The large-scale organization of metabolic networks

A highly active synthetic auxin response element (AuxRE), referred to as DR5, was created by performing site-directed mutations in a natural composite AuxRE found in the soybean GH3 promoter. DR5 consisted of tandem direct repeats of 11 bp that included the auxin-responsive TGTCTC element. The DR5 AuxRE showed greater auxin responsiveness than a natural composite AuxRE and the GH3 promoter when assayed by transient expression in carrot protoplasts or in stably transformed Arabidopsis seedlings, and it provides a useful reporter gene for studying auxin-responsive transcription in wild-type plants and mutants. An auxin response transcription factor, ARF1, bound with specificity to the DR5 AuxRE in vitro and interacted with Aux/IAA proteins in a yeast two-hybrid system. Cotransfection experiments with natural and synthetic AuxRE reporter genes and effector genes encoding Aux/IAA proteins showed that overexpression of Aux/IAA proteins in carrot protoplasts resulted in specific repression of TGTCTC AuxRE reporter gene expression.

Aux/IAA proteins repress expression of reporter genes containing natural and highly active synthetic auxin response elements.

1. Introductory remarks 2. Convolvulacaea 2. Scrophulariaceae, Gesneriaceae, Labiatae, etc. 4. Cruciferae, Papaveraceae, Resedaceae, etc. 5. Geraniaceae, Leguminosae, Onagraceae, etc. 6. Solanaceae, Primulaceae, Polygoneae, etc. 7. Summary of the heights and weights of the crossed and self-fertilised plants 8. Difference between crossed and self-fertilised plants in constitutional vigour and in other respects 9. The effects of cross-fertilisation and self-fertilisation on the production of seeds 10. Means of fertilisation 11. The habits of insects in relation to the fertilisation of flowers 12. General results Index.

The Effects of Cross and Self Fertilisation in the Vegetable Kingdom: CONVOLVULACEÆ

The plant hormone abscisic acid (ABA) plays a major role in seed maturation and germination, as well as in adaptation to abiotic environmental stresses. ABA promotes stomatal closure by rapidly altering ion fluxes in guard cells. Other ABA actions involve modifications of gene expression, and the analysis of ABA-responsive promoters has revealed a diversity of potential cis-acting regulatory elements. The nature of the ABA receptor(s) remains unknown. In contrast, combined biophysical, genetic, and molecular approaches have led to considerable progress in the characterization of more downstream signaling elements. In particular, substantial evidence points to the importance of reversible protein phosphorylation and modifications of cytosolic calcium levels and pH as intermediates in ABA signal transduction. Exciting advances are being made in reassembling individual components into minimal ABA signaling cascades at the single-cell level.

Abscisic acid signal transduction

A molecular approach to investigate auxin signaling in plants has led to the identification of several classes of early/primary auxin response genes. Within the promoters of these genes, cis elements that confer auxin responsiveness (referred to as auxin-response elements or AuxREs) have been defined, and a family of trans-acting transcription factors (auxin-response factors or ARFs) that bind with specificity to AuxREs has been characterized. A family of auxin regulated proteins referred to as Aux/IAA proteins also play a key role in regulating these auxin-response genes. Auxin may regulate transcription on early response genes by influencing the types of interactions between ARFs and Aux/IAAs.

https://link.springer.com/content/pdf/10.1023%2FA%3A1015207114117.pdf

Auxin-responsive gene expression: genes, promoters and regulatory factors.

SELF-INCOMPATIBILITY in flowering plants is often controlled by a single nuclear gene (the S-gene) having several alleles1. This gene prevents fertilization by self-pollen or by pollen bearing either of the two S- alleles expressed in the style. Sequence analysis shows that three alleles of the S gene of Nicotiana alata encode style glycoproteins2,3 with regions of defined homology. Two of the homologous regions also show precise homology with ribonucleases T2 (ref. 4) and Rh (ref. 5). We report here that glycoproteins corresponding to the S1, S2, S3, S6 and S7 alleles isolated from style extracts of N. alata6 are ribonucleases. These style S-gene-encoded glycoproteins account for most of the ribonuclease activity recovered from style extracts. The ribonuclease specific activity of style extracts of the self-incompatible species N. alata is 100–1,000-fold higher than that of the related self-compatible species N. tabacum. These observations implicate ribonuclease activity in the mechanism of gametophytic self-incompatibility.

Style self-incompatibility gene products of Nicotlana alata are ribonucleases

Many angiosperms employ self-incompatibility systems to prevent inbreeding. The simple genetics of such systems have made them attractive models of plant cellular communication. Implicit in the single locus genetics is that only one or a few gene products are necessary for recognition and rejection of incompatible pollen. Results in the sporophytic system of the Brassicaceae suggest that different S-locus products are responsible for the pollen and pistil parts of the recognition and rejection response. In solanaeceous plants, which have a gametophytic self-incompatibility system, the S locus product responsible for the pollen portion of the interaction has not been identified, but ribonucleases encoded by the S-locus (S-RNases) are strongly implicated in the style part of the recognition and rejection reaction. In Nicotiana alata, pollen recognition and rejection occur if its S-allele matches either S-allele in the style. The putative stylar S-RNase is abundant in the transmitting tract, and pollen rejection may be related to action of S-RNase on pollen RNAs. Efforts to understand the molecular basis for pollen recognition and rejection have been limited by the lack of a system for manipulating and expressing S-RNases. Here we use the promoter of a style-expressed gene from tomato to obtain high levels of S-RNase expression in transgenic Nicotiana. Recognition and rejection of N. alata pollen S-alleles occur faithfully in the transgenic plants. Our results show that S-RNases alone are sufficient for pollen rejection in this system.

S-RNase expressed in transgenic Nicotiana causes S-allele-specific pollen rejection

GAMETOPHYTIC self-incompatibility is a genetically based system of cellular recognition in plants1. It prevents fertilization by pollen bearing an S-allele identical to either of the two S-alleles present in the female sporophytic tissues. Self-incompatibility in the Solanaceae has been especially well studied and several S-allele specific style glycoproteins identified2–6. In addition, complementary DNAs for nine style S-glycoproteins have been sequenced7–10 and have homology with two fungal ribonucleases. Recently five Nicotiana alata S-glycoproteins were shown to be RNases (S-RNases)11. We now report that S-allele specific degradation of pollen RNA occurs in vivo. After incompatible, but not after compatible pollinations, pollen RNA becomes degraded. This specificity cannot be demonstrated in vitro using isolated S-RNases and pollen RNAs. Our results support a model in which the gametophytic self-incompatibility system in N. alata acts through a cytotoxic mechanism directed against pollen RNA.

Self-incompatibility in Nicotiana alata involves degradation of pollen rRNA

Pollen-pistil interactions are crucial for controlling plant mating. For example, S-RNase-based self-incompatibility prevents inbreeding in diverse angiosperm species. S-RNases are thought to function as specific cytotoxins that inhibit pollen that has an S-haplotype that matches one of those in the pistil. Thus, pollen and pistil factors interact to prevent mating between closely related individuals. Other pistil factors, such as HT-B, 4936-factor and the 120 kDa glycoprotein, are also required for pollen rejection but do not contribute to S-haplotype-specificity per se. Here we show that S-RNase is taken up and sorted to a vacuolar compartment in the pollen tubes. Antibodies to the 120 kDa glycoprotein label the compartment membrane. When the pistil does not express HT-B or 4936-factor, S-RNase remains sequestered, unable to cause rejection. Similarly, in wild-type pistils, compatible pollen tubes degrade HT-B and sequester S-RNase. We suggest that S-RNase trafficking and the stability of HT-B are central to S-specific pollen rejection.

Compartmentalization of S-RNase and HT-B degradation in self-incompatible Nicotiana

We have characterized a soybean gene cluster that encodes a group of auxin-regulated RNAs (small auxin up RNAs). DNA sequencing of a portion of the locus reveals five homologous genes, spaced at intervals of about 1.25 kilobases and transcribed in alternate directions. At least three of the genes are transcriptionally regulated by auxin. An increase in the rate of transcription is detected 10 min after application of auxin to soybean elongating hypocotyl sections. Each of the genes contains an open reading frame that could encode a protein of 9 kilodaltons to 10.5 kilodaltons. Sequence comparisons among the five genes reveal several areas of high homology. Two regions of high homology begin about 250 base pairs upstream of the open reading frames and two regions of homology have been identified in sequences downstream of the open reading frames. One of the latter sequences occurs in the 39-untranslated region of the RNAs. The other occurs far downstream, 618 base pairs to 741 base pairs from the stop codon. Conservation of these sequences among the five different genes suggests that they may be important for the regulation of expression of the genes.

Bruce McClure

Papers

Style self-incompatibility gene products of Nicotlana alata are ribonucleases

S-RNase expressed in transgenic Nicotiana causes S-allele-specific pollen rejection

Self-incompatibility in Nicotiana alata involves degradation of pollen rRNA

Compartmentalization of S-RNase and HT-B degradation in self-incompatible Nicotiana

Transcription, organization, and sequence of an auxin-regulated gene cluster in soybean.