Overexpression of RNA-binding bacterial chaperones in rice leads to stay-green phenotype, improved yield and tolerance to salt and drought stresses

Abstract

Genes encoding bacterial cold shock proteins A (CspA, 213 bp) and B (CspB, 216 bp) were isolated from Escherichia coli strain K12, which showed 100% homology with gene sequences isolated from other bacterial species. In silico domain, analysis showed eukaryotic conserved cold shock domain (CSD) and ribonuclease-binding domain (RBD) indicating that they bind to RNA and are involved in temperature stress tolerance. Overexpression of these two genes in E. coli resulted in higher growth in presence of 200 mM NaCl and 300 mM mannitol. Western blot confirmed the translational products of the two genes. Seedlings of indica rice were transformed with Agrobacterium tumefaciens containing pCAMBIA1301 CspA and CspB genes. Transgene integration was confirmed by β-glucuronidase (GUS) histochemical assay, polymerase chain reaction (PCR) amplification, and gene copy number by Southern blotting. Chlorophyll, proline, Na+, and K+ contents were higher in transgenics exposed to 150 mM NaCl and drought (imposed by withholding water) stresses during floral initiation stage. Catalase (CAT), superoxide dismutase (SOD), and guaiacol peroxidase (GPX) activities increased, while malondialdehyde (MDA) content was low in transgenics. Transgenics displayed increased root, shoot, and panicle lengths, root dry mass, and a distinct stay-green (SGR) phenotype. Higher transcript levels of CspA, CspB, SGR, chlorophyllase, isopentenyl adenine transferase 1 (IPT1), 9-cis-epoxycarotenoid dioxygenase (NCED), SOD, and sirtuin 1 (SIRT1) genes were observed in transgenics compared to wild type plants (WT) under multiple stresses. Present work indicates that bacterial chaperone proteins are capable of imparting SGR phenotype, salt and drought stress tolerance alongside grain improvement.