Characterization of cCe13, a Member of the Pepper Endo‐β‐1, 4‐Glucanase Multigene Family

In pepper plants the enzyme endo-beta-1,4-glucanase (EGase) is encoded by a multigene family. Here is described the characterization of cCel3, a cDNA which codes for the third EGase known so far in this plant. The known members of this family are present as single copy genes, as demonstrated by a Southern analysis of the genomic DNA. Analysis of the expression of cCel3 demonstrates that the highest levels of the cCel3 mRNA are found in abscission zones of leaves and flowers activated by treatments with the plant hormone ethylene. However, the amounts of the cCel3 transcripts (detectable only by RT-PCR) are always much lower than those of cCel2 so that the latter can be regarded as the "abscission" EGase while cCel3 is likely to perform a role ancillary to that of cCel2

Expression of an Endopolygalacturonase Gene During Growth and Abscission of Peach Fruits

Polygalacturonase (PG, EC 3.2.1.15) is one of the cell wall hydrolases involved in the cell separation processes which occur during ripening of some fleshy fruits (FISHER and BENNETT 1991), as well as during abscission of leaves and fruits (HUBERMANN and GOREN 1979; RASC~O et al. 1985; TAYLOR et al. 1990; BONGHI et al. 1992; TAYLOR et al. 1993). In particular, endopolygalacturonase of tomato fruit is the most widely known form of this enzyme, having been characterized at molecular and biochemical level (see literature in ZHENG et al. 1992). Endopolygalacturonase activity has also been found in peach during both abscission and development of fruits, while no such activity could be detected during leaf abscission (BONGHI et al. 1992; ZANCHIN et al. 1993). A few years ago it was observed that polyclonal antibodies raised against a tomato fruit PG (i.e., PGZA) recognized, in soft ripe fruits of peach, a polypeptide with molecular mass similar to that of PG2A. Furthermore, using as a probe a cDNA coding for tomato fruit endopolygalacturonase, the same researchers were able to clone and characterize a 3.5 kb fragment of peach genomic DNA (LEE et al. 1990). On the basis of sequence analysis it was concluded that, besides an unidentified sequence, it contained about the 3' half of a gene which showed, in the coding regions, extensive homology with the tomato PG gene. According to the same researchers, such homology could explain the observed cross-reaction between the antibody to tomato fruit PG and a peach polypeptide, which was therefore suggested to be a peach endopolygalacturonase and the product of the partly characterized gene (LEE et al. 1990). In tomato the gene encoding the fruit endopolygalacturonase seems to be expressed during the fruit ripening, but not during the leaf abscission. In fact, despite a significative rise in PG activity, an antibody to fruit PG did not recognize any leaf abscission protein. Moreover, a cDNA encoding a tomato fruit PG gave no hybridization to mRNA obtained from activated abscission zones of tomato leaves (TAYLOR et al. 1990). In peach it has recently been shown that a cDNA coding for tomato fruit PG hybridized to mRNA obtained from fruit abscission zones but not from leaf ones where, in any case, no PG activity had been detected (BONGHI et al. 1992). In peach, cell separation events which show an involvement of endopolygalacturonase, are not restricted to fruit softening and abscission. Recently, it has been found that PG activity can also be detected throughout the fruit growth ( ZANCHIN et al. in press). On the basis of the above findings we considered it of some interest to see whether the endopolygalacturonase activity, observed in the course of different cell separation events in peach, is due to expression of the partly known PG gene (LEE et al. 1990) or, as already observed in tomato (TAYLOR et al. 1990), only some of that activity can be ascribed to expression of that gene

Transcriptional regulatory networks controlling woolliness in peach in response to preharvest gibberellin application and cold storage

BACKGROUND: Postharvest fruit conservation relies on low temperatures and manipulations of hormone metabolism to maintain sensory properties. Peaches are susceptible to chilling injuries, such as ‘woolliness’ that is caused by juice loss leading to a ‘wooly’ fruit texture. Application of gibberellic acid at the initial stages of pit hardening impairs woolliness incidence, however the mechanisms controlling the response remain unknown. We have employed genome wide transcriptional profiling to investigate the effects of gibberellic acid application and cold storage on harvested peaches. RESULTS: Approximately half of the investigated genes exhibited significant differential expression in response to the treatments. Cellular and developmental process gene ontologies were overrepresented among the differentially regulated genes, whereas sequences in cell death and immune response categories were underrepresented. Gene set enrichment demonstrated a predominant role of cold storage in repressing the transcription of genes associated to cell wall metabolism. In contrast, genes involved in hormone responses exhibited a more complex transcriptional response, indicating an extensive network of crosstalk between hormone signaling and low temperatures. Time course transcriptional analyses demonstrate the large contribution of gene expression regulation on the biochemical changes leading to woolliness in peach. CONCLUSION: Overall, our results provide insights on the mechanisms controlling the complex phenotypes associated to postharvest textural changes in peach and suggest that hormone mediated reprogramming previous to pit hardening affects the onset of chilling injuries. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12870-015-0659-2) contains supplementary material, which is available to authorized users

The Peach RGF/GLV Signaling Peptide pCTG134 Is Involved in a Regulatory Circuit That Sustains Auxin and Ethylene Actions

In vascular plants the cell-to-cell interactions coordinating morphogenetic and physiological processes are mediated, among others, by the action of hormones, among which also short mobile peptides were recognized to have roles as signals. Such peptide hormones (PHs) are involved in defense responses, shoot and root growth, meristem homeostasis, organ abscission, nutrient signaling, hormone crosstalk and other developmental processes and act as both short and long distant ligands. In this work, the function of CTG134, a peach gene encoding a ROOT GROWTH FACTOR/GOLVEN-like PH expressed in mesocarp at the onset of ripening, was investigated for its role in mediating an auxin-ethylene crosstalk. In peach fruit, where an auxin-ethylene crosstalk mechanism is necessary to support climacteric ethylene synthesis, CTG134 expression peaked before that of ACS1 and was induced by auxin and 1-methylcyclopropene (1-MCP) treatments, whereas it was minimally affected by ethylene. In addition, the promoter of CTG134 fused with the GUS reporter highlighted activity in plant parts in which the auxin-ethylene interplay is known to occur. Arabidopsis and tobacco plants overexpressing CTG134 showed abnormal root hair growth, similar to wild-type plants treated with a synthetic form of the sulfated peptide. Moreover, in tobacco, lateral root emergence and capsule size were also affected. In Arabidopsis overexpressing lines, molecular surveys demonstrated an impaired hormonal crosstalk, resulting in a re-modulated expression of a set of genes involved in both ethylene and auxin synthesis, transport and perception. These data support the role of pCTG134 as a mediator in an auxin-ethylene regulatory circuit and open the possibility to exploit this class of ligands for the rational design of new and environmental friendly agrochemicals able to cope with a rapidly changing environment

A microarray approach to identify genes involved in seed-pericarp cross-talk and development in peach

<p>Abstract</p> <p>Background</p> <p>Field observations and a few physiological studies have demonstrated that peach embryogenesis and fruit development are tightly coupled. In fact, attempts to stimulate parthenocarpic fruit development by means of external tools have failed. Moreover, physiological disturbances during early embryo development lead to seed abortion and fruitlet abscission. Later in embryo development, the interactions between seed and fruit development become less strict. As there is limited genetic and molecular information about seed-pericarp cross-talk and development in peach, a massive gene approach based on the use of the μPEACH 1.0 array platform and quantitative real time RT-PCR (qRT-PCR) was used to study this process.</p> <p>Results</p> <p>A comparative analysis of the transcription profiles conducted in seed and mesocarp (cv Fantasia) throughout different developmental stages (S1, S2, S3 and S4) evidenced that 455 genes are differentially expressed in seed and fruit. Among differentially expressed genes some were validated as markers in two subsequent years and in three different genotypes. Seed markers were a LTP1 (lipid transfer protein), a PR (pathogenesis-related) protein, a prunin and LEA (Late Embryogenesis Abundant) protein, for S1, S2, S3 and S4, respectively. Mesocarp markers were a RD22-like protein, a serin-carboxypeptidase, a senescence related protein and an Aux/IAA, for S1, S2, S3 and S4, respectively.</p> <p>The microarray data, analyzed by using the HORMONOMETER platform, allowed the identification of hormone-responsive genes, some of them putatively involved in seed-pericarp crosstalk. Results indicated that auxin, cytokinins, and gibberellins are good candidates, acting either directly (auxin) or indirectly as signals during early development, when the cross-talk is more active and vital for fruit set, whereas abscisic acid and ethylene may be involved later on.</p> <p>Conclusions</p> <p>In this research, genes were identified marking different phases of seed and mesocarp development. The selected genes behaved as good seed markers, while for mesocarp their reliability appeared to be dependent upon developmental and ripening traits. Regarding the cross-talk between seed and pericarp, possible candidate signals were identified among hormones.</p> <p>Further investigations relying upon the availability of whole genome platforms will allow the enrichment of a marker genes repertoire and the elucidation of players other than hormones that are involved in seed-pericarp cross-talk (i.e. hormone peptides and microRNAs).</p

Biotecnologie in classe: un intervento di peer tutoring con studenti della scuola secondaria

The article presents a school project implemented in the framework of “Piano Nazionale Lauree Scientifiche”, using a Student Voice approach: a group of high school students facilitated a laboratory focused on biotechnology, playing the role of tutor for their peers. A qualitative research was carried out in order to answer to the following questions: what are the students’ perceptions about the teaching and learning method used, namely the peer tutoring? What are the students’ perceptions about the topic of biotechnology, with specific focus on GMOs and what is the impact of the laboratory experience to guide students’ university future choices? The research aimed at investigating strengths and the critical aspects of this project, in order to improve future editions. Findings showed that the two active learning methods used, i.e. the peer tutoring and laboratory, allowed students to experience leadership and to develop their competences, by managing a complex task with responsibility and autonomy within a teamwork.L’articolo presenta l’esperienza di un progetto scolastico svolto con un approccio Student Voice, in cui un gruppo di studenti liceali hanno assunto il ruolo di tutor per i propri compagni, gestendo un laboratorio sulle biotecnologie, promosso all’interno del Piano Nazionale Lauree Scientifiche. L’esperienza è stata oggetto di una ricerca qualitativa, mossa dai seguenti interrogativi: quali sono le percezioni degli studenti in merito al metodo didattico utilizzato e cioè il peer tutoring? Quali sono le percezioni degli studenti in merito ai contenuti proposti, cioè le biotecnologie ed in particolare gli OGM, nonché il contributo dato dal laboratorio per l’orientamento alla scelta universitaria? La finalità era raccogliere i punti di forza e le criticità di tale esperienza per migliorare le future edizioni. L’indagine ha confermato come i due metodi di didattica attiva utilizzati e cioè il peer tutoring ed il laboratorio hanno permesso ai ragazzi di fare una esperienza di protagonismo e sviluppo di competenze, in cui sono stati capaci di gestire con responsabilità ed autonomia un compito complesso all’interno di un lavoro di squadra

A PLENA-like gene of peach is involved in carpel formation and subsequent transformation into a fleshy fruit

MADS-box genes have been shown to play a role in the formation of fruits, both in Arabidopsis and in tomato. In peach, two C-class MADS-box genes have been isolated. Both of them are expressed during flower and mesocarp development. Here a detailed analysis of a gene that belongs to the PLENA subfamily of MADS-box genes is shown. The expression of this PLENA-like gene (PpPLENA) increases during fruit ripening, and its ectopic expression in tomato plants causes the transformation of sepals into carpel-like structures that become fleshy and ripen like real fruits. Interestingly, the transgenic berries constitutively expressing the PpPLENA gene show an accelerated ripening, as judged by the expression of genes that are important for tomato fruit ripening. It is suggested that PpPLENA might interfere with the endogenous activity of TAGL1, thereby activating the fruit ripening pathway earlier compared with wild-type tomato plants

Transcriptomic and photosynthetic analyses of Synechocystis sp. PCC6803 and Chlorogloeopsis fritschii sp. PCC6912 exposed to an M-dwarf spectrum under an anoxic atmosphere

IntroductionCyanobacteria appeared in the anoxic Archean Earth, evolving for the first time oxygenic photosynthesis and deeply changing the atmosphere by introducing oxygen. Starting possibly from UV-protected environments, characterized by low visible and far-red enriched light spectra, cyanobacteria spread everywhere on Earth thanks to their adaptation capabilities in light harvesting. In the last decade, few cyanobacteria species which can acclimate to far-red light through Far-Red Light Photoacclimation (FaRLiP) have been isolated. FaRLiP cyanobacteria were thus proposed as model organisms to study the origin of oxygenic photosynthesis as well as its possible functionality around stars with high far-red emission, the M-dwarfs. These stars are astrobiological targets, as their longevity could sustain life evolution and they demonstrated to host rocky terrestrial-like exoplanets within their Habitable Zone.MethodsWe studied the acclimation responses of the FaRLiP strain Chlorogloeopsis fritschii sp. PCC6912 and the non-FaRLiP strain Synechocystis sp. PCC6803 to the combination of three simulated light spectra (M-dwarf, solar and far-red) and two atmospheric compositions (oxic, anoxic). We first checked their growth, O2 production and pigment composition, then we studied their transcriptional responses by RNA sequencing under each combination of light spectrum and atmosphere conditions.Results and discussionPCC6803 did not show relevant differences in gene expression when comparing the responses to M-dwarf and solar-simulated lights, while far-red caused a variation in the transcriptional level of many genes. PCC6912 showed, on the contrary, different transcriptional responses to each light condition and activated the FaRLiP response under the M-dwarf simulated light. Surprisingly, the anoxic atmosphere did not impact the transcriptional profile of the 2 strains significantly. Results show that both cyanobacteria seem inherently prepared for anoxia and to harvest the photons emitted by a simulated M-dwarf star, whether they are only visible (PCC6803) or also far-red photons (PCC6912). They also show that visible photons in the simulated M-dwarf are sufficient to keep a similar metabolism with respect to solar-simulated light.ConclusionResults prove the adaptability of the cyanobacterial metabolism and enhance the plausibility of finding oxygenic biospheres on exoplanets orbiting M-dwarf stars

The Italian National Project of Astrobiology-Life in Space-Origin, Presence, Persistence of Life in Space, from Molecules to Extremophiles

The \u2018\u2018Life in Space\u2019\u2019 project was funded in the wake of the Italian Space Agency\u2019s proposal for the development of a network of institutions and laboratories conceived to implement Italian participation in space astrobiology experiments