- © 2008 American Society of Plant Biologists
Sex Differences in Plastid DNA Integration into the Nucleus
The plastid (chloroplast) genome of higher plants has been reduced to approximately 130 genes, while its cyanobacterial ancestor is estimated to have contained more than 3,000 genes. Thus, many of the ancestral endosymbiont genes were either lost or transferred to the nucleus during evolution. Indeed, thousands of functional nuclear genes in Arabidopsis (Arabidopsis thaliana) are derived from the endosymbiont genome. Sheppard et al. (pp. 328–336) examine the question of whether there are sex-specific differences in the rates of plastid DNA relocation. Male gametes of most angiosperm species undergo a programmed elimination of plastids during pollen development, a process that underpins maternal inheritance of organellar genes. The authors have monitored the cellular location of a kanamycin resistance gene tailored for nuclear expression (35SneoSTLS2) in the progeny of reciprocal crosses of tobacco (Nicotiana tabacum) in which, at the start of the experiments, the reporter gene was confined either to the male or the female parental plastid genome. Among 146,000 progeny from crosses where the transplastomic parent was male, 13 transposition events were identified, whereas only one atypical transposition was identified in a screen of 273,000 transplastomic ovules. Thus, the transfer of plastid DNA to the nucleus takes place far more frequently in the male germline than in the female germline. Some plant biotechnologists have advocated the placement of transgenes in the plastid genome to prevent transgene escape through pollen dispersal. This study demonstrates that the frequency of plastid transgene relocation to the nucleus in the male germline is at least an order of magnitude higher than in the female germline. Hence, plastid transgenesis alone does not provide complete transgene containment in tobacco, and additional safeguards may be necessary to eliminate all possibility of transgene escape.
Protein Farnesylation and Plant Innate Immunity
Pathogen-specific defense is mediated by Resistance (R) proteins, which recognize pathogenic effector molecules. The activation of R proteins initiates signaling events that usually culminate in programmed death of cells at the site of infection and containment of the invading pathogen (the hypersensitive response). Most known R proteins belong to the NBS-LRR class, with carboxyl-terminal Leu-rich repeats (LRRs) and a central nucleotide binding site (NBS) domain. Although many R genes have been cloned, the signaling events downstream of R-protein activation remain elusive. The proper localization of defense signaling components and their interaction with other proteins are imperative for successful defense responses, and these often depend on posttranslational modifications. To search for additional components required for R-protein signaling, Goritschnig et al. (pp. 348–357) took advantage of the plant autoimmune model suppressor of npr1 constitutive1 (snc1), a gain-of-function allele of an NBS-LRR R gene. In addition to having constitutive resistance against virulent bacterial and oomycete pathogens, the snc1 mutant also displays increased levels of endogenous salicylic acid and constitutive expression of PR genes. In a screen for modifiers of snc1 (MOS), the authors have identified mos8, a genetic suppressor of snc1. Interestingly, mos8 turns out to be a novel allele of ERA1 (for ENHANCED RESPONSE TO ABSCISIC ACID1), which encodes the protein farnesyltransferase β-subunit. Protein farnesylation involves attachment of C15-prenyl residues to the carboxyl termini of specific target proteins, and has been shown to be important in development and hormonal responses. mos8 affects basal resistance against virulent pathogens as well as some R-protein-mediated resistance responses. Thus, the farnesylation of proteins in response to biotic stresses adds another layer of complexity to the signaling network of plant innate immunity.
Viroids and RNA Silencing
Self-replicating RNAs known as viroids are the smallest (only 246 to 401 nucleotides long!) infectious agents in plants. The mechanism(s) by which these pathogenic RNAs interact with hosts to induce disease symptoms is uncertain. The discovery of small RNAs that regulate host and nonhost gene expression in eukaryotes has led to a new hypothesis of viroid pathogenesis that is based on RNA silencing. RNA silencing is a sequence-specific RNA-inactivation mechanism in eukaryotes that guides chromatin modification and gene regulation. Many small RNA pathways in plants have a requirement for RDR6, an RNA-dependent RNA polymerase. Gómez et al. (pp. 414–423) reasoned that if the symptoms following viroid infection are mediated by RNA silencing, then the following predictions should be fulfilled: (1) the inhibition of the RNA silencing activity should be associated with a decrease in the severity of symptoms, and (2) infected plants with deficiencies in the RNA silencing pathway would either be asymptomatic or show a reduction in the severity of symptoms. In an attempt to validate these predictions, they used a symptomatic transgenic line of tobacco (Nicotiana benthamiana) that expresses and processes dimeric forms of Hop stunt viroid (HSVd) into the biologically active monomeric circular and linear forms. The HSVd-expressing tobacco plants were used as stocks in grafting assays with an rdr6i line, in which RDR6 is constitutively silenced. The authors demonstrate that the expression of symptoms in tobacco plants is independent of HSVd accumulation levels, but dependent on an active state of the viroid-specific RNA silencing pathway. The scion of rdr6i plants remained asymptomatic when grafted onto symptomatic plants, despite an accumulation of a high level of mature forms of HSVd, indicating the requirement of RDR6 for the viroid-induced symptom production. Thus, the symptoms induced by HSVd infection in tobacco are dependent on RDR6 activity, a key component of diverse RNA silencing pathways, and their severity is associated with efficient viroid-specific RNA silencing activity.
Proteomics of Seed Aging
Seed aging refers to the progressive loss of germination vigor over time. Seed aging is of both ecological and agricultural interest, and is especially troublesome to programs devoted to the preservation of germplasm. Previous genetic studies have revealed that seed longevity is a multigenic trait. In this issue, Rajjou et al. (pp. 620–641) relate the results of proteomic studies aimed at elucidating the molecular biology underlying seed aging in Arabidopsis. The authors used a hydration/dehydration protocol known as controlled deterioration that is widely believed to mimic natural aging. Germination tests showed a progressive decrease of germination vigor depending on the duration of controlled deterioration. Proteomic analyses revealed that seed deterioration was accompanied by a massive increase in carbonylated proteins. There were also decreases in dehydrins and other desiccation-associated proteins as well as increases in certain glycolytic enzymes. The levels of β-mercaptopyruvate sulfurtransferase, an enzyme that functions in cyanide detoxification, also decreased. This finding suggests a possible role for sulfur metabolism in the maintenance of seed vigor. The authors conclude that that the essential mechanisms for seed vigor include translational capacity, the ability to mobilize seed storage reserves in a controlled manner, and detoxification efficiency.
Leaf Development in a C4 Species Lacking Kranz Anatomy
C4 photosynthesis was once thought to be inextractably linked to Kranz anatomy, a morphology in which leaf chlorenchyma is separated into mesophyll cells containing phosphoenolpyruvate carboxylase (PEPC) and into bundle sheath cells containing Rubisco. This concept was turned on its head when three succulent species in the family Chenopodiaceae, including Bienertia sinuspersici, were shown to have a unique photosynthetic mechanism in which C4 photosynthesis occurs within individual photosynthetic cells. This single-celled C4 photosynthesis is possible because B. sinuspersici develops two cytoplasmic compartments in its chlorenchyma cells: a large central cytoplasmic compartment packed with chloroplasts and mitochondria, and a peripheral layer of cytoplasm with chloroplasts. The peripheral cytoplasm is analogous to the mesophyll cells of typical C4 plants. Atmospheric CO2, upon entry into chlorenchyma cells, is incorporated into the C4 acid oxaloacetate by PEPC in the peripheral cytoplasm. The central cytoplasmic compartment, on the other hand, is analogous to the bundle sheath cell of typical C4 plants. Rubisco, localized exclusively in the central cytoplasmic compartment, fixes the CO2 released by the organic acids produced by PEPC. The studies of Lara et al. (pp. 593–610) shed light on the timing and pattern of expression of photosynthetic genes in B. sinuspersici as well as the stability of its unusual cytoplasmic architecture. They report that Rubisco subunits, and enzymes of the glycolate pathway, accumulate more rapidly than enzymes associated with the C4 cycle, suggesting a progressive development of C4 photosynthesis. They also report that when branches containing mature leaves were enclosed in darkness for one month, the cytoplasmic spatial domains were maintained.
Insights into the Function of an Outer Chloroplast Membrane Protein
The Omp85 (Outer membrane protein, 85 kD) superfamily of β-barrel proteins is found in chloroplasts, mitochondria, and Gram-negative bacteria. The Omp85 proteins in bacteria and mitochondria mediate biogenesis of other β-barrel proteins, and are indispensable for viability. Chloroplasts contain at least two distinct types of Omp85 homologs, namely, Toc75 (Translocon at the outer envelope membrane of chloroplasts, 75 kD) and OEP80 (Outer Envelope Protein, 80 kD). Since both homologs exist in extant cyanobacteria, these proteins presumably arose from a common ancestor in the original endosymbiont. Indeed, detailed phylogenetic analyses suggested that Toc75 and OEP80 diverged early in the evolution of chloroplasts. Toc75 functions as a preprotein translocation channel during chloroplast import, but the role of OEP80 remains elusive. Patel et al. (pp. 235–245) demonstrate that AtOEP80 is essential for viability, and reveal that the N-terminal part of the protein is not required for its biogenesis or function.
