Transcriptional regulation of plant phosphate transporters.
(17/2576)
Phosphorus is acquired by plant roots primarily via the high-affinity inorganic phosphate (Pi) transporters. The transcripts for Pi transporters are highly inducible upon Pi starvation, which also results in enhanced Pi uptake when Pi is resupplied. Using antibodies specific to one of the tomato Pi transporters (encoded by LePT1), we show that an increase in the LePT1 transcript under Pi starvation leads to a concurrent increase in the transporter protein, suggesting a transcriptional regulation for Pi acquisition. LePT1 protein accumulates rapidly in tomato roots in response to Pi starvation. The level of transporter protein accumulation depends on the Pi concentration in the medium, and it is reversible upon resupply of Pi. LePT1 protein accumulates all along the roots under Pi starvation and is localized primarily in the plasma membranes. These results clearly demonstrate that plants increase their capacity for Pi uptake during Pi starvation by synthesis of additional transporter molecules. (+info)
Mutagenesis of amino acids at two tomato ringspot nepovirus cleavage sites: effect on proteolytic processing in cis and in trans by the 3C-like protease.
(18/2576)
Tomato ringspot nepovirus (ToRSV) encodes two polyproteins that are processed by a 3C-like protease at specific cleavage sites. Analysis of ToRSV cleavage sites identified previously and in this study revealed that cleavage occurs at conserved Q/(G or S) dipeptides. In addition, a Cys or Val is found in the -2 position. Amino acid substitutions were introduced in the -6 to +1 positions of two ToRSV cleavage sites: the cleavage site between the protease and putative RNA-dependent RNA polymerase, which is processed in cis, and the cleavage site at the N-terminus of the movement protein, which is cleaved in trans. The effect of the mutations on proteolytic processing at these sites was tested using in vitro translation systems. Substitution of conserved amino acids at the -2, -1, and +1 positions resulted in a significant reduction in proteolytic processing at both cleavage sites. The effects of individual substitutions were stronger on the cleavage site processed in trans than on the one processed in cis. The cleavage site specificity of the ToRSV protease is discussed in comparison to that of related proteases. (+info)
Interaction of NPR1 with basic leucine zipper protein transcription factors that bind sequences required for salicylic acid induction of the PR-1 gene.
(19/2576)
The Arabidopsis thaliana NPR1 has been shown to be a key regulator of gene expression during the onset of a plant disease-resistance response known as systemic acquired resistance. The npr1 mutant plants fail to respond to systemic acquired resistance-inducing signals such as salicylic acid (SA), or express SA-induced pathogenesis-related (PR) genes. Using NPR1 as bait in a yeast two-hybrid screen, we identified a subclass of transcription factors in the basic leucine zipper protein family (AHBP-1b and TGA6) and showed that they interact specifically in yeast and in vitro with NPR1. Point mutations that abolish the NPR1 function in A. thaliana also impair the interactions between NPR1 and the transcription factors in the yeast two-hybrid assay. Furthermore, a gel mobility shift assay showed that the purified transcription factor protein, AHBP-1b, binds specifically to an SA-responsive promoter element of the A. thaliana PR-1 gene. These data suggest that NPR1 may regulate PR-1 gene expression by interacting with a subclass of basic leucine zipper protein transcription factors. (+info)
Quantitative immunofluorescence of regulated eps gene expression in single cells of Ralstonia solanacearum.
(20/2576)
Ralstonia solanacearum, a phytopathogenic bacterium, uses an environmentally sensitive and complex regulatory network to control expression of multiple virulence genes. Part of this network is an unusual autoregulatory system that produces and senses 3-hydroxypalmitic acid methyl ester. In culture, this autoregulatory system ensures that expression of virulence genes, such as those of the eps operon encoding biosynthesis of the acidic extracellular polysaccharide, occurs only at high cell density (>10(7) cells/ml). To determine if regulation follows a similar pattern within tomato plants, we first developed a quantitative immunofluorescence (QIF) method that measures the relative amount of a target protein within individual bacterial cells. For R. solanacearum, QIF was used to determine the amount of beta-galactosidase protein within wild-type cells containing a stable eps-lacZ reporter allele. When cultured cells were examined to test the method, QIF accurately detected both low and high levels of eps gene expression. QIF analysis of R. solanacearum cells recovered from stems of infected tomato plants showed that expression of eps during pathogenesis was similar to that in culture. These results suggest that there are no special signals or conditions within plants that override or short-circuit the regulatory processes observed in R. solanacearum in culture. Because QIF is a robust, relatively simple procedure that uses generally accessible equipment, it should be useful in many situations where gene expression in single bacterial cells must be determined. (+info)
Multiple epoxide hydrolases in Alternaria alternata f. sp. lycopersici and their relationship to medium composition and host-specific toxin production.
(21/2576)
The production of Alternaria alternata f. sp. lycopersici host-specific toxins (AAL toxins) and epoxide hydrolase (EH) activity were studied during the growth of this plant-pathogenic fungus in stationary liquid cultures. Media containing pectin as the primary carbon source displayed peaks of EH activity at day 4 and at day 12. When pectin was replaced by glucose, there was a single peak of EH activity at day 6. Partial characterization of the EH activities suggests the presence of three biochemically distinguishable EH activities. Two of them have a molecular mass of 25 kDa and a pI of 4.9, while the other has a molecular mass of 20 kDa and a pI of 4.7. Each of the EH activities can be distinguished by substrate preference and sensitivity to inhibitors. The EH activities present at day 6 (glucose) or day 12 (pectin) are concomitant with AAL toxin production. (+info)
Gene for a protein capable of enhancing lateral root formation.
(22/2576)
Analysis of genes preferentially expressed in hairy roots caused by infection with Agrobacterium rhizogenes has provided insights into the regulation of lateral root formation. A hairy root preferential cDNA, HR7, has been cloned from hairy roots of Hyoscyamus niger. HR7 encodes a novel protein partially homologous to a metallocarboxypeptidase inhibitor and is expressed exclusively in the primordium and base of lateral roots in hairy roots. Overexpression of HR7 in transgenic roots of H. niger dramatically enhances the frequency of lateral root formation. The results of this study indicate that expression of HR7 plays a critical role in initiating lateral root formation. (+info)
Identification of replication specificity determinants in two strains of tomato leaf curl virus from New Delhi.
(23/2576)
We used two strains of tomato leaf curl virus from New Delhi to investigate specificity in replication of their cognate genomes. The strains share 94% sequence identity and are referred to as severe and mild on the basis of symptoms on tomato and tobacco. Replication assays in tobacco protoplasts and plants showed that a single amino acid change, Asn10 to Asp in the N terminus of Rep protein, determines specificity for replication of the two strains based upon its interaction with the origin of replication (ori) sequences. The change of Asp10 to Asn in Rep protein of the mild strain coupled with point mutations at the 3rd and 10th nucleotides of the 13-mer binding site altered its replication ability, resulting in increased levels of virus accumulation. Similarly, changing Asn10 to Asp in Rep protein of the severe strain impaired replication of the virus and altered its severe phenotype in plants. Site-directed mutations made in ori and Asn10 of Rep protein suggested that Asn10 recognizes the third base pair of the putative binding site sequence GGTGTCGGAGTC in the severe strain. (+info)
Constitutively active Pto induces a Prf-dependent hypersensitive response in the absence of avrPto.
(24/2576)
Resistance in tomato to Pseudomonas syringae pv tomato (avrPto) is conferred by the gene Pto in a gene-for-gene relationship. A hypersensitive disease resistance response (HR) is elicited when Pto and avrPto are expressed experimentally within the same plant cell. The kinase capability of Pto was required for AvrPto-dependent HR induction. Systematic mutagenesis of the activation segment of Pto kinase confirmed the homologous P+1 loop as an AvrPto-binding determinant. Specific amino acid substitutions in this region led to constitutive induction of HR upon expression in the plant cell in the absence of AvrPto. Constitutively active Pto mutants required kinase capability for activity, and were unable to interact with proteins previously shown to bind to wild-type Pto. The constitutive gain-of-function phenotype was dependent on a functional Prf gene, demonstrating activation of the cognate disease resistance pathway and precluding a role for Prf upstream of Pto. (+info)