Rice tungro bacilliform virus (RTBV) replicates only in phloem cells in infected rice plants and its promoter drives strong phloem‐specific reporter gene expression in transgenic rice plants. We isolated a cDNA encoding a basic leucine zipper (bZIP) protein, RF2a, which binds to the Box II cis element that is important for expression from the promoter. RF2a, which stimulates Box II‐dependent transcription in a homologous in vitro transcription system, accumulates in nuclei of phloem and certain other cell types in shoots, but is found at only very low levels in roots. Transgenic antisense plants in which RF2a accumulation was suppressed had normal roots but stunted, twisted leaves with small, disorganized vascular bundles, an enlarged sclerenchyma and large air spaces. We propose that the RTBV promoter exploits a host transcription factor that is critical for leaf tissue differentiation and vascular development for its expression.

No abstract

BROTHER OF LUX ARRHYTHMO (BOA) is a GARP family transcription factor in Arabidopsis thaliana and is regulated by circadian rhythms. Transgenic lines that constitutively overexpress BOA exhibit physiological and developmental changes, including delayed flowering time and increased vegetative growth under standard growing conditions. Arabidopsis circadian clock protein CIRCADIAN CLOCK ASSOCIATED1 (CCA1) binds to the evening element of the BOA promoter and negatively regulates its expression. Furthermore, the period of BOA rhythm was shortened in cca1-11, lhy-21 (for LATE ELONGATED HYPOCOTYL), and cca1-11 lhy-21 genetic backgrounds. BOA binds to the promoter of CCA1 through newly identified promoter binding sites and activates the transcription of CCA1 in vivo and in vitro. In transgenic Arabidopsis lines that overexpress BOA, the period length of CCA1 rhythm was increased and the amplitude was enhanced. Rhythmic expression of other clock genes, including LHY, GIGANTEA (GI), and TIMING OF CAB EXPRESSION1 (TOC1), was altered in transgenic lines that overexpress BOA. Rhythmic expression of BOA was also affected in mutant lines of toc1-1, gi-3, and gi-4. Results from these studies indicate that BOA is a critical component of the regulatory circuit of the circadian clock.

No abstract

RF2a is a bZIP transcription factor that regulates expression of the promoter of rice tungro bacilliform badnavirus. RF2a is predicted to include three domains that contribute to its function. The results of transient assays with mutants of RF2a from which one or more domains were removed demonstrated that the acidic domain was essential for the activation of gene expression, although the proline-rich and glutamine-rich domains each played a role in this function. Studies using fusion proteins of different functional domains of RF2a with the 2C7 synthetic zinc finger DNA-binding domain showed that the acidic region is a relatively strong activation domain, the function of which is dependent on the context in which the domain is placed. Data from transgenic plants further supported the conclusion that the acidic domain was important for maintaining the biological function of RF2a. The severe stunting symptoms of rice tungro disease are caused by infection of rice tungro bacilliform virus (RTBV), 1 a double-stranded DNA badnavirus. Understanding the transcriptional regulation of RTBV is an important factor to elucidate the basis of the disease. RTBV carries a single, vascular tissue-specific promoter with several defined DNA cis-elements (1-4). Box II, one of the DNA cis-elements in the promoter, is essential for phloem-specific expression of the promoter (3, 4).A bZIP type rice host transcription factor, RF2a, was identified by its interaction with Box II (5). Furthermore, overexpression of RF2a in transgenic plants is sufficient to activate expression of RTBV promoter in other than vascular tissues (6).Temporal and spatial regulation of gene expression relies largely on the function of gene-specific transcription factors and is achieved by the activity of multiple proteins that bind to regulatory elements and with other proteins to alter basal rates of transcription initiation and/or elongation (7-9). A typical gene-specific eukaryotic transcription factor includes a DNAbinding domain and one or more domains that influence the activation or repression of transcription through interactions with general transcription factors, co-factors, chromatin remodeling complexes, and components of RNA polymerase II holoenzyme, among others (7, 10 -13). Transacting domains are often characterized as having a high content of specific amino acids, including domains rich in the acidic amino acids, proline or glutamine (14 -16). Acidic domains have been reported to possess activation functions that include interactions with TATA-binding proteins (TBP) (13, 17), TBP-associated factors (TAFs) (18), TFIIA (19), TFIIB (20, 21), other general transcription complexes (13,22), and co-factors (12). Proline-rich and glutamine-rich domains typically act through interactions with TBP, TAFs, and other co-factors (14). Although proline-rich and glutamine-rich domains act as activation domains in most of the cases, they can also function as repression domains (23,24). RF2a contains three putative transacting domains, namely proline-rich and ac...

No abstract

The promoter from rice tungro bacilliform badnavirus (RTBV) is expressed only in phloem tissues in transgenic rice plants. RF2a, a b-Zip protein from rice, is known to bind to the Box II cis element near the TATA box of the promoter. Here, we report that the full-length RTBV promoter and a truncated fragment E of the promoter, comprising nucleotides ؊164 to ؉45, result in phloemspecific expression of ␤-glucuronidase (GUS) reporter genes in transgenic tobacco plants. When a fusion gene comprising the cauliflower mosaic virus 35S promoter and RF2a cDNA was coexpressed with the GUS reporter genes, GUS activity was increased by 2-20-fold. The increase in GUS activity was positively correlated with the amount of RF2a, and the expression pattern of the RTBV promoter was altered from phloem-specific to constitutive. Constitutive expression of RF2a did not induce morphological changes in the transgenic plants. In contrast, constitutive overexpression of the b-ZIP domain of RF2a had a strong effect on the development of transgenic plants. These studies suggest that expression of the b-Zip domain can interfere with the function of homologues of RF2a that regulate development of tobacco plants. R egulation of transcription is achieved by the activity of multiple proteins that bind to regulatory elements, many of which are upstream of the promoters and alter basal rates of transcription initiation and͞or elongation (1, 2). To understand the mechanisms of tissue-specific and constitutive gene expression in plants, a number of promoters and transcription factors have been studied in recent years (3-16). It was shown that constitutive promoters, such as the cauliflower mosaic virus 35S promoter (17) and the promoter from cassava vein mosaic virus (14) are modular in organization and multiple cis elements. These elements, with specific transcription factors, apparently interact in an additive and͞or synergistic manner to confer gene expression in all plant tissues. Similarly, tissue-specific promoters contain multiple elements that contribute to promoter activity in both positive and negative ways (4-6, 8, 12, 18).The rice tungro bacilliform badnavirus (RTBV) promoter (19,20) and the transcription factors that interact it may serve as a model system to study plant tissue-specific gene expression. The virus, which replicates solely in phloem tissues, has a single promoter that is active in transfected protoplasts and is phloemspecific in transgenic rice plants (7,10,21,22).Within the fragment E of the promoter (nucleotides Ϫ164 to ϩ45), multiple cis elements were identified as being required for phloem-specific gene expression (7,10,15,23). A b-ZIP type transcription factor, RF2a, was isolated from rice and bound to Box II, a crucial cis element of the promoter. RF2a activates transcription from the RTBV promoter in an in vitro transcription system derived from rice cell cultures (11). Moreover, studies in transgenic rice plants suggested that RF2a is involved in the development of vascular tissues (11).We report here the functi...

No abstract