CLONING AND PROPERTIES OF A RICE GENE ENCODING PHENYLALANINE AMMONIA-LYASE

Phenylalanine ammonia-lyase (PAL) genomic sequences were isolated from a rice (Oryza sativa L.) genomic library using a PCR-amplified rice P AL DNA fragment as a probe. There is a small family of PAL genes in th e rice genome. The nucleotide sequence of one PAL gene, ZB8, was deter mined. The ZB8 gene is 4660 bp in length and consists of two exons and one intron. It encodes a polypeptide of 710 amino acids. The transcri ption start site was 137 bp upstream from the translation initiation s ite. Rice PAL transcripts accumulated to a high level in stems, with l ower levels in roots and leaves. Wounding of leaf tissues induced ZB8 PAL transcripts to a high level. In rice suspension-cultured cells tre ated with fungal cell wall elicitors, the ZB8 PAL transcript increased within 30 min and reached maximum levels in 1-2 h. The transcription of the ZB8 gene was investigated by fusing its promoter to the reporte r gene beta-glucuronidase (GUS) and transforming the construct into ri ce and tobacco plants, as well as rice suspension-cultured cells. High levels of GUS activity were observed in stems, moderate levels in roo ts and low levels in leaves of transgenic rice and tobacco plants. His tochemical analysis indicated that in transgenic rice the promoter was active in root apical tips, lateral root initiation sites, and vascul ar and epidermal tissues of stems and roots. In rice flowers, high GUS activity was observed in floral shoots, receptacles, anthers and fila ments, occasionally GUS activity was also detected in lemma and awn ti ssues. In tobacco flowers, high GUS activity was detected in the pink part of petals. Consistent with the activity of endogenous PAL transcr ipts, wounding of rice and tobacco leaf tissues induced GUS activity f rom low basal levels. Tobacco mosaic virus (TMV) infection of tobacco leaves induced GUS activity to a high level. Fungal cell wall elicitor s strongly induced GUS activity and GUS transcripts to high levels in transgenic rice suspension-cultured cells. We demonstrated that the pr omoter of ZB8 gene is both developmentally regulated and stress-induci ble.