The role of COP1/SPA in light signaling: Growth control, cell-cell communication and functional conservation in plants

Light is one of the most important environmental factors affecting almost all stages of plant growth and development. Arabidopsis SPA and COP1 proteins act as repressors of light signaling in darkness. Members of the SPA protein family (SPA1-SPA4) can physically interact with COP1 and, together, they constitute a functional E3-ubiquitin ligase complex. The COP1/SPA complex regulates seedling development, stomata differentiation, leaf size and photoperiodic flowering in Arabidopsis by targeting transcription factors such as HY5, HFR1, CO etc. for degradation. In the present study, I investigated in which tissues SPA1 needs to be expressed to regulate different plant developmental processes. To this end, I expressed a GUS-SPA1 fusion protein under the control of various tissue-specific promoters (phloem, leaf-mesophyll, epidermis, meristem and root) in a spa mutant background and analyzed the transgenic plants for complementation of the spa mutant phenotype. The results show that SPA1 functions exclusively in the phloem to regulate photoperiodic flowering suggesting that SPA1 acts cell-autonomously in the phloem to target its substrate CO for degradation. To regulate the leaf size, SPA1 acts in both the phloem and the leaf mesophyll, but not in the epidermis indicating non-cell autonomous effects in SPA1-dependent leaf size regulation. Moreover, phloem-specific expression of SPA1 has major effects on seedling development in both darkness and light. Eventually, stomata differentiation and epidermal pavement cell shape are also regulated by phloem-specific functions of SPA1. These results indicate that cell-cell communication plays a very important role in SPA1-regulated plant developmental processes. SPA proteins and, therefore, the COP1/SPA complexes are plant specific. However, the function of COP1 and SPA proteins are not known in plant species other than the dicot Arabidopsis. In a second project, I examined the functionality of the COP1 and SPA proteins from the moss Physcomitrella and the monocot rice in Arabidopsis. To this end, I expressed the open reading frames of rice and Physcomitrella COP1 and SPA homologs in Arabidopsis cop1 and spa mutant plants, respectively, and then analyzed the transgenic plants for complementation of the respective mutant phenotypes. Rice and Physcomitrella COP1 homologs were functional in Arabidopsis, whereas SPA homologs from these species were not functional, suggesting a conserved basic mechanism of action of COP1, but functional divergence of SPA proteins during plant evolution. Interestingly, Physcomitrella COP1 and SPA proteins interact in vitro suggesting the possibility of formation of a COP1/SPA complex early in evolution

A New Advanced Backcross Tomato Population Enables High Resolution Leaf QTL Mapping and Gene Identification.

Quantitative Trait Loci (QTL) mapping is a powerful technique for dissecting the genetic basis of traits and species differences. Established tomato mapping populations between domesticated tomato (Solanum lycopersicum) and its more distant interfertile relatives typically follow a near isogenic line (NIL) design, such as the S. pennellii Introgression Line (IL) population, with a single wild introgression per line in an otherwise domesticated genetic background. Here, we report on a new advanced backcross QTL mapping resource for tomato, derived from a cross between the M82 tomato cultivar and S. pennellii This so-called Backcrossed Inbred Line (BIL) population is comprised of a mix of BC2 and BC3 lines, with domesticated tomato as the recurrent parent. The BIL population is complementary to the existing S. pennellii IL population, with which it shares parents. Using the BILs, we mapped traits for leaf complexity, leaflet shape, and flowering time. We demonstrate the utility of the BILs for fine-mapping QTL, particularly QTL initially mapped in the ILs, by fine-mapping several QTL to single or few candidate genes. Moreover, we confirm the value of a backcrossed population with multiple introgressions per line, such as the BILs, for epistatic QTL mapping. Our work was further enabled by the development of our own statistical inference and visualization tools, namely a heterogeneous hidden Markov model for genotyping the lines, and by using state-of-the-art sparse regression techniques for QTL mapping

The tomato receptor CuRe1 senses a cell wall protein to identify Cuscuta as a pathogen

Parasitic plants of the genus Cuscuta penetrate shoots of host plants with haustoria and build a connection to the host vasculature to exhaust water, solutes and carbohydrates. Such infections usually stay unrecognized by the host and lead to harmful host plant damage. Here, we show a molecular mechanism of how plants can sense parasitic Cuscuta. We isolated an 11 kDa protein of the parasite cell wall and identified it as a glycine-rich protein (GRP). This GRP, as well as its minimal peptide epitope Crip21, serve as a pathogen-associated molecular pattern and specifically bind and activate a membrane-bound immune receptor of tomato, the Cuscuta Receptor 1 (CuRe1), leading to defense responses in resistant hosts. These findings provide the initial steps to understand the resistance mechanisms against parasitic plants and further offer great potential for protecting crops by engineering resistance against parasitic plants

Randomised study on single stage laparo-endoscopic rendezvous (intra-operative ERCP) procedure versus two stage approach (Pre-operative ERCP followed by laparoscopic cholecystectomy) for the management of cholelithiasis with choledocholithiasis

Introduction : The ′Rendezvous′ technique consists of laparoscopic cholecystectomy (LC) standards with intra-operative cholangiography followed by endoscopic sphincterotomy. The sphincterotome is driven across the papilla through a guidewire inserted by the transcystic route. In this study, we intended to compare the two methods in a prospective randomised trial. Materials And Methods: From 2005 to 2012, we enrolled 83 patients with a diagnosis of cholecysto-choledocolithiasis. They were randomised into two groups. In ′group-A′,41 patients were treated with two stages management, first by pre-operative endoscopic retrograde cholangiopancreatography (ERCP) and common bile duct (CBD) clearance and second by LC. In ′group-B′, 42 patients were treated with LC and intra-operative cholangiography; and when diagnosis of choledocholithiasis was confirmed, patients had undergone one stage management of by Laparo-endoscopic Rendezvous technique. Results: In arm-A and arm-B groups, complete CBD clearance was achieved in 29 and 38 patients, respectively. Failure of the treatment in arm-A was 29% and in arm-B was 9.5%. In arm-A, selective CBD cannulation was achieved in 33 cases (80.5%) and in arm-B in 39 cases (93%). In arm-Agroup, post-ERCP hyperamylasia was presented in nine patients (22%) and severe pancreatitis in five patients (12%) versus none of the patients (0%) in arm-B group, respectively. Mean post-operative hospital stay in arm-A and arm-B groups are 10.9 and 6.8 days, respectively. Conclusion: One stage laparo-endoscopic rendezvous approach increases selective cannulation of CBD, reduces post-ERCP pancreatitis, reduces days of hospital stay, increases patient′s compliance and prevents unnecessary intervention to CBD

Auxin represses stomatal development in dark-grown seedlings via Aux/IAA proteins

Stomatal development is tightly regulated through internal and external factors that are integrated by a complex signalling network. Light represents an external factor that strongly promotes stomata formation. Here, we show that auxin-resistant aux/iaa mutants, e.g. axr3-1, exhibit a de-repression of stomata differentiation in dark-grown seedlings. The higher stomatal index in dark-grown axr3-1 mutants when compared with the wild type is due to increased cell division in the stomatal lineage. Excessive stomata in dark-grown seedlings were also observed in mutants defective in auxin biosynthesis or auxin perception and in seedlings treated with the polar auxin transport inhibitor NPA. Consistent with these findings, exogenous auxin repressed stomata formation in light-grown seedlings. Taken together, these results indicate that auxin is a negative regulator of stomatal development in dark-grown seedlings. Epistasis analysis revealed that axr3-1 acts genetically upstream of the bHLH transcription factors SPCH, MUTE and FAMA, as well as the YDA MAP kinase cascade, but in parallel with the repressor of photomorphogenesis COP1 and the receptor-like protein TMM. The effect of exogenous auxin required the ER family of leucine-rich repeat receptor-like kinases, suggesting that auxin acts at least in part through the ER family. Expression of axr3-1 in the stomatal lineage was insufficient to alter the stomatal index, implying that cell-cell communication is necessary to mediate the effect of auxin. In summary, our results show that auxin signalling contributes to the suppression of stomatal differentiation observed in dark-grown seedlings