Sporopollenin monomer biosynthesis in arabidopsis

Land plants have evolved aliphatic biopolymers that protect their cell surfaces against dehydration, pathogens, and chemical and physical damage. In flowering plants, a critical event during pollen maturation is the formation of the pollen surface structure. The pollen wall consists essentially of the microspore-derived intine and the sporophyte-derived exine. The major component of the exine is termed sporopollenin, a complex biopolymer. The chemical composition of sporopollenin remains poorlycharacterized because it is extremely resistant to chemical and biological degradation procedures. Recent characterization of Arabidopsis thaliana genes and corresponding enzymes involved in exine formation has demonstrated that the sporopollenin polymer consists of phenolic and fatty acid-derived constituents that are covalently coupled by ether and ester linkages. This review illuminates the outlines of a biosynthetic pathway involved in generating monomer constituents of the sporopollenin biopolymer component of the pollen wall.     

Differential inhibition of indole-3-acetic acid and tryptophan biosynthesis by indole analogues. I. Tryptophan dependent IAA biosynthesis

Maize liquid endosperm extracts contain the enzymes necessary for all of the steps of the plant IAA biosynthetic pathway from tryptophan, and provide a means to assay the pathway in vitro. We have analyzed the reactions in the presence of a series of indole and indole-like analogues in order to evaluate the potential of these compounds to act as inhibitors of IAA biosynthesis. Such inhibitors will be useful to investigate the tryptophan to IAA pathway, to determine the precursors and intermediates involved, and to select for mutants in this process. A number of such compounds were tested using in vitro enzyme assays for both the tryptophan dependent IAA biosynthesis pathway and for tryptophan synthase activity. Some compounds showed strong inhibition of IAA biosynthesis while having only a slight effect on the reaction rate of tryptophan synthase . These results: (1) show that IAA biosynthesis can be selectively inhibited relative to tryptophan biosynthesis; (2) suggest potential ways to screen for IAA biosynthetic pathway mutations in plants; and (3) provide additional tools for studies of IAA biosynthesis in plants.     

Novel seed lipid phenotypes in combinations of mutants altered in fatty acid biosynthesis inArabidopsis

Summary The seed fatty acid (FA) composition of various single mutant combinations ofArabidopsis thaliana that affect FA biosynthesis has been examined. Double mutant combinations offae, a mutation affecting CIS elongation, and a series of four other FA biosynthetic mutants were synthesized. The four other single mutants were:fad2 andfad3, which are deficient in 181 and 182 desaturation, respectively;fab1, which is elevated in 160 and decreased in 181; andfab2, which is elevated in 180 and decreased in 181. The superimposition of two blocks in the FA biosynthetic pathway leads to dramatic changes in the FA content of the double mutants. The tenArabidopsis stocks analyzed to date (wild-type, five single mutants, and four double mutants) make seed oils with a wide range of FA compositions, and illustrate the diversity of oils it is possible to obtain from a single plant species.     

Quantitative analysis of IAA in DR5::GUS transgenic arabidopsis plants

In the study of auxin transport, transgenic constructs, including DR5::GUS, are widely used for visualization of phytohormone localization. Previously we proposed a method for quantitative evaluation of the IAA content by histochemical staining for glucuronidase activity. In this work, this method was complemented by quantitative data on the content of IAA in plants obtained by gas chromatography-mass spectrometry (GC/MS), which allowed more accurate characterization of the lateral IAA gradient arising at the Arabidopsis thaliana (L.) Heynh (ecotype Columbia 0) root gravistimulation. Applied method of IAA analysis, combining GC/MS and histochemistry, can be used for quantitatification of the other plant hormone distribution in transgenic plants with the GUS reporter.     

Tryptophan biosynthesis in the marine luminous bacterium Vibrio harveyi.

Tryptophan biosynthetic enzyme levels in wild-type Vibrio harveyi and a number of tryptophan auxotrophs of this species were coordinately regulated over a 100-fold range of specific activities. The tryptophan analog indoleacrylic acid evoked substantial derepression of the enzymes in wild-type cells. Even higher enzyme levels were attained in auxotrophs starved for tryptophan, regardless of the location of the block in the pathway. A derepressed mutant selected by resistance to 5-fluorotryptophan was found to have elevated basal levels of trp gene expression; these basal levels were increased only two- to threefold by tryptophan limitation. The taxonomic implications of these and other biochemical results support previous suggestions that the marine luminous bacteria are more closely related to enteric bacteria than to other gram-negative taxa.     

色氨酸, 天然产物生物合成中的重要结构单元

氨基酸作为生物体内组成生命物质的小分子化合物, 在天然产物生物合成中扮演了非常重要的作用。色氨酸含有一个独特的吲哚环, 相对复杂的吲哚环平面结构使得色氨酸相比其他氨基酸具有更多的修饰空间。在微生物天然产物生物合成研究中, 色氨酸及其衍生物经常作为组成模块参与到天然产物的生物合成中, 本文概述了色氨酸几种不同的生物修饰方式, 包括烷基化修饰、卤化修饰、羟基化修饰、以及吲哚环的开环重排反应等。分析并总结色氨酸在天然产物生物合成中的作用可以增加我们对天然产物结构多样性的认识和推动天然产物生物合成机制的研究。     

Differential IAA dose response relations of the axr1 and axr2 mutants of Arabidopsis

In comparison to wild type Arabidopsis thaliana, the auxin resistant mutants axr1 and axr2 exhibit reduced inhibition of root elongation in response to auxins. Several auxin-regulated physiological processes are also altered in the mutant plants. When wild-type, axr1 and axr2 seedlings were grown in darkness on media containing indoleacetic acid (IAA), promotion of root growth was observed at low concentrations of IAA (10^?11 to 10^?7M) in 5-day-old axr2 seedlings, but not in axr1 or wild-type seedlings. In axr1 there was little or no measurable root growth response over the same concentration range. In wild type, root growth was inhibited at concentrations greater than 10^?10M and no detectable root growth response was observed at lower concentrations. In addition, production of lateral roots in response to IAA increased in axr2 seedlings and decreased in axr1 seedlings relative to wild type. Promotion of root elongation and initiation of lateral roots in axr2 seedlings in response to auxin indicate that axr2 seedlings are able to perceive and respond to IAA.     

Tryptophan biosynthesis genes trpEGC in the thermoacidophilic archaebacterium Sulfolobus solfataricus.

A DNA fragment containing the trpEGC gene cluster was isolated from the thermoacidophilic archaebacterium Sulfolobus solfataricus. The products of trpE, trpG, and trpC from S. solfataricus were compared to the homologous products from a eukaryote, a eubacterium, and two archaebacteria, namely, a methanogen and an extreme halophile. They appeared to be equally related to the proteins from Escherichia coli and Saccharomyces cerevisiae, the percentages of conserved amino acids being roughly the same as those measured when comparing the eubacterial and eukaryotic sequences directly. These percentages did not rise significantly when a comparison with the proteins from Haloferax volcanii was drawn, while a slightly closer relationship with the proteins from Methanococcus thermoautotrophicum was found.     

Isolation and characterization of mutants blocked in T-2 toxin biosynthesis

Mutants of Fusarium sporotrichioides NRRL 3299 that were blocked or altered in the biosynthesis of the trichothecene T-2 toxin were generated by UV treatment and identified by a rapid screen in which monoclonal antibodies to T-2 were used. Three stable mutants were isolated and chemically characterized. Two mutants accumulated diacetoxyscirpenol, which suggests that they were defective in the step required for the addition of a hydroxyl group to the C-8 position in the trichothecene core structure. The third mutant appeared to be partially blocked at an early step or regulatory point in the pathway. This represents the first isolation of mutants in a trichothecene biosynthetic pathway.     

Arabidopsis sfd mutants affect plastidic lipid composition and suppress dwarfing, cell death, and the enhanced disease resistance phenotypes resulting from the deficiency of a fatty acid desaturase

A loss-of-function mutation in the Arabidopsis SSI2/FAB2 gene, which encodes a plastidic stearoyl-acyl-carrier protein desaturase, has pleiotropic effects. The ssi2 mutant plant is dwarf, spontaneously develops lesions containing dead cells, accumulates increased salicylic acid (SA) levels, and constitutively expresses SA-mediated, NPR1-dependent and -independent defense responses. In parallel, jasmonic acid-regulated signaling is compromised in the ssi2 mutant. In an effort to discern the involvement of lipids in the ssi2-conferred developmental and defense phenotypes, we identified suppressors of fatty acid (stearoyl) desaturase deficiency (sfd) mutants. The sfd1, sfd2, and sfd4 mutant alleles suppress the ssi2-conferred dwarfing and lesion development, the NPR1-independent expression of the PATHOGENESIS-RELATED1 (PR1) gene, and resistance to Pseudomonas syringae pv maculicola. The sfd1 and sfd4 mutant alleles also depress ssi2-conferred PR1 expression in NPR1-containing sfd1 ssi2 and sfd4 ssi2 plants. By contrast, the sfd2 ssi2 plant retains the ssi2-conferred high-level expression of PR1. In parallel with the loss of ssi2-conferred constitutive SA signaling, the ability of jasmonic acid to activate PDF1.2 expression is reinstated in the sfd1 ssi2 npr1 plant. sfd4 is a mutation in the FAD6 gene that encodes a plastidic omega6-desaturase that is involved in the synthesis of polyunsaturated fatty acid-containing lipids. Because the levels of plastid complex lipid species containing hexadecatrienoic acid are depressed in all of the sfd ssi2 npr1 plants, we propose that these lipids are involved in the manifestation of the ssi2-conferred phenotypes.     

Detection of the IAA Biosynthetic Pathway from Tryptophan via Indole-3-Acetamide in Bradyrhizobium spp.

The IAA biosynthetic pathway of tryptophan to IAA via IAM wasdetected in Bradyrhizobium spp. (slow-growing Rhizobium) butnot in Rhizobium spp. (fast-growing Rhizobium). A simple methodusing rapid HPLC analysis to measure the conversion from NAMto NAA was developed to detect indole-3-acetamide hydrolaseactivity in cultures of bacteria. Most of the Bradyrhizobiumstrains produce large amounts of NAA converted from NAM underour assay conditions. In addition, GC/MS analysis of purifiedextracts from cultures of B.japonicum wild-type strain J1063,grown in a tryptophan-supplemented liquid medium, demonstratedthe presence of IAM and IAA. The results strongly suggest thatbiosynthesis of IAA in Bradyrhizobium spp. involves the samepathway as that operating in Pseudomonas savastanoi and Agrobacteriumtumefaciens. (Received December 25, 1988; Accepted May 18, 1988)     

Melanotic mutants in Drosophila: pathways and phenotypes

Mutations in >30 genes that regulate different pathways and developmental processes are reported to cause a melanotic phenotype in larvae. The observed melanotic masses were generally linked to the hemocyte-mediated immune response. To investigate whether all black masses are associated with the cellular immune response, we characterized melanotic masses from mutants in 14 genes. We found that the melanotic masses can be subdivided into melanotic nodules engaging the hemocyte-mediated encapsulation and into melanizations that are not encapsulated by hemocytes. With rare exception, the encapsulation is carried out by lamellocytes. Encapsulated nodules are found in the hemocoel or in association with the lymph gland, while melanizations are located in the gut, salivary gland, and tracheae. In cactus mutants we found an additional kind of melanized mass containing various tissues. The development of these tissue agglomerates is dependent on the function of the dorsal gene. Our results show that the phenotype of each mutant not only reflects its connection to a particular genetic pathway but also points to the tissue-specific role of the individual gene.     

Overexpression of the non-canonical Aux/IAA genes causes auxin-related aberrant phenotypes in Arabidopsis

Degradation of Aux/IAA proteins which are triggered by the ubiquitin ligase complex containing the auxin F-box receptors (AFBs), is thought to be the primary reaction of auxin signaling. Upon auxin perception, AFBs bind domain II of Aux/IAA proteins that is conserved in most of the 29 family members in Arabidopsis. However, IAA20 and IAA30 lack domain II. Furthermore, IAA31, which forms a single clade with IAA20 and IAA30 in Aux/IAA protein family, has a partially conserved domain II, which contains an amino acid substitution that would cause a dominant mutation of Aux/IAA genes. It has been shown that the half-lives of these proteins are much longer than those of the canonical Aux/IAA proteins. We generated overexpression lines (OXs) of IAA20 , IAA30 and IAA31 by the use of cauliflower mosaic virus 35S promoter to better understand the molecular function of atypical Aux/IAA proteins in Arabidopsis. OXs of the three genes exhibited similar auxin-related aberrant phenotypes, with IAA20 OX showing the most severe defects: Some of them showed a semi-dwarf phenotype; gravitropic growth orientation was often affected in hypocotyl and root; vasculature of cotyledons was malformed; the primary root stopped growing soon after germination because of collapse of root apical meristem. IAA 20 and IAA30 were early auxin inducible, but IAA31 was not. These results showed that the wild-type genes of the three Aux/IAAs could disturb auxin physiology when ectopically overexpressed.