Induction of Somatic Embryogenesis in Plants: Different Players and Focus on WUSCHEL and WUS-RELATED HOMEOBOX (WOX) Transcription Factors

In plants, other cells can express totipotency in addition to the zygote, thus resulting in embryo differentiation; this appears evident in apomictic and epiphyllous plants. According to Haberlandt's theory, all plant cells can regenerate a complete plant if the nucleus and the membrane system are intact. In fact, under in vitro conditions, ectopic embryos and adventitious shoots can develop from many organs of the mature plant body. We are beginning to understand how determination processes are regulated and how cell specialization occurs. However, we still need to unravel the mechanisms whereby a cell interprets its position, decides its fate, and communicates it to others. The induction of somatic embryogenesis might be based on a plant growth regulator signal (auxin) to determine an appropriate cellular environment and other factors, including stress and ectopic expression of embryo or meristem identity transcription factors (TFs). Still, we are far from having a complete view of the regulatory genes, their target genes, and their action hierarchy. As in animals, epigenetic reprogramming also plays an essential role in re-establishing the competence of differentiated cells to undergo somatic embryogenesis. Herein, we describe the functions of WUSCHEL-RELATED HOMEOBOX (WOX) transcription factors in regulating the differentiation-dedifferentiation cell process and in the developmental phase of in vitro regenerated adventitious structures

Erinea in the 'Ansonica' grapevine cultivar: trichome complement, histological effects and analysis of chlorophyll fluorescence in affected leaves

Grapevine leaves are usually characterized by trichomes, specialized epidermal cells. They are interesting in ampelography and also important for the plant ecological responses in biotic and abiotic interactions. In nature, the trichome development is a genetic trait but it can be modified by pests as eriophyid mites. Colomerus vitis is quite common and its economic value is sometime substantial. Here, we studied the leaf erineum induced by C. vitis on 'Ansonica' ('Inzolia'), an important grapevine cultivar characterized by a low level of leaf trichome coating. To date, the interaction between C. vitis and grape has been investigated in few pedo-climatic conditions and no data are reported in 'Ansonica'. Therefore, our objectives were: (1) the analysis, in a Tuscan environment, of the morphology and histology of trichomes in 'Ansonica' leaves unaffected or affected by C. vitis; (2) evaluation, in mature leaves, of the effects of the mite both on pigment content and chlorophyll a fluorescence parameters. 'Ansonica' was devoid of glandular trichomes but it has been established the presence of few simple trichomes strictly associated with the veins. In the erineal sectors, a dense proliferation of simple trichomes in the abaxial epidermis and the development of hyperplasia in the adaxial surface were observed. Moreover, the leaf sections in the erineal regions were thicker due to an abnormal development of the lacunar parenchyma, and trichome proliferation was also extended to interveinal regions. Leaves with erinea showed a deficient content of carotenoids, in comparison to unaffected leaves. In 'Ansonica' leaves, C. vitis induced a decrease in the steady-state operational efficiency of photosystem II associated to a reduction in photochemical quenching and an increase in non-photochemical quenching values. In leaves with erinea, the reduction of photosystem II efficiency was extended to foliar areas not directly affected by galls. The collected results highlight that 'Ansonica' is susceptible to attacks by C. vitis and in the case of widespread leaf attacks the productive damage should not be underestimated

Molecular aspects of zygotic embryogenesis in sunflower (Helianthus annuus L.): correlation of positive histone marks with HaWUS expression and putative link HaWUS/HaL1L

Main conclusion The link HaWUS/HaL1L, the opposite transcriptional behavior, and the decrease/increase in positive histone marks bond to both genes suggest an inhibitory effect of WUS on HaL1L in sunflower zygotic embryos. In Arabidopsis, a group of transcription factors implicated in the earliest events of embryogenesis is the WUSCHELRELATED HOMEOBOX (WOX) protein family including WUSCHEL (WUS) and other 14 WOX protein, some of which contain a conserved WUS-box domain in addition to the homeodomain. WUS transcripts appear very early in embryogenesis, at the 16-cell embryo stage, but gradually become restricted to the center of the developing shoot apical meristem (SAM) primordium and continues to be expressed in cells of the niche/organizing center of SAM and floral meristems to maintain stem cell population. Moreover, WUS has decisive roles in the embryonic program presumably promoting the vegetative-to-embryonic transition and/or maintaining the identity of the embryonic stem cells. However, data on the direct interaction between WUS and key genes for seed development (as LEC1 and L1L) are not collected. The novelty of this report consists in the characterization of Helianthus annuus WUS (HaWUS) gene and in its analysis regarding the pattern of the methylated lysine 4 (K4) of the Histone H3 and of the acetylated histone H3 during the zygotic embryo development. Also, a parallel investigation was performed for HaL1L gene since two copies of the WUS-binding site (WUSATA), previously identified on HaL1L nucleotide sequence, were able to be bound by the HaWUS recombinant protein suggesting a not described effect of HaWUS on HaL1L transcription

A CYCLOIDEA-like gene mutation in sunflower determines an unusual floret type able to produce filled achenes at the periphery of the pseudanthium

The pseudanthium of sunflower (Helianthus annuus L.) consists of two floret types: zygomorphic sterile ray florets and actinomorphic hermaphrodite disc florets. In the tubular ray flower (turf) mutant, the loss-of-function of a CYCLOIDEA (CYC) gene generates hermaphrodite tubular-like ray florets that replace the normal sterile ray florets. We evaluated whether tubular-like ray florets have a multifaceted set of floral traits and the presence of heteromorphic seeds in the turf inflorescence. During early stages of floral ontogeny, primordia of both tubular-like ray florets and typical ray florets displayed a comparable shape. In contrast, during later stages of development, the form of tubular-like ray floret primordia was most similar to disc floret primordia. In mature tubular-like ray florets, corolla and ovary had both ray and disc floret characteristics but also displayed distinct identity traits. In open-pollinated tubular-like ray florets, the seed set was low, but a noteworthy increase of filled achenes was obtained by hand pollination. Wild type ray achenes were always empty. Embryos of tubular-like ray florets were shorter and lighter than the embryos of disc florets but able to produce fertile plants. In conclusion, the different identity characteristics combined in tubular-like ray florets of the mutant evolved a capitulum type not described in the genus Helianthus

Genotipizzazione delle accessioni della collezione ‘Vitiarium’

The 231 accessions of the most famous grapevine collection in Tuscany, the ‘Vitiarium’, have been genotyped analysing the polymorphism of 14 SSR loci. The analysis reveal 130 varieties belong to 28 international and 30 Italian varieties, 30 Tuscan local varieties and 42 unique genotypes, exclusive of the ‘Vitiarium’. Several synonyms, homonyms and misnomer have been identified. All data will be uploaded in the Italian Vitis Database

Surgical site infection after gastrointestinal surgery in high-income, middle-income, and low-income countries: a prospective, international, multicentre cohort study

Background: Surgical site infection (SSI) is one of the most common infections associated with health care, but its importance as a global health priority is not fully understood. We quantified the burden of SSI after gastrointestinal surgery in countries in all parts of the world. Methods: This international, prospective, multicentre cohort study included consecutive patients undergoing elective or emergency gastrointestinal resection within 2-week time periods at any health-care facility in any country. Countries with participating centres were stratified into high-income, middle-income, and low-income groups according to the UN's Human Development Index (HDI). Data variables from the GlobalSurg 1 study and other studies that have been found to affect the likelihood of SSI were entered into risk adjustment models. The primary outcome measure was the 30-day SSI incidence (defined by US Centers for Disease Control and Prevention criteria for superficial and deep incisional SSI). Relationships with explanatory variables were examined using Bayesian multilevel logistic regression models. This trial is registered with ClinicalTrials.gov, number NCT02662231. Findings: Between Jan 4, 2016, and July 31, 2016, 13 265 records were submitted for analysis. 12 539 patients from 343 hospitals in 66 countries were included. 7339 (58·5%) patient were from high-HDI countries (193 hospitals in 30 countries), 3918 (31·2%) patients were from middle-HDI countries (82 hospitals in 18 countries), and 1282 (10·2%) patients were from low-HDI countries (68 hospitals in 18 countries). In total, 1538 (12·3%) patients had SSI within 30 days of surgery. The incidence of SSI varied between countries with high (691 [9·4%] of 7339 patients), middle (549 [14·0%] of 3918 patients), and low (298 [23·2%] of 1282) HDI (p < 0·001). The highest SSI incidence in each HDI group was after dirty surgery (102 [17·8%] of 574 patients in high-HDI countries; 74 [31·4%] of 236 patients in middle-HDI countries; 72 [39·8%] of 181 patients in low-HDI countries). Following risk factor adjustment, patients in low-HDI countries were at greatest risk of SSI (adjusted odds ratio 1·60, 95% credible interval 1·05–2·37; p=0·030). 132 (21·6%) of 610 patients with an SSI and a microbiology culture result had an infection that was resistant to the prophylactic antibiotic used. Resistant infections were detected in 49 (16·6%) of 295 patients in high-HDI countries, in 37 (19·8%) of 187 patients in middle-HDI countries, and in 46 (35·9%) of 128 patients in low-HDI countries (p < 0·001). Interpretation: Countries with a low HDI carry a disproportionately greater burden of SSI than countries with a middle or high HDI and might have higher rates of antibiotic resistance. In view of WHO recommendations on SSI prevention that highlight the absence of high-quality interventional research, urgent, pragmatic, randomised trials based in LMICs are needed to assess measures aiming to reduce this preventable complication

CYCLOIDEA 2 Clade Genes: Key Players in the Control of Floral Symmetry, Inflorescence Architecture, and Reproductive Organ Development

Undoubted lines of evidence point out that members of CYCLOIDEA (CYC) 2 clade are essential players to control flower symmetry and, amusingly, also are determinants of capitula architecture (pseudanthium). In several species, CYC-like genes influence the androecium patterning, but to date, the function of these genes in the development of gynoecium organs is less clear. In this review, we first reported details about floral symmetry and an overview of genes and molecular mechanisms regulating the development of zygomorphism in different angiosperm lineages (e.g., basal and core eudicots and monocots). Then, we paid emphasis on the role of CYC-like genes in the development of heterogamous inflorescence of sunflower as well as other Asteraceae and some species within the Dipsacaceae family. Helianthus annuus is particularly attractive because it represents a useful model to study the role of CYC-like genes on shaping floral corolla as well as the differentiation of reproductive organs in different flowers of pseudanthia. A special attention was reserved to inflorescence morphology mutants of sunflower (i.e., Chrysanthemoids2 and tubular ray flower) because they provide useful information on the role of CYC-like genes in the radiate capitulum evolution. Finally, we discuss data from literature to suggest that CYC-like genes are also co-opted to regulate stamen and carpel differentiation likely throughout their interaction with the cell cycle and flower organ identity genes. The recruitment of reproductive organs in ray flowers also supports the phylogenetic origin of a radiate inflorescence of sunflower from a discoid capitulum and suggests that in sterile zygomorphic ray flower primordia the latent identity to differentiate both microsporangium and macrosporangium was conserved

Usual and unusual development of the dicot leaf: Involvement of transcription factors and hormones

Morphological diversity exhibited by higher plants is essentially related to the tremendous variation of leaf shape. With few exceptions, leaf primordia are initiated postembryonically at the flanks of a group of undifferentiated and proliferative cells within the shoot apical meristem (SAM) in characteristic position for the species and in a regular phyllotactic sequence. Auxin is critical for this process, because genes involved in auxin biosynthesis, transport, and signaling are required for leaf initiation. Down-regulation of transcription factors (TFs) and cytokinins are also involved in the light-dependent leaf initiation pathway. Furthermore, mechanical stresses in SAM determine the direction of cell division and profoundly influence leaf initiation suggesting a link between physical forces, gene regulatory networks and biochemical gradients. After the leaf is initiated, its further growth depends on cell division and cell expansion. Temporal and spatial regulation of these processes determines the size and the shape of the leaf, as well as the internal structure. A complex array of intrinsic signals, including phytohormones and TFs control the appropriate cell proliferation and differentiation to elaborate the final shape and complexity of the leaf. Here, we highlight the main determinants involved in leaf initiation, epidermal patterning, and elaboration of lamina shape to generate small marginal serrations, more deep lobes or a dissected compound leaf. We also outline recent advances in our knowledge of regulatory networks involved with the unusual pattern of leaf development in epiphyllous plants as well as leaf morphology aberrations, such as galls after pathogenic attacks of pests

Isolation and molecular analysis of two R2R3-MYB genes from sunflower (Helianthus annuus)

Abstract MYB factors comprise one of the largest family of transcription factors (TFs) characterised by a highly conserved DNA-binding domain: the MYB domain. From sunflower genome, we have isolated two R2R3-type MYB TFs (Ha-R2R3-MYB1 and Ha-R2R3-MYB2) that show homology with the functional domain of most R2R3-MYB proteins of other species. The R2 (53 amino acids) and R3 (51 amino acids) motifs of sunflower MYB TFs contain typical amino acids, including a series of highly conserved tryptophan residues, which play a key role in sequence-specific DNA binding. In the MYB domain, both genes (Ha-R2R3-MYB1 and Ha-R2R3-MYB2) exhibit the highly conserved splicing arrangement of three exons and two introns. Using in situ hybridization, a weak-level of Ha-R2R3-MYB1 transcription was uniformly spread in shoot apical meristem (SAM) as well as in axillary meristem (AM). By contrast, Ha-R2R3-MYB2 transcription was strongly restricted to a small domain within the boundary zone separating the SAM and the leaf primordia, suggesting that Ha-R2R3-MYB2 may be involved in an early step of AM development

Molecular cloning, phylogenetic analysis, and expression patterns of LATERAL SUPPRESSOR-LIKE and REGULATOR OF AXILLARY MERISTEM FORMATION-LIKE genes in sunflower (Helianthus annuus L.)

The wild sunflower (Helianthus annuus) plants develop a highly branched form with numerous small flowering heads. The origin of a no branched sunflower, producing a single large head, has been a key event in the domestication process of this species. The interaction between hormonal factors and several genes organizes the initiation and outgrowth of axillary meristems (AMs). From sunflower, we have isolated two genes putatively involved in this process, LATERAL SUPPRESSOR (LS)-LIKE (Ha-LSL) and REGULATOR OF AXILLARY MERISTEM FORMATION (ROX)-LIKE (Ha-ROXL), encoding for a GRAS and a bHLH transcription factor (TF), respectively. Typical amino acid residues and phylogenetic analyses suggest that Ha-LSL and Ha-ROXL are the orthologs of the branching regulator LS and ROX/LAX1, involved in the growth habit of both dicot and monocot species. qRT-PCR analyses revealed a high accumulation of Ha-LSL transcripts in roots, vegetative shoots, and inflorescence shoots. By contrast, in internodal stems and young leaves, a lower amount of Ha-LSL transcripts was observed. A comparison of transcription patterns between Ha-LSL and Ha-ROXL revealed some analogies but also remarkable differences; in fact, the gene Ha-ROXL displayed a low expression level in all organs analyzed. In situ hybridization (ISH) analysis showed that Ha-ROXL transcription was strongly restricted to a small domain within the boundary zone separating the shoot apical meristem (SAM) and the leaf primordia and in restricted regions of the inflorescence meristem, beforehand the separation of floral bracts from disc flower primordia. These results suggested that Ha-ROXL may be involved to establish a cell niche for the initiation of AMs as well as flower primordia. The accumulation of Ha-LSL transcripts was not restricted to the boundary zones in vegetative and inflorescence shoots, but the mRNA activity was expanded in other cellular domains of primary shoot apical meristem as well as AMs. In addition, Ha-LSL transcript accumulation was also detected in leaves and floral primordia at early stages of development. These results were corroborated by qRT-PCR analyses that evidenced high levels of Ha-LSL transcripts in very young leaves and disc flowers, suggesting a role of Ha-LSL for the early outgrowth of lateral primordia