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Petunia, the goose, learns that possessing knowledge doesn't mean carrying a book around constantly.

In 1973, Jaffe identified and characterized the phenomenon of thigmomorphogenesis, also referred to as mechanical stress (MS) or mechanical stimulation in plants. Previous studies on petunia plants demonstrated that MS significantly affects growth dynamics. As a response to MS, petunias exhibit increased levels of indole-3-acetic acid (IAA) oxidase and peroxidase, although the active transport of endogenous IAA remains unaffected. Furthermore, earlier research has shown that MS inhibits the synthesis of IAA and gibberellin (GA 3 ), with noticeable effects on the 14th day of mechanical stimulation. The current experiment made on Petunia  ×  atkinsiana 'Pegasus Special Burgundy Bicolor’ focused on evaluating the morphological and physiological responses to MS, along with the expression of specific touch-responsive genes such as GH3.1, which is involved in auxin metabolism, and calmodulins (CaMs), playing an important role in stress responses. GH3.1 expression was found to be negatively correlated with IAA synthesis while positively correlated with GAs synthesis and IAA oxidase activity. Variable expression patterns were observed in the calmodulins: CAM53 and CAM81 expression positively correlated with IAA synthesis and plant height, whereas CAM72 expression was positively associated with GAs levels and IAA oxidase activity in plants touched 80× per day, but all of them were negatively related to IAA content and shoot increment, while positively related to GAs synthesis and IAA oxidase activity.

The Lc petunia system, which displays enhanced, light-induced vegetative pigmentation, was used to investigate how high light affects anthocyanin biosynthesis, and to assess the effects of anthocyanin pigmentation upon photosynthesis. Lc petunia plants displayed intense purple anthocyanin pigmentation throughout the leaves and stems when grown under high-light conditions, yet remain acyanic when grown under shade conditions. The coloured phenotypes matched with an accumulation of anthocyanins and flavonols, as well as the activation of the early and late flavonoid biosynthetic genes required for flavonol and anthocyanin production. Pigmentation in Lc petunia only occurred under conditions which normally induce a modest amount of anthocyanin to accumulate in wild-type Mitchell petunia [Petunia axillarisx(Petunia axillarisxPetunia hybrida cv. 'Rose of Heaven')]. Anthocyanin pigmentation in Lc petunia leaves appears to screen underlying photosynthetic tissues, increasing light saturation and light compensation points, without reducing the maximal photosynthetic assimilation rate (Amax). In the Lc petunia system, where the bHLH factor Leaf colour is constitutively expressed, expression of the bHLH (Lc) and WD40 (An11) components of the anthocyanin regulatory system were not limited, suggesting that the high-light-induced anthocyanin pigmentation is regulated by endogenous MYB transcription factors.

Background Development ornamental varieties with enhanced floral traits and distinctive characteristics is a central goal in floriculture. Anthocyanins, the principal flavonoid pigments in floral tissues, are synthesized via the well-characterized flavonoid biosynthesis pathway. Chalcone isomerase ( CHI ) catalyzes the first committed reaction in the flavonoid biosynthetic pathway. Method In this study, we investigated the effects of CHI suppression via RNA interference (RNAi) in three Petunia hybrida cultivars exhibiting distinct petal colors. Leaf discs from three Petunia hybrida lines exhibiting distinct petal colors (blue, pink, and purple) were inoculated with Agrobacterium tumefaciens harboring the recombinant plasmid pBI121 containing the RNAi construct targeting Phchi . Transgenic plants were regenerated on MS medium via microshoots emerging from wounded explant regions and verified by PCR using two primer pairs specific to the CHI silencing vector. Gene expression analysis was conducted for chi and associated pigment biosynthesis genes, including flavanone 3-hydroxylase (F3H) , flavonoid 3′-hydroxylase (F3′H) , flavonoid 3′,5′-hydroxylase (F3′5′H) , and dihydroflavonol 4-reductase (DFR) , comparing transgenic and wild-type plants. Chalcone and naringenin contents were quantified using a NanoDrop ELISA plate reader. Data analysis was performed using SPSS version 16, and mean comparisons were evaluated via T-test at a significance level of α = 0.05. Results Transformation efficiencies for CHI -silenced Petunia lines were 57.89%, 72.0% and 84.0% for the pink, blue, and purple phenotypes, respectively. Phenotypic evaluation revealed not only altered pigment distributions but also novel floral morphologies, such as tetramerous corollas and twisted tubular petal margins, in the CHI -suppressed lines. Quantitative metabolic profiling demonstrated a significant reduction in naringenin levels across all transgenic lines, whereas, chalcone accumulation was significantly elevated in a subset of lines. The expression of CHI gene was markedly reduced in all lines carrying the CHI -RNAi construct; however, transcript levels did not differ significantly between the aberrant floral phenotypes and the control. Transcript levels of key downstream flavonoid biosynthesis pathway genes, including F3H , F3'H , F3'5'H , and DFR were significantly attenuated in CHI -silenced lines compared with wild-type controls. This concerted down‐regulation indicates that suppression of the CHI ‐catalyzed step exerts a broader repressive effect on subsequent pathway components. Despite the overall repression of flavonoid-biosynthetic genes in CHI -silenced lines, certain novel phenotypes maintained or even increased expression of specific downstream enzymes. Notably, blue-flowered transformants exhibited F3'H transcript levels comparable to wild-type control, and pink-flowered lines retained F3'5'H expression. These exceptions highlight the nuanced, tissue-specific regulatory responses triggered by CHI suppression. Conclusion Collectively, our results demonstrate that RNAi- mediated silencing of CHI gene is an effective strategy for generating novel pigmentation patterns and floral morphology in Petunia hybrida . Moreover, this work provides critical insights into how targeted manipulation of a single enzymatic step can influence the broader transcriptional network governing anthocyanin biosynthesis.

Carotenoids and carotenoid cleavage products play an important and integral role in plant development. The Decreased apical dominance1 (Dad1)/PhCCD8 gene of petunia (Petunia hybrida) encodes a hypothetical carotenoid cleavage dioxygenase (CCD) and ortholog of the MORE AXILLARY GROWTH4 (MAX4)/AtCCD8 gene. The dad1-1 mutant allele was inactivated by insertion of an unusual transposon (Dad-one transposon), and the dad1-3 allele is a revertant allele of dad1-1. Consistent with its role in producing a graft-transmissible compound that can alter branching, the Dad1/PhCCD8 gene is expressed in root and shoot tissue. This expression is upregulated in the stems of the dad1-1, dad2, and dad3 increased branching mutants, indicating feedback regulation of the gene in this tissue. However, this feedback regulation does not affect the root expression of Dad1/PhCCD8. Overexpression of Dad1/PhCCD8 in the dad1-1 mutant complemented the mutant phenotype, and RNA interference in the wild type resulted in an increased branching phenotype. Other differences in phenotype associated with the loss of Dad1/PhCCD8 function included altered timing of axillary meristem development, delayed leaf senescence, smaller flowers, reduced internode length, and reduced root growth. These data indicate that the substrate(s) and/or product(s) of the Dad1/PhCCD8 enzyme are mobile signal molecules with diverse roles in plant development.

Phosphorus and nitrogen are essential nutrient elements that are needed by plants in large amounts. The arbuscular mycorrhizal symbiosis between plants and soil fungi improves phosphorus and nitrogen acquisition under limiting conditions. On the other hand, these nutrients influence root colonization by mycorrhizal fungi and symbiotic functioning. This represents a feedback mechanism that allows plants to control the fungal symbiont depending on nutrient requirements and supply. Elevated phosphorus supply has previously been shown to exert strong inhibition of arbuscular mycorrhizal development. Here, we address to what extent inhibition by phosphorus is influenced by other nutritional pathways in the interaction between Petunia hybrida and R. irregularis. We show that phosphorus and nitrogen are the major nutritional determinants of the interaction. Interestingly, the symbiosis-promoting effect of nitrogen starvation dominantly overruled the suppressive effect of high phosphorus nutrition onto arbuscular mycorrhiza, suggesting that plants promote the symbiosis as long as they are limited by one of the two major nutrients. Our results also show that in a given pair of symbiotic partners (Petunia hybrida and R. irregularis), the entire range from mutually symbiotic to parasitic can be observed depending on the nutritional conditions. Taken together, these results reveal complex nutritional feedback mechanisms in the control of root colonization by arbuscular mycorrhizal fungi.

Petal fusion in petunia (Petunia x hybrida) results from lateral expansion of the five initially separate petal primordia, forming a ring-like primordium that determines further development. Here, we show that MAEWEST (MAW) and CHORIPETALA SUZANNE (CHSU) are required for petal and carpel fusion, as well as for lateral outgrowth of the leaf blade. Morphological and molecular analysis of maw and maw chsu double mutants suggest that polarity defects along the adaxial/abaxial axis contribute to the observed reduced lateral outgrowth of organ primordia. We show that MAW encodes a member of the WOX (WUSCHEL-related homeobox) transcription factor family and that a partly similar function is redundantly encoded by WOX1 and PRESSED FLOWER (PRS) in Arabidopsis thaliana, indicating a conserved role for MAW/WOX1/PRS genes in regulating lateral organ development. Comparison of petunia maw and Arabidopsis wox1 prs phenotypes suggests differential recruitment of WOX gene function depending on organ type and species. Our comparative data together with previous reports on WOX gene function in different species identify the WOX gene family as highly dynamic and, therefore, an attractive subject for future evo-devo studies.

Animal-mediated pollination is essential in plant reproductive biology and is often associated with pollination syndromes, sets of floral traits, such as color, scent, shape, or nectar content. Selection by pollinators is often considered a key factor in floral evolution and plant speciation. Our aim is the identification and characterization of the genetic changes that caused the evolution of divergent pollination syndromes in closely related plant species. We focus on ANTHOCYANIN2 (AN2), a well-defined myb-type transcription factor that is a major determinant of flower color variation between Petunia integrifolia and Petunia axillaris. Analysis of sequence variation in AN2 in wild P. axillaris accessions showed that loss-of-function alleles arose at least five times independently. DNA sequence analysis was complemented by functional assays for pollinator preference using genetic introgressions and transgenics. These results show that AN2 is a major determinant of pollinator attraction. Therefore, changes in a single gene cause a major shift in pollination biology and support the notion that the adaptation of a flowering plant to a new pollinator type may involve a limited number of genes of large effect. Gene identification and analysis of molecular evolution in combination with behavioral and ecological studies can ultimately unravel the evolutionary genetics of pollination syndromes.

Background The ALOG ( Arabidopsis LSH1 and Oryza G1) family of proteins, namely DUF640 (domain of unknown function 640) domain proteins, were found in land plants. Functional characterization of a few ALOG members in model plants such as Arabidopsis and rice suggested they play important regulatory roles in plant development. The information about its evolution, however, is largely limited, and there was no any report on the ALOG genes in Petunia , an important ornamental species. Results The ALOG genes were identified in four species of Petunia including P. axillaris , P. inflata , P. integrifolia, and P. exserta based on the genome and/or transcriptome databases, which were further confirmed by cloning from P. hybrida ‘W115’ (Mitchel diploid), a popular laboratorial petunia line susceptible to genetic transformation. Phylogenetic analysis indicated that Petunia ALOG genes (named as LSHs according to their closest Arabidopsis homologs) were grouped into four clades, which can be further divided into eight groups, and similar exon - intron structure and motifs are reflected in the same group. The PhLSH genes of hybrid petunia ‘W115’ were mainly derived from P. axillaris . The qPCR analysis revealed distinct spatial expression patterns among them suggesting potentially functional diversification. Moreover, over-expressing PhLSH7a and PhLSH7b in Arabidopsis uncovered their functions in the development of both vegetative and reproductive organs. Conclusions Petunia genome includes 11 ALOG genes that can be divided into eight distinct groups, and they also show different expression patterns. Among these genes, PhLSH7b and PhLSH7a play significant roles in plant growth and development, especially in fruit development. Our results provide new insight into the evolution of ALOG gene family and have laid a good foundation for the study of petunia LSH gene in the future.

Key message Site-directed mutagenesis of nitrate reductase genes using direct delivery of purified Cas9 protein preassembled with guide RNA produces mutations efficiently in Petunia   ×   hybrida protoplast system. The clustered, regularly interspaced, short palindromic repeat (CRISPR)-CRISPR associated endonuclease 9 (CRISPR/Cas9) system has been recently announced as a powerful molecular breeding tool for site-directed mutagenesis in higher plants. Here, we report a site-directed mutagenesis method targeting Petunia nitrate reductase ( NR ) gene locus. This method could create mutations efficiently using direct delivery of purified Cas9 protein and single guide RNA (sgRNA) into protoplast cells. After transient introduction of RNA-guided endonuclease (RGEN) ribonucleoproteins (RNPs) with different sgRNAs targeting NR genes, mutagenesis at the targeted loci was detected by T7E1 assay and confirmed by targeted deep sequencing. T7E1 assay showed that RGEN RNPs induced site-specific mutations at frequencies ranging from 2.4 to 21 % at four different sites (NR1, 2, 4 and 6) in the PhNR gene locus with average mutation efficiency of 14.9 ± 2.2 %. Targeted deep DNA sequencing revealed mutation rates of 5.3–17.8 % with average mutation rate of 11.5 ± 2 % at the same NR gene target sites in DNA fragments of analyzed protoplast transfectants. Further analysis from targeted deep sequencing showed that the average ratio of deletion to insertion produced collectively by the four NR-RGEN target sites (NR1, 2, 4, and 6) was about 63:37. Our results demonstrated that direct delivery of RGEN RNPs into protoplast cells of Petunia can be exploited as an efficient tool for site-directed mutagenesis of genes or genome editing in plant systems.