ Specificity / Q000235:genetics; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012639:Seeds / Q000235:genetics / Q000254:growth & development*; D013347:Subcellular Fractions / Q000201:enzymology; D014170:Transformation, Genetic
chemical_list: D026581:Isoaspartic Acid; D009687:Nuclear Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase