Multiple cis regulatory elements for maximal expression of the cauliflower mosaic virus 35S promoter in transgenic plants.

The 35S promoter is a major promoter of the cauliflower mosaic virus that infects crucifers. This promoter is still active when excised from cauliflower mosaic virus and integrated into the nuclear genome of transgenic tobacco. Previous work has shown that the -343 to -46 upstream fragment is responsible for the majority of the 35S promoter strength (Odell, J.

Transcriptional interaction between the promoters of the maize chloroplast genes which encode the beta subunit of ATP synthase and the large subunit of ribulose 1,5-bisphosphate carboxylase.

The genes encoding the beta subunit of ATP synthase and the large subunit of ribulose 1,5-bisphosphate carboxylase are located on opposite strands of the maize chloroplast genome. Their transcription start sites are separated by a 159 bp sequence that includes the promoters for both genes. The effects of deleting or modifying one of the two promoters on transcription from the adjacent, unaltered promoter were assessed in vitro using maize chloroplast extracts to transcribe cloned maize DNA templates.

Binding site requirements for pea nuclear protein factor GT-1 correlate with sequences required for light-dependent transcriptional activation of the rbcS-3A gene.

Nuclear protein factor GT-1 binds to sequence boxes II, III, II* and III* upstream of the light-responsive pea rbcS-3A gene. We have shown previously that box II and box III are required for expression of rbcS-3A when redundant elements upstream of -170 (relative to the transcription start site) are removed. Here we present evidence that deletion and substitution mutations downstream of -170 which eliminate expression also decrease binding.

Dissection of 5' upstream sequences for selective expression of the Nicotiana plumbaginifolia rbcS-8B gene.

We have previously isolated and characterized a gene (rbcS-8B) from the wild-type species Nicotiana plumbaginifolia, encoding the small subunit of ribulose-1,5-bisphosphate carboxylase. Using transgenic N. plumbaginifolia as a host, we found that a 5' upstream region (-1038 to +32) of rbcS-8B contains all the sequences required for organ-specific and light-dependent expression of the gene.

Abscisic acid and water-stress induce the expression of a novel rice gene.

We have identified a novel rice gene, called RAB 21, which is induced when plants are subject to water-stress. This gene encodes a basic, glycine-rich protein (mol. wt 16,529) which has a duplicated domain structure.

Transcription of the wheat chloroplast gene that encodes the 32 kd polypeptide.

We have mapped and cloned the wheat chloroplast gene (psbA) that encodes the 32 kd polypeptide of Photosystem II. The psbA gene is located in the large single copy region adjacent to one inverted repeat and is transcribed toward the latter. The sequence of the 5' end of the wheat gene is homologous with dicot psbA genes.

Localization and conditional redundancy of regulatory elements in rbcS-3A, a pea gene encoding the small subunit of ribulose-bisphosphate carboxylase.

Expression of the pea rbcS-3A gene, one of a family of genes encoding the small subunit of ribulose-bisphosphate carboxylase [EC 4.1.1.

Characterization of a chloroplast sequence-specific DNA binding factor.

The large subunit of ribulose 1,5-bisphosphate carboxylase (rbcL) and the beta subunit of chloroplast ATP synthase (atpB) are encoded by divergently transcribed genes on the plastid genome. We have identified DNA binding factors specific for sequences located in the intergenic region between these two genes. Soluble plastid extracts from pea or whole cell extracts from maize protected a maize chloroplast DNA probe containing the 160-base pair region between the 5' ends of rbcL and atpB genes from exonuclease III digestion between positions -16 and -101 relative to the rbcL gene transcription start site.

Analysis of the mechanism of protection in transgenic plants expressing the potato virus X coat protein or its antisense RNA.

Transgenic tobacco plants engineered to express either the potato virus X (PVX) coat protein (CP+) or the antisense coat protein transcript (CP-antisense) were protected from infection by PVX, as indicated by reduced lesion numbers on inoculated leaves, delay or absence of systemic symptom development and reduction in virus accumulation in both inoculated and systemic leaves. The extent of protection observed in CP+ plants primarily depended upon the level of expression of the coat protein. Plants expressing antisense RNA were protected only at low inoculum concentrations.

Cell-specific expression of pyruvate, pi dikinase : in situ mRNA hybridization and immunolocalization labeling of protein in wheat seed.

Pyruvate, Pi dikinase (PPDK) is a key enzyme in the C4 photosynthetic pathway. However, its metabolic role in C3 plants remains uncertain. Northern blot analyses of PPDK mRNAs from wheat leaves and seeds probed with maize PPDK cDNA indicates the presence of organ-specific mRNAs.

Artificial combination of two cis-regulatory elements generates a unique pattern of expression in transgenic plants.

We show that a 36-base-pair-long upstream fragment from the soybean hsp17.3-B gene comprising two partly overlapping heat-shock element (HSE)-like sequences can confer heat inducibility to a reporter gene in transgenic tobacco. The heat-shock response does not display organ specificity and is not affected by light.

Chloroplast DNA gyrase and in vitro regulation of transcription by template topology and novobiocin.

We have examined the effects of novobiocin and template topology on the transcription of two chloroplast genes encoding the large subunit of ribulose 1,5-bisphosphate carboxylase (rbcL) and the beta subunit of the chloroplast ATPase (atpB), in an in vitro transcription system. The template topology was monitored by agarose gel electrophoresis while the in vitro transcripts were determined by 5' S1 nuclease analysis under identical conditions. We discovered that our chloroplast transcription extracts contain a DNA gyrase activity and a chromatographically separable topoisomerase I activity.

Sequence-specific interactions of a pea nuclear factor with light-responsive elements upstream of the rbcS-3A gene.

Pea nuclear extracts were used in gel retardation assays and DNase I footprinting experiments to identify a protein factor that specifically interacts with regulatory DNA sequences upstream of the pea rbcS-3A-gene. This factor, designated GT-1, binds to two short sequences (boxes II and III) in the -150 region that are known to function as light-responsive elements (LREs) in transgenic tobacco. Binding of GT-1 to homologous sequences further upstream (boxes II and III in the -220 region) indicates that these boxes comprise the redundant LRE that functions in vivo when boxes II and III are deleted.

The 5'-proximal region of the wheat Cab-1 gene contains a 268-bp enhancer-like sequence for phytochrome response.

We have previously reported that the expression of the wheat Cab-1 gene is subject to phytochrome regulation and a 1.8-kb 5' upstream sequence of this gene is sufficient for the regulated expression. To delineate sequences for the phytochrome response we analyzed a series of 5' deletion mutants as well as chimeric gene constructs comprising different sequences of the Cab-1 upstream region in transgenic tobacco seedlings.

Targeting of bacterial chloramphenicol acetyltransferase to mitochondria in transgenic plants.

Most mitochondrial proteins are encoded by nuclear genes and are synthesized as precursors containing a presequence at the N terminus. In yeast and in mammalian cells, the function of the presequence in mitochondrial targeting has been revealed by chimaeric gene studies. Fusion of a mitochondrial presequence to a foreign protein coding sequence enables the protein to be imported into mitochondria in vitro as well as in vivo.

Sequences in the pea rbcS-3A gene have homology to constitutive mammalian enhancers but function as negative regulatory elements.

The pea rbcS-3A gene, which codes for a key component of the photosynthetic machinery, requires light for its expression. Analysis of chimeric constructs in transgenic tobacco plants has shown that a 280-bp fragment from the 5' noncoding region can act as a light-inducible transcriptional enhancer. Further characterization of this enhancer identifies a 58-bp sequence containing two regulatory elements that can decrease transcription in the dark.

Plant cells do not properly recognize animal gene polyadenylation signals.

We have introduced chimeric genes containing polyadenylation signals from a human gene and two animal virus genes into tobacco cells. We see, in all three cases, inefficient and 'aberrant' utilization of the foreign polyadenylation signals. We find that a chimeric gene carrying the polyadenylation site of the human growth hormone gene is polyadenylated at three sites in the vicinity of the site that is polyadenylated in human cells.

Monocot and dicot pre-mRNAs are processed with different efficiencies in transgenic tobacco.

A gene encoding the small subunit of ribulose 1,5-bisphosphate carboxylase (rbcS) in wheat, a monocot plant, was transferred to tobacco, a dicot plant. The wheat gene is not expressed in transgenic tobacco under the control of its own promoter, but when transcription is driven by a viral promoter, several wheat transcripts accumulate. These include both spliced and unspliced transcripts, which are polyadenylated at multiple novel sites in the wheat 3' flanking region.

Expression dynamics of the pea rbcS multigene family and organ distribution of the transcripts.

We have determined the nucleotide sequence of two members (rbcS-3A and -3C) of the pea nuclear gene family encoding the small subunit (rbcS) of ribulose-1,5-bisphosphate carboxylase. Both rbcS-3A and -3C are interrupted by two introns located at the same positions as those of the other three pea rbcS genes. Compared with the other pea rbcS genes the rbcS-3C gene has the most divergent 5'- and 3'-flanking sequences while the rbcS-3A gene has a larger and highly divergent intron 1.

Phytochrome-controlled expression of a wheat Cab gene in transgenic tobacco seedlings.

We have monitored changes in the chlorophyll a/b-binding protein (Cab) mRNA levels in etiolated wheat leaves exposed to light of different wavelengths by Northern blot hybridizations and 5' S1 nuclease protection assays. Accumulation of the Cab mRNA and the specific Cab-1 transcript is regulated by phytochrome. Transcript levels are elevated by red light and the red enhancement can be partially reversed by far-red light.

Organ-specific and light-induced expression of plant genes.

Light plays a pivotal role in the development of plants. The photoregulation of plant genes involves recognition of light quality and quantity by phytochrome and other light receptors. Two gene families, rbcS and Cab, which code for abundant proteins active in photosynthesis, the small subunit of ribulose bisphosphate carboxylase and the chlorophyll a/b binding protein, show a 20-to 50-fold increase in transcript abundance in the light.

Developmental regulation of two genes encoding ribulose-bisphosphate carboxylase small subunit in pea and transgenic petunia plants: Phytochrome response and blue-light induction.

We describe the effects of light quality on the light-induced expression of two genes (rbcS-3A and rbcS-3C) encoding the ribulose-1,5-bisphosphate carboxylase small subunit of pea plants. These two genes exhibit distinctive photoresponses depending on the developmental stages of the leaves. In etiolated primary leaves, changes in the rbcS-3A and rbcS-3C transcript levels are regulated by phytochrome.

Photoregulated expression of a pea rbcS gene in leaves of transgenic plants.

A 2.4-kb pea genomic fragment, containing a member (rbcS-E9) of the multigene family encoding the small subunit (rbcS) of ribulose-1,5-bisphosphate carboxylase, was inserted into a non-oncogenic, Ti-plasmid vector and introduced into the genomes of Petunia hybrida (Mitchell) and Nicotiana tabacum (SR1) plants by in vitro transformation. Petunia and tobacco plants containing the introduced pea rbcS-E9 gene were regenerated from protoplasts.