A Subtle Interplay Between Three Pex11 Proteins Shapes De Novo Formation and Fission of Peroxisomes

The organization of eukaryotic cells into membrane-bound compartments must be faithfully sustained for survival of the cell. A subtle equilibrium exists between the degradation and the proliferation of organelles. Commonly, proliferation is initiated by a membrane remodeling process. Here, we dissect the function of proteins driving organelle proliferation in the particular case of peroxisomes. These organelles are formed either through a growth and division process from existing peroxisomes or de novo from the endoplasmic reticulum (ER). Among the proteins involved in the biogenesis of peroxisomes, peroxins, members of the Pex11 protein family participate in peroxisomal membrane alterations. In the yeast Saccharomyces cerevisiae, the Pex11 family consists of three proteins, Pex11p, Pex25p and Pex27p. Here we demonstrate that yeast mutants lacking peroxisomes require the presence of Pex25p to regenerate this organelle de novo. We also provide evidence showing that Pex27p inhibits peroxisomal function and illustrate that Pex25p initiates elongation of the peroxisomal membrane. Our data establish that although structurally conserved each of the three Pex11 protein family members plays a distinct role. While ScPex11p promotes the proliferation of peroxisomes already present in the cell, ScPex25p initiates remodeling at the peroxisomal membrane and ScPex27p acts to counter this activity. In addition, we reveal that ScPex25p acts in concert with Pex3p in the initiation of de novo peroxisome biogenesis from the ER

Early detection of grapevine graft incompatibility: insights into translocated and virus-induced incompatibility

In vineyards to control phylloxera (Daktulosphaira vitifolia Ficth) attacks in Vitis vinifera L., heterografted vines are planted using American vines hybrids as rootstocks. However, graft incompatibilities can affect grape yield and plant longevity. Thus, to identify early graft incompatibility factors, we established in vitro micrografting protocols coupled with histology and histochemistry analysis in grapevine graft combinations of known compatibility behavior. The histochemical characterization of the graft union revealed irregular cell arrangement, slower vascular differentiation, persistence of the necrotic layer, accumulation of starch, and lower differentiation of phloem cells in hetero- compared to homografts, indicating the presence of translocated incompatibility symptoms. We highlight the utility of evaluating the graft interface cellular arrangement and starch content via calcofluor and I2KI staining, respectively, as allowed to identify the graft combinations with lower graft success. Wounded and grafted Syrah plantlets pointed out an impaired sucrose distribution in these plants and levels of Grapevine Rupestris Stem Pitting associated Virus (GRSPaV) infections correlated with graft (un)-success in two Syrah clones micrografted onto 110-Ritcher rootstock. Furthermore, silencing GRSPaV before grafting increased graft success rates. We propose that grapevine graft incompatibility is mainly a virus-induced phenomenon that can arise even in certified plants

Two-Dimensional Patterning by a Trapping/Depletion Mechanism: The Role of TTG1 and GL3 in Arabidopsis Trichome Formation

Trichome patterning in Arabidopsis serves as a model system to study how single cells are selected within a field of initially equivalent cells. Current models explain this pattern by an activator–inhibitor feedback loop. Here, we report that also a newly discovered mechanism is involved by which patterning is governed by the removal of the trichome-promoting factor TRANSPARENT TESTA GLABRA1 (TTG1) from non-trichome cells. We demonstrate by clonal analysis and misexpression studies that Arabidopsis TTG1 can act non-cell-autonomously and by microinjection experiments that TTG1 protein moves between cells. While TTG1 is expressed ubiquitously, TTG1–YFP protein accumulates in trichomes and is depleted in the surrounding cells. TTG1–YFP depletion depends on GLABRA3 (GL3), suggesting that the depletion is governed by a trapping mechanism. To study the potential of the observed trapping/depletion mechanism, we formulated a mathematical model enabling us to evaluate the relevance of each parameter and to identify parameters explaining the paradoxical genetic finding that strong ttg1 alleles are glabrous, while weak alleles exhibit trichome clusters

In vitro early detection of grapevine virus-induced graft incompatibility in Syrah/R110: the involvement of GRSPaV

In grapevines, incompatibility manifests in short-term graft failure or long-term decline of vineyards both causing economic losses to nurseries and growers. Here, we assessed the suitability of in vitro systems coupled with histochemistry analysis as early detection methods for grapevine incompatibility using certified homografted (i.e., a graft between the same genotype) and heterografted (i.e., a graft between two genotypes) grapevine micrografts, with known graft compatibility response when grafted onto the worldwide used rootstock 110R (V. berlandieri x V. rupestris). To prove the involvement of Grapevine rupestris stem pitting-associated virus (GRSPaV) in Syrah/110R incompatibility, we silenced GRSPaV transcripts by ectopically applying a GRSPaV siRNA-inducing dsRNA

Circadian, carbon, and light control of expansion growth and leaf movement

We used Phytotyping4D to investigate the contribution of clock and light signaling to the diurnal regulation of rosette expansion growth and leaf movement in Arabidopsis (Arabidopsis thaliana). Wild-type plants and clock mutants with a short (lhycca1) and long (prr7prr9) period were analyzed in a T24 cycle and in T-cycles that were closer to the mutants’ period. Wild types also were analyzed in various photoperiods and after transfer to free-running light or darkness. Rosette expansion and leaf movement exhibited a circadian oscillation, with superimposed transients after dawn and dusk. Diurnal responses were modified in clock mutants. lhycca1 exhibited an inhibition of growth at the end of night and growth rose earlier after dawn, whereas prr7prr9 showed decreased growth for the first part of the light period. Some features were partly rescued by a matching T-cycle, like the inhibition in lhycca1 at the end of the night, indicating that it is due to premature exhaustion of starch. Other features were not rescued, revealing that the clock also regulates expansion growth more directly. Expansion growth was faster at night than in the daytime, whereas published work has shown that the synthesis of cellular components is faster in the day than at nighttime. This temporal uncoupling became larger in short photoperiods and may reflect the differing dependence of expansion and biosynthesis on energy, carbon, and water. While it has been proposed that leaf expansion and movement are causally linked, we did not observe a consistent temporal relationship between expansion and leaf movement.This work was supported by the Max Planck Society, the European Union (collaborative project TiMet under contract no. 245143), and an International Max Planck Research School stipend (to D.B.)

The Phloem-Delivered RNA Pool Contains Small Noncoding RNAs and Interferes with Translation1[W][OA]

In plants, the vascular tissue contains the enucleated sieve tubes facilitating long-distance transport of nutrients, hormones, and proteins. In addition, several mRNAs and small interfering RNAs/microRNAs were shown to be delivered via sieve tubes whose content is embodied by the phloem sap (PS). A number of these phloem transcripts are transported from source to sink tissues and function at targeted tissues. To gain additional insights into phloem-delivered RNAs and their potential role in signaling, we isolated and characterized PS RNA molecules distinct from microRNAs/small interfering RNAs with a size ranging from 30 to 90 bases. We detected a high number of full-length and phloem-specific fragments of noncoding RNAs such as tRNAs, ribosomal RNAs, and spliceosomal RNAs in the PS of pumpkin (Cucurbita maxima). In vitro assays show that small quantities of PS RNA molecules efficiently inhibit translation in an unspecific manner. Proof of concept that PS-specific tRNA fragments may interfere with ribosomal activity was obtained with artificially produced tRNA fragments. The results are discussed in terms of a functional role for long distance delivered noncoding PS RNAs

MPB2C, a Microtubule-Associated Protein, Regulates Non-Cell-Autonomy of the Homeodomain Protein KNOTTED1[W][OA]

Plasmodesmata establish a pathway for the intercellular trafficking of viral movement proteins and endogenous non-cell-autonomous proteins, such as the two closely related meristem-maintaining KNOTTED1-like homeobox (KNOX) proteins Zea mays KNOTTED1 (KN1) and Arabidopsis thaliana SHOOTMERISTEMLESS (STM). KNOX family members are DNA binding proteins that regulate the transcriptional activity of target genes in conjunction with BEL1-like homeodomain proteins. It has been shown previously, using in vivo transport assays, that the C-terminal domain of KN1, including the homeodomain, is necessary and sufficient for cell-to-cell transport through plasmodesmata. Here, using interaction and coexpression assays, we demonstrate that the microtubule-associated and viral movement protein binding protein MPB2C from Nicotiana tabacum, and its homolog in Arabidopsis, At MPB2C, are KN1/STM binding factors. Interaction between the MPB2C proteins and KN1/STM was mapped to the KN1 homeodomain, a region not essential for heterodimerization with BEL1. Expression of MPB2C in single cells prevented KN1 cell-to-cell movement. Furthermore, in vivo trichome rescue studies established that MPB2C negatively regulates KN1 association to plasmodesmata and, consequently, cell-to-cell transport. These findings are discussed in terms of the role played by MPB2C proteins in regulating the cell-to-cell trafficking of homeodomain proteins in plants

A Phenotypic Search on Graft Compatibility in Grapevine

Grafting is the most used propagation method in viticulture and is the unique control strategy against Phylloxera. Nevertheless, its practice remains limited mainly due to inconsistent graft success and difficulties in predicting graft compatibility responses of proposed scion–rootstock combinations, slowing down the selection of elite rootstocks. Aiming to identify optimal phenotypic parameters related to graft (in)compatibility, we used four clones of two grapevine cultivars that show different compatibility behavior when grafted onto the same rootstock. Several physiological parameters, internal anatomy of the graft union, chlorophyll fluorescence, and pigment contents of homo- and heterografts were monitored in a nursery-grafting context. The measurements highlighted enhanced performance of the heterografts due to rooting difficulties of Vitis vinifera homografts. This suggests that in viticulture, homografts should only be used as compatibility controls regarding qualitative attributes. By observing the internal anatomy of the union, we found that grapevines might require longer times for graft healing than anticipated. While Affinity Coefficients were not informative to assess incompatibility, leaf chlorophyll concentration analysis proved to be a more sensitive indicator of stress than the analysis of chlorophyll fluorescence. Overall, we conclude that graft take correlated best with callus formation at the graft junction three weeks after grafting.Publishe