In order to prevent genome instability, cells need to be protected by a number of repair mechanisms, including DNA double‐strand break (DSB) repair. The extent to which DSB repair, biased towards deletions or insertions, contributes to evolutionary diversification of genome size is still under debate. We analyzed mutation spectra in Arabidopsis thaliana and in barley (Hordeum vulgare) by PacBio sequencing of three DSB‐targeted loci each, uncovering repair via gene conversion, single strand annealing (SSA) or nonhomologous end‐joining (NHEJ). Furthermore, phylogenomic comparisons between A. thaliana and two related species were used to detect naturally occurring deletions during Arabidopsis evolution. Arabidopsis thaliana revealed significantly more and larger deletions after DSB repair than barley, and barley displayed more and larger insertions. Arabidopsis displayed a clear net loss of DNA after DSB repair, mainly via SSA and NHEJ. Barley revealed a very weak net loss of DNA, apparently due to less active break‐end resection and easier copying of template sequences into breaks. Comparative phylogenomics revealed several footprints of SSA in the A. thaliana genome. Quantitative assessment of DNA gain and loss through DSB repair processes suggests deletion‐biased DSB repair causing ongoing genome shrinking in A. thaliana, whereas genome size in barley remains nearly constant.

In the hyperaccumulator Arabidopsis halleri, the zinc (Zn) vacuolar transporter MTP1 is a key component of hypertolerance. Because protein sequences and functions are highly conserved between A. halleri and Arabidopsis thaliana, Zn tolerance in A. halleri may reflect the constitutively higher MTP1 expression compared with A. thaliana, based on copy number expansion and different cis regulation. Three MTP1 promoters were characterized in A. halleri ecotype I16. The comparison with the A. thaliana MTP1 promoter revealed different expression profiles correlated with specific cis‐acting regulatory elements. The MTP1 5′ untranslated region, highly conserved among A. thaliana, Arabidopsis lyrata and A. halleri, contains a dimer of MYB‐binding motifs in the A. halleri promoters absent in the A. thaliana and A. lyrata sequences. Site‐directed mutagenesis of these motifs revealed their role for expression in trichomes. A. thaliana mtp1 transgenic lines expressing AtMTP1 controlled by the native A. halleri promoter were more Zn‐tolerant than lines carrying mutations on MYB‐binding motifs. Differences in Zn tolerance were associated with different distribution of Zn among plant organs and in trichomes. The different cis‐acting elements in the MTP1 promoters of A. halleri, particularly the MYB‐binding sites, are probably involved in the evolution of Zn tolerance.

Many plant cells retain their totipotency when cultured in vitro. The regulation of shoot regeneration from in vitro culture involves a number of gene products, but the nature of the associated post-transcriptional events remains largely unknown. Here, the post-transcriptional regulator ARGONAUTE10 (AGO10), a protein which is specifically expressed in the explant during the period when pro-shoot apical meristems (SAMs) are forming, has been known to inhibit shoot regeneration. In in vitro cultured explants of the loss-of-function mutant ago10, a much larger than normal number of SAMs was formed and, in these, the stem cell marker genes WUSCHEL, CLAVATA3 and SHOOT MERISTEMLESS were all strongly expressed. AGO10 repressed the accumulation of the microRNAs miR165/166, thereby up-regulating a suite of HD-ZIP III genes. The overproduction of miR166 was shown to promote shoot regeneration, while the absence of miR165/166 message resulted in a blockage to shoot regeneration and only a partial rescue of the phenotype of the ago10 mutant. The major conclusion was that the shoot regeneration inhibition determined by AGO10 functions via the repression of miR165/166.

Isofucosterol is a major 4-demethylsterol which has an ethylidene group at C-24 in Arabidopsis thaliana. To evaluate the presence of brassinosteroids (BRs) with the same carbon skeleton as that of isofucosterol, a large quantity of A. thaliana was extracted and purified. GC-MS/selected ion monitoring analysis verified that 6-deoxohomodolichosterone and homodolichosterone are present in Arabidopsis. An enzyme solution prepared from wild type Arabidopsis successfully mediated conversion of 6-deoxohomodolichosterone to homodolichosterone. However, a double mutant cyp85a1/ cyp85a2 could not catalyze the conversion, implying that in A. thaliana the C-6 oxidation of 6-deoxohomodolichosterone to homodolichosterone seems to be catalyzed by CYP85A1 and/or CYP85A2. In yeast, both heterologously expressed CYP85A1 and CYP85A2 catalyzed the C-6 oxidation of 6- deoxohomodolichosterone to homodolichosterone, but the conversion rate in CYP85A2/V60/WAT21 was significantly higher than that in CYP85A1/V60/WAT21, indicating that C-6 oxidation of 6-deoxohomodolichosterone to homodolichosterone is mainly catalyzed by CYP85A2 in A. thaliana. Taken together, this study strongly suggests that a biosynthetic pathway for the production of 6-deoxohomodolichosterone and homodolichosterone is functional, and CYP85As have important roles in 24-ethylidene biosynthesis in A. thaliana.

A pair of Arabidopsis thaliana resistance proteins, RPS4 and RRS1, recognizes the cognate Avr effector from the bacterial pathogens Pseudomonas syringae pv. tomato expressing avrRps4 (Pst-avrRps4), Ralstonia solanacearum, and the fungal pathogen Colletotrichum higginsianum and leads to defense signaling activation against the pathogens. In the present study, we analyzed 14 A. thaliana accessions for natural variation in Pst-avrRps4 and C. higginsianum susceptibility, and found new compatible and incompatible Arabidopsis–pathogen interactions. We first found that A. thaliana accession Cvi-0 is susceptible to Pst-avrRps4. Interestingly, the genome sequence assembly indicated that Cvi-0 lost both RPS4 and RRS1, but not RPS4B and RRS1B, compared to the reference genome sequence from A. thaliana accession Col-0. On the other hand, the natural variation analysis of RPS4 alleles from various Arabidopsis accessions revealed that one amino-acid change, Y950H, is responsible for the loss of resistance to Pst-avrRps4 and C. higginsianum in RLD-0. Our data indicate that the amino acid change, Y950H, in RPS4 resulted in the loss of both RPS4 and RRS1 functions and resistance to pathogens.

Although plant development is highly reproducible, some stochasticity exists. This developmental stochasticity may be caused by noisy gene expression. Here we analyze the fluctuation of protein expression in Arabidopsis thaliana. Using the photoconvertible KikGR marker, we show that the protein expressions of individual cells fluctuate over time. A dual reporter system was used to study extrinsic and intrinsic noise of marker gene expression. We report that extrinsic noise is higher than intrinsic noise and that extrinsic noise in stomata is clearly lower in comparison to several other tissues/cell types. Finally, we show that cells are coupled with respect to stochastic protein expression in young leaves, hypocotyls and roots but not in mature leaves. Our data indicate that stochasticity of gene expression can vary between tissues/cell types and that it can be coupled in a non-cell-autonomous manner.

[SPA] El reloj circadiano es un mecanismo que proporciona ventajas adaptativas y está presente en todos los organismos. El reloj circadiano en plantas, como se describe en Arabidopsis (Arabidopsis thaliana), está formado por una serie de bucles interconectados y retroalimentados, pero no existe mucha información sobre su función en procesos como la emisión de compuestos volátiles orgánicos. En el presente trabajo analizaremos la emisión de volátiles en dos condiciones experimentales (medio ambientales?) en la planta modelo boca de dragón (Antirrhinum majus). [ENG] The circadian clock is a mechanism that provides adaptive advantages being present in all living organisms. The circadian clock in plants, as described in Arabidopsis (Arabidopsis thaliana), is comprised of a series of interlocked regulatory feedback loops, but there is not much information about its function on processes such as the emission of volatile organic compounds. In this study, we will analyze volatile emission under two different environmental conditions in snapdragon (Antirrhinum majus). | Agradecemos a la Dr. María José Roca el soporte ofrecido con los análisis GC-MS. El trabajo se realiza dentro del proyecto MICINN BFU-45148-R.

As the major resource of reactive oxygen species (ROS), the NADPH oxidases (Rbohs) have been shown to play important roles in plant cells under normal growth and stress conditions. Although many family members of Rbohs were studied, little is known about the function of RbohI in Arabidopsis thaliana. Here, we report that exogenous ABA application decreases RbohI expression and mannitol significantly increases RbohI expression at transcript level. The RbohI transcripts were strongly detected in dry seeds and roots. The loss-of-function mutant rbohI exhibited sensitivity to ABA and mannitol stress during germination. Furthermore, the lateral root growth of rbohI was severely inhibited after treatment with mannitol stress. Overexpression of RbohI in Arabidopsis significantly improves the drought tolerance. Moreover, more H2O2 accumulated in RbohI overexpressors than in wild type plants in response to mannitol stress. Our conclusion is that AtRbohI functions in drought-stress response in Arabidopsis thaliana.

Pseudomonas syringae pv. tomato strain DC3000 (Pto DC3000), which causes bacterial speck disease of tomato, has been used as a model pathogen because of its pathogenicity on Arabidopsis thaliana. Here, we demonstrate a rapid and reliable flood-inoculation method based on young Arabidopsis seedlings grown on one-half strength MS medium. We also describe a method to evaluate the bacterial growth in Arabidopsis.