Gene expression in physically impeded maize roots

Abstract

Two approaches were used to search for genes which respond to physical impedance. First, cDNA clones induced by mechanical stress or drought stress of other plant species were hybridized to mRNA from maize root tips. The results showed that only two clones, TCH1 induced by wind stress in Arabidopsis, and LP2 induced by drought stress in pine, had high homology with the RNA in maize root tips, but they did not reveal an inducible pattern of expression in the impeded maize roots tips. Second, a cDNA library was constructed from mMRNA from a 10 min physical impedance treatment of maize roots tips and was differentially screened with radioactive labeled cDNA probes synthesized using mRNA extracted from stressed and non-stressed maize roots tips. Three clones, PIIGI, pIIG2, and pIRG3, were identified as responding to a 10 min physical impedance stress. The first two cDNA clones (PIIGI and pIIG2), whose expressions were induced in a 10 min physical impedance treatment, were characterized further. cDNA PIIGI contains 678 hp with an open reading frame which specifies a polypeptide of 129 amino acid residues which showed 97% similarity at the nucleic acid level to maize root cortical cell delineating protein. Northern analysis with cDNA PIIGI as a probe showed that the expression was strongly induced by the 10 min physical impedance treatment and genomic Southern analysis showed that a relatively conserved gene family exists in maize. The CDNA pIIG2 has a nucleotide sequence of 830 bp with an open reading frame which specifies a polypeptide of 210 amino acid residues, but in a search of the GENBANK database it did not show significant homology with any identified gene of known function. Genomic Southern hybridization using cDNApIIG2 found duplicated loci in maize but single loci in rice. The third cDNA clone pIRG3, 800 bp, whose expression was reduced about 33% by 10 to 30 min physical impedance, is identical to the partial sequence of maize "calreticulin!' gene by GENBANK search.