The first enzyme of the phenylpropanoid pathway, Phe ammonia-lyase (PAL), is encoded by four genes in Arabidopsis thaliana. Whereas PAL function is well established in various plants, an insight into the functional significance of individual gene family members is lacking. We show that in the absence of clear phenotypic alterations in the Arabidopsis pal1 and pal2 single mutants and with limited phenotypic alterations in the pal1 pal2 double mutant, significant modifications occur in the transcriptome and metabolome of the pal mutants. The disruption of PAL led to transcriptomic adaptation of components of the phenylpropanoid biosynthesis, carbohydrate metabolism, and amino acid metabolism, revealing complex interactions at the level of gene expression between these pathways. Corresponding biochemical changes included a decrease in the three major flavonol glycosides, glycosylated vanillic acid, scopolin, and two novel feruloyl malates coupled to coniferyl alcohol. Moreover, Phe overaccumulated in the double mutant, and the levels of many other amino acids were significantly imbalanced. The lignin content was significantly reduced, and the syringyl/guaiacyl ratio of lignin monomers had increased. Together, from the molecular phenotype, common and specific functions of PAL1 and PAL2 are delineated, and PAL1 is qualified as being more important for the generation of phenylpropanoids.

Upon the incidence of DNA stress, the ataxia telangiectasia-mutated (ATM) and Rad3-related (ATR) signaling kinases activate a transient cell cycle arrest that allows cells to repair DNA before proceeding into mitosis. Although the ATM-ATR pathway is highly conserved over species, the mechanisms by which plant cells stop their cell cycle in response to the loss of genome integrity are unclear. We demonstrate that the cell cycle regulatory WEE1 kinase gene of Arabidopsis thaliana is transcriptionally activated upon the cessation of DNA replication or DNA damage in an ATR-or ATM-dependent manner, respectively. In accordance with a role for WEE1 in DNA stress signaling, WEE1-deficient plants showed no obvious cell division or endoreduplication phenotype when grown under nonstress conditions but were hypersensitive to agents that impair DNA replication. Induced WEE1 expression inhibited plant growth by arresting dividing cells in the G2-phase of the cell cycle. We conclude that the plant WEE1 gene is not rate-limiting for cycle progression under normal growth conditions but is a critical target of the ATR-ATM signaling cascades that inhibit the cell cycle upon activation of the DNA integrity checkpoints, coupling mitosis to DNA repair in cells that suffer DNA damage.

Plastids are DNA-containing organelles unique to plant cells. In Arabidopsis, one-third of the genes required for embryo development encode plastid-localized proteins. To help understand the role of plastids in embryogenesis and postembryonic development, we characterized proteins of the mitochondrial transcription termination factor (mTERF) family, which in animal models, comprises DNA-binding regulators of mitochondrial transcription. Of 35 Arabidopsis mTERF proteins, 11 are plastid-localized. Genetic complementation shows that at least one plastidic mTERF, BELAYA SMERT' (BSM), is required for embryogenesis. The main postembryonic phenotypes of genetic mosaics with the bsm mutation are severe abnormalities in leaf development. Mutant bsm cells are albino, are compromised in growth, and suffer defects in global plastidic gene expression. The bsm phenotype could be phenocopied by inhibition of plastid translation with spectinomycin. Plastid translation is essential for cell viability in dicotyledonous species such as tobacco but not in monocotyledonous maize. Here, genetic interactions between BSM and the gene encoding plastid homomeric acetyl-CoA carboxylase ACC2 suggest that there is a functional redundancy in malonyl-CoA biosynthesis that permits bsm cell survival in Arabidopsis. Overall, our results indicate that biosynthesis of malonyl-CoA and plastid-derived systemic growth-promoting compounds are the processes that link plant development and plastid gene expression. organellar gene expression | splicing | ClpPR protease | myrosinase | digitonin

The 26S proteasome is an ATP-dependent eukaryotic protease responsible for degrading many important cell regulators, especially those conjugated with multiple ubiquitins. Bound on both ends of the 20S core protease is a multisubunit regulatory particle that plays a crucial role in substrate selection by an as yet unknown mechanism(s). Here, we show that the RPN12 subunit of the Arabidopsis regulatory particle is involved in cytokinin responses. A T-DNA insertion mutant that affects RPN12a has a decreased rate of leaf formation, reduced root elongation, delayed skotomorphogenesis, and altered growth responses to exogenous cytokinins, suggesting that the mutant has decreased sensitivity to the hormone. The cytokinin-inducible genes CYCD3 and NIA1 are upregulated constitutively in rpn12a-1 , indicating that feedback-inhibitory mechanisms also may be altered. rpn12a-1 seedlings also showed changes in auxin-induced growth responses, further illustrating the close interaction between auxin and cytokinin regulation. In yeast, RPN12 is necessary for the G1/S and G2/M transitions of the cell cycle, phases that have been shown to be under cytokinin control in plants. We propose that RPN12a is part of the Arabidopsis 26S proteasome that controls the stability of one or more of the factors involved in cytokinin regulation. INTRODUCTIONRegulated protein turnover provides a mechanism to adjust rapidly to changing ligand concentrations and/or environmental conditions and is essential for many signal response pathways. In eukaryotes, the ubiquitin/26S proteasome pathway is particularly important, being responsible for removing most short-lived intracellular proteins (Hershko and Ciechanover, 1998;Callis and Vierstra, 2000). In this proteolytic pathway, proteins committed for degradation first are modified by the covalent attachment of multiple ubiquitins. This conjugation is directed by an ATP-dependent reaction cascade involving the sequential action of E1s, E2s, and E3s, which ultimately attach one or more ubiquitins to appropriate targets. In most cases, the ubiquitinated proteins then are recognized and degraded by the 26S proteasome, a multisubunit ATP-dependent protease with broad substrate specificity.The 26S proteasome is a 2-MD complex assembled from two particles: the 20S core particle (CP) and the 19S regulatory particle (RP) (Voges et al., 1999). The proteolytic activities reside within the central chamber of the 28-subunit CP, whereas the functions that direct substrate recognition, unfolding, and subsequent entry into the 20S particle reside within the Ͼ 18-subunit RP. The RP can be divided further in two subcomplexes, the lid and base (Glickman et al., 1998). The base contains six ATPase subunits, RPT1 to RPT6, which presumably use ATP hydrolysis to unfold target proteins, and three non-ATPase subunits, RPN1, RPN2, and RPN10. The lid contains nine additional RPN subunits (RPN3, RPN5 to RPN9, and RPN11 to RPN13). Many of the lid RPN subunits share sequence motifs with components of the COP9/signalosome and EIF3 co...

The 26S proteasome is an essential protease complex responsible for removing most short-lived intracellular proteins, especially those modified with polyubiquitin chains. We show here that an Arabidopsis mutant expressing an altered RPN10 subunit exhibited a pleiotropic phenotype consistent with specific changes in 26S proteasome function. rpn10-1 plants displayed reduced seed germination, growth rate, stamen number, genetic transmission through the male gamete, and hormone-induced cell division, which can be explained partially by a constitutive downregulation of the key cell cycle gene CDKA;1 . rpn10-1 also was more sensitive to abscisic acid (ABA), salt, and sucrose stress and to DNA-damaging agents and had decreased sensitivity to cytokinin and auxin. Most of the phenotypes can be explained by a hypersensitivity to ABA, which is reflected at the molecular level by the selective stabilization of the short-lived ABA-signaling protein ABI5. Collectively, these results indicate that RPN10 affects a number of regulatory processes in Arabidopsis likely by directing specific proteins to the 26S proteasome for degradation. A particularly important role may be in regulating the responses to signals promulgated by ABA.

AtATM3, an ATP-binding cassette transporter of Arabidopsis (Arabidopsis thaliana), is a mitochondrial protein involved in the biogenesis of iron-sulfur clusters and iron homeostasis in plants. Our gene expression analysis showed that AtATM3 is upregulated in roots of plants treated with cadmium [Cd(II)] or lead (II); hence, we investigated whether this gene is involved in heavy metal tolerance. We found that AtATM3-overexpressing plants were enhanced in resistance to Cd, whereas atatm3 mutant plants were more sensitive to Cd than their wild-type controls. Moreover, atatm3 mutant plants expressing 35S promoter-driven AtATM3 were more resistant to Cd than wild-type plants. Since previous reports often showed that the cytosolic glutathione level is positively correlated with heavy metal resistance, we measured nonprotein thiols (NPSH) in these mutant plants. Surprisingly, we found that atatm3 contained more NPSH than the wild type under normal conditions. AtATM3-overexpressing plants did not differ under normal conditions, but contained less NPSH than wild-type plants when exposed to Cd(II). These results suggest a role for AtATM3 in regulating cellular NPSH level, a hypothesis that was further supported by our gene expression study. Genetic or pharmacological inhibition of glutathione biosynthesis led to the elevated expression of AtATM3, whereas expression of the glutathione synthase gene GSH1 was increased under Cd(II) stress and in the atatm3 mutant. Because the closest homolog of AtATM3 in fission yeast (Schizosaccharomyces pombe), HMT1, is a vacuolar membrane-localized phytochelatin-Cd transporter, it is tempting to speculate that glutathione-Cd(II) complexes formed in the mitochondria are exported by AtATM3. In conclusion, our data show that AtATM3 contributes to Cd resistance and suggest that it may mediate transport of glutamine synthetaseconjugated Cd(II) across the mitochondrial membrane.

African oil palm has the highest productivity amongst cultivated oleaginous crops. Species can constitute a single crop capable to fulfill the growing global demand for vegetable oils, which is estimated to reach 240 million tons by 2050. Two types of vegetable oil are extracted from the palm fruit on commercial scale. The crude palm oil and kernel palm oil have different fatty acid profiles, which increases versatility of the crop in industrial applications. Plantations of the current varieties have economic life-span around 25–30 years and produce fruits around the year. Thus, predictable annual palm oil supply enables marketing plans and adjustments in line with the economic forecasts. Oil palm cultivation is one of the most profitable land uses in the humid tropics. Oil palm fruits are the richest plant source of pro-vitamin A and vitamin E. Hence, crop both alleviates poverty, and could provide a simple practical solution to eliminate global pro-vitamin A deficiency. Oil palm is a perennial, evergreen tree adapted to cultivation in biodiversity rich equatorial land areas. The growing demand for the palm oil threatens the future of the rain forests and has a large negative impact on biodiversity. Plant science faces three major challenges to make oil palm the key element of building the future sustainable world. The global average yield of 3.5 tons of oil per hectare (t) should be raised to the full yield potential estimated at 11–18t. The tree architecture must be changed to lower labor intensity and improve mechanization of the harvest. Oil composition should be tailored to the evolving needs of the food, oleochemical and fuel industries. The release of the oil palm reference genome sequence in 2013 was the key step toward this goal. The molecular bases of agronomically important traits can be and are beginning to be understood at the single base pair resolution, enabling gene-centered breeding and engineering of this remarkable crop.

A chimeric construct containing the Nicotiana plumbaginifolia beta‐1,3‐glucanase gn1 gene was introduced into Nicotiana tabacum SR1 to produce high levels of the enzyme constitutively. We determined that the GN1 protein represents a basic beta‐1,3‐glucanase isoform which accumulates into the vacuoles of the transgenic plants. Analysis of the progeny of the transgenic plant with the highest levels of gn1 expression revealed an unexpected phenomenon of gene suppression. Plants hemizygous for the T‐DNA locus contained high levels of gn1 mRNA and exhibited a 14‐fold higher beta‐1,3‐glucanase activity than untransformed plants. However, the expression of gn1 was completely suppressed in the homozygous plants: no corresponding mRNA or protein could be detected. This suppression mechanism occurs at a post‐transcriptional level and is under developmental control. In addition, by generating haploid plants we found that this silencing phenomenon is not dependent on allelic interaction between T‐DNA copies present at the same locus of homologous chromosomes, but rather is correlated with the transgene dose in the plant genome. We postulate that high doses of GN1 protein relative to the level(s) of other still unknown plant products could trigger the cellular processes directed to suppress gn1 expression.