Plant-insect interactions.

Recent research shows partially overlapping signal transduction pathways controlling responses to wounding, insects, and pathogens. Chemical and behavioral assays show that plants release herbivore-specific volatiles, and that parasitic wasps can distinguish between these emission patterns. QTL mapping and candidate gene studies are beginning to identify polymorphic resistance genes, and ecological analyses provide information on the physiological and fitness costs of resistance. Such multidisciplinary approaches can elucidate the physiological causes and ecological consequences of plant-herbivore interactions.

Opposite Regulation of the Copy Number and the Expression of Plastid and Mitochondrial Genes by Light and Acetate in the Green Flagellate Chlorogonium.

In the unicellular green alga Chlorogonium elongatum (Chlamydomonadaceae), the formation of both the photosynthetic and the respiratory apparatus is under the control of light and acetate. Autotrophically cultured cells possess a 3-fold higher copy number of the plastid genes rbcL and psbA than cells cultivated in the dark with acetate (heterotrophic cells). Under mixotrophic conditions (light and acetate), both genes are present at an intermediate level. This pattern is repeated at the mRNA level. The amounts of rbcL and psbA mRNAs are approximately 3-fold higher in autotrophic cells than in heterotrophic ones and are intermediate in mixotrophic cells. As expected, the copy number of the nuclear-encoded rbcS gene is constant irrespective of the applied culture conditions. RbcS mRNA, however, is 7-fold more frequent in autotrophic than in heterotrophic cells. Again, mixotrophic cells show an intermediate level. In contrast to genes encoding plastid proteins, the copy number and transcript level of the mitochondrial cob gene are approximately 5-fold higher in heterotrophic cells than in autotrophic ones. As before, mixotrophic cells take an intermediate position. Therefore, light and acetate control the genes involved in the formation of either the photosynthetic or the respiratory apparatus in a coordinated but opposite manner.

The apocytochrome-b gene in Chlorogonium elongatum (Chlamydomonadaceae): an intronic GIY-YIG ORF in green algal mitochondria.

The mitochondrial cob gene from the green alga Chlorogonium elongatum (Chlamydomonadaceae) is interrupted by two group-I introns each containing an open reading frame in-phase with the upstream exon. One of these ORFs belongs to the LAGLI-DADG family, the other to the GIY-YIG family. The latter has not yet been identified in any mitochondrial genome except those from fungi. The Chlorogonium ORFs are similar to ORFs encoded by fungal introns that are located at an identical position within the gene, and to the ORF encoded by the mobile intron in the Chlamydomonas smithii cob gene.

The mitochondrial genome of Chlorogonium elongatum inferred from the complete sequence.

The 22,704-bp circular mitochondrial DNA (mtDNA) of the chlamydomonad alga Chlorogonium elongatum was completely cloned and sequenced. The genome encodes seven proteins of the respiratory electron transport chain, subunit 1 of the cytochrome oxidase complex (cox1), apocytochrome b (cob), five subunits of the NADH dehydrogenase complex (nad1, nad2, nad4, nad5, and nad6), a set of three tRNAs (Q, W, M), and the large (LSU)- and small (SSU)-subunit ribosomal RNAs. Six group-I introns were found, two each in the cox1, cob, and nad5 genes. In each intron an open reading frame (ORF) related to maturases or endonucleases was identified. Both the LSU and the SSU rRNA genes are split into fragments intermingled with each other and with other genes. Although the average A + T content is 62.2%, GC-rich clusters were detected in intergenic regions, in variable domains of the rRNA genes, and in introns and intron-encoded ORFs. A comparison of the genome maps reveals that C. elongatum and Chlamydomonas eugametos mtDNAs are more closely related to one another than either is to Chlamydomonas reinhardtii mtDNA.

Comparative genomics and regulatory evolution: conservation and function of the Chs and Apetala3 promoters.

DNA sequence variations of chalcone synthase (Chs) and Apetala3 gene promoters from 22 cruciferous plant species were analyzed to identify putative conserved regulatory elements. Our comparative approach confirmed the existence of numerous conserved sequences which may act as regulatory elements in both investigated promoters. To confirm the correct identification of a well-conserved UV-light-responsive promoter region, a subset of Chs promoter fragments were tested in Arabidopsis thaliana protoplasts. All promoters displayed similar light responsivenesses, indicating the general functional relevance of the conserved regulatory element. In addition to known regulatory elements, other highly conserved regions were detected which are likely to be of functional importance. Phylogenetic trees based on DNA sequences from both promoters (gene trees) were compared with the hypothesized phylogenetic relationships (species trees) of these taxa. The data derived from both promoter sequences were congruent with the phylogenies obtained from coding regions of other nuclear genes and from chloroplast DNA sequences. This indicates that promoter sequence evolution generally is reflective of species phylogeny. Our study also demonstrates the great value of comparative genomics and phylogenetics as a basis for functional analysis of promoter action and gene regulation.

Genetic control of natural variation in Arabidopsis glucosinolate accumulation.

Glucosinolates are biologically active secondary metabolites of the Brassicaceae and related plant families that influence plant/insect interactions. Specific glucosinolates can act as feeding deterrents or stimulants, depending upon the insect species. Hence, natural selection might favor the presence of diverse glucosinolate profiles within a given species. We determined quantitative and qualitative variation in glucosinolates in the leaves and seeds of 39 Arabidopsis ecotypes. We identified 34 different glucosinolates, of which the majority are chain-elongated compounds derived from methionine. Polymorphism at only five loci was sufficient to generate 14 qualitatitvely different leaf glucosinolate profiles. Thus, there appears to be a modular genetic system regulating glucosinolate profiles in Arabidopsis. This system allows the rapid generation of new glucosinolate combinations in response to changing herbivory or other selective pressures. In addition to the qualitative variation in glucosinolate profiles, we found a nearly 20-fold difference in the quantity of total aliphatic glucosinolates and were able to identify a single locus that controls nearly three-quarters of this variation.

A gene controlling variation in Arabidopsis glucosinolate composition is part of the methionine chain elongation pathway.

Arabidopsis and other Brassicaceae produce an enormous diversity of aliphatic glucosinolates, a group of methionine (Met)-derived plant secondary compounds containing a beta-thio-glucose moiety, a sulfonated oxime, and a variable side chain. We fine-scale mapped GSL-ELONG, a locus controlling variation in the side-chain length of aliphatic glucosinolates. Within this locus, a polymorphic gene was identified that determines whether Met is extended predominantly by either one or by two methylene groups to produce aliphatic glucosinolates with either three- or four-carbon side chains. Two allelic mutants deficient in four-carbon side-chain glucosinolates were shown to contain independent missense mutations within this gene. In cell-free enzyme assays, a heterologously expressed cDNA from this locus was capable of condensing 2-oxo-4-methylthiobutanoic acid with acetyl-coenzyme A, the initial reaction in Met chain elongation. The gene methylthioalkylmalate synthase1 (MAM1) is a member of a gene family sharing approximately 60% amino acid sequence similarity with 2-isopropylmalate synthase, an enzyme of leucine biosynthesis that condenses 2-oxo-3-methylbutanoate with acetyl-coenzyme A.

Induced plant defense responses against chewing insects. Ethylene signaling reduces resistance of Arabidopsis against Egyptian cotton worm but not diamondback moth.

The induction of plant defenses by insect feeding is regulated via multiple signaling cascades. One of them, ethylene signaling, increases susceptibility of Arabidopsis to the generalist herbivore Egyptian cotton worm (Spodoptera littoralis; Lepidoptera: Noctuidae). The hookless1 mutation, which affects a downstream component of ethylene signaling, conferred resistance to Egyptian cotton worm as compared with wild-type plants. Likewise, ein2, a mutant in a central component of the ethylene signaling pathway, caused enhanced resistance to Egyptian cotton worm that was similar in magnitude to hookless1. Moreover, pretreatment of plants with ethephon (2-chloroethanephosphonic acid), a chemical that releases ethylene, elevated plant susceptibility to Egyptian cotton worm. By contrast, these mutations in the ethylene-signaling pathway had no detectable effects on diamondback moth (Plutella xylostella) feeding. It is surprising that this is not due to nonactivation of defense signaling, because diamondback moth does induce genes that relate to wound-response pathways. Of these wound-related genes, jasmonic acid regulates a novel beta-glucosidase 1 (BGL1), whereas ethylene controls a putative calcium-binding elongation factor hand protein. These results suggest that a specialist insect herbivore triggers general wound-response pathways in Arabidopsis but, unlike a generalist herbivore, does not react to ethylene-mediated physiological changes.