Primary structure of a novel barley gene differentially expressed in immature aleurone layers

As a direct approach to elucidate the molecular biology of barley aleurone cell development, we differentially screened an aleurone cDNA library made from poly(A)+ RNA of immature grains for clones representing transcripts present in the aleurone but not in the starchy endosperm. For one of these clones, B22E, which hybridies to a 0.7 kb transcript, Northern and in situ hybridization revealed that expression is under complex spatial, temporal and hormonal control in barley grains. cDNAs corresponding to B22E transcripts were isolated from aleurone/pericarp and embryo of developing grains, and from germinating scutella. Among these were the nearly full-length aleurone/pericarp clone pB22E.a16 (541 bp). cDNAs matching the sequence of this clone (type 1 transcript) were found for all tissues investigated. In addition, cDNAs with an extra 12 bp insertion (type 2 transcript) were obtained from germinating scutella. The two different transcripts can encode novel barley proteins of 115 and 119 amino acids, respectively. A gene designated B22EL8 was isolated and sequenced; it encodes the type 1 B22E transcript and contains two introns of 145 and 125 bp. Particle bombardment of barley aleurone with a B22EL8 promoter-GUS (beta-glucuronidase) construct demonstrates that the promoter (3 kb) is active in developing barley grains. The promoter is not, however, active in the seeds of tobacco plants transgenic for the B22EL8 gene, indicating the existence of sequences specific for monocots. A comparison of 1.4 kb of upstream sequence of B22E with the maize c1 promoter reveals a number of short, identical sequences which may be responsible for aleurone cell-specific gene transcription.