Alien are highly repeated plant transposable elements characterized by their small size (approx. 400 bp), high A + T content, target site specificity, potential to form stable secondary structures and possession of a conserved 28-bp terminal inverted repeat (TIR). Besides the TIR, they contain subterminal inverted repeat motifs (SIRM), as well as the 5'-CATGCAT domain which has been reported to be a cis-acting regulatory element of gene expression in some plant species. Although they were first identified in the intron of the bell pepper (Capsicum annuum) Sn-2 gene and in the promoter region of the potato starch phosphorylase-encoding gene, Alien arranged in tandem are present in the promoter of patatin class-II genes. PCR on the bell pepper genomic DNA using the Alien TIR consensus sequence as primer yielded DNA fragments of nearly 400 bp. These fragments have characteristics of transposable elements and contain numerous motifs reminiscent of Alien elements. Importantly, PCR on genomic DNA extracts from various monocotyledonous and dicotyledonous plants using the TIR consensus sequence as primer and subsequent hybridization with different Alien probes revealed that these elements are ubiquitously present and highly repeated in the genomes of higher plants.

The basic PR-1 gene, CABPR1, accumulates in pepper leaf tissues during pathogen infection as well as after ethylene treatment. We isolated and functionally characterized the CABPR1 promoter region in tobacco leaves to identify the cis-acting regulatory sequences that are involved in CABPR1 gene expression. Constructs harboring the 5'-serially deleted CABPR1 promoter, which was fused to the beta-glucuronidase (GUS) gene, were evaluated for their promoter activity in the tobacco leaves. The CABPR1 promoter of 1670 bp in size was locally or systemically induced during a compatible interaction with Pseudomonas syringae pv. tabaci. The CABPR1 promoter also was differentially activated by treatment with ethylene, salicylic acid, nitric oxide, high salinity, drought and low temperature. The expression of the pepper transcription factors, CAZFP1 and CARAV1, activated the CABPR1 promoter. Analyses of a series of 5'-deletions of the CABPR1 promoter indicated that novel cis-acting elements essential for induction by pathogen and abiotic elicitors are localized in the region between -1670 bp and -1466 bp upstream from the translation start site. These results suggest that CABPR1 promoter is essential for regulating CABPR1 gene expression in response to pathogen, abiotic and environmental stresses, possibly by transactivating the CAZFP1 and CARAV1 transcription factors.

We find that salicylic acid and ethephon treatment in hot pepper increases the expression of a putative basic/leucine zipper (bZIP) transcription factor gene, CabZIP1. CabZIP1 mRNA is expressed ubiquitously in various organs. The green fluorescent protein-fused transcription factor, CabZIP1::GFP, can be specifically localized to the nucleus, an action that is consistent with the presence of a nuclear localization signal in its protein sequence. Transient overexpression of the CabZIP1 transcription factor results in an increase in PR-1 transcripts level in Nicotiana benthamiana leaves. Using chromatin immunoprecipitation, we demonstrate that CabZIP1 binds to the G-box elements in native promoter of the hot pepper pathogenesis-related protein 1 (CaPR-1) gene in vivo. Taken together, our results suggest that CabZIP1 plays a role as a transcriptional regulator of the CaPR-1 gene.