Reproductive potential of the functionally female native Croatian grapevine 'Grk bijeli'

A native Croatian grapevine 'Grk bijeli', sharing a parent-offspring relationships with 'Tribidrag' (aka 'Zinfandel'), is grown exclusively on the Adriatic island of Korčula. It is one of the grape cultivars with female-only functional flowers, causing reduced fertilisation and leads to problems in grape production and wine quality. A typical cluster of 'Grk bijeli' at maturity contains fully developed seeded berries, in addition to a highly variable proportion of undersized seedless berries. The aim of this study was to identify the cause of the reduced reproduction potential of 'Grk bijeli' in order to establish a better growing environment for improved yield and grape quality. 'Grk bijeli' female gametophyte develops normaly and at maturity contains both egg and central cell, together with two synergid cells and three antipodal cells. On the other hand, 'Grk bijeli' pollen grains show developmental deviations. Specifically, most of the pollen grains undergo the first pollen mitosis and contain one vegetative cell and one generative cell, while 20 % of ungerminating grains also accomplish the second pollen mitosis, giving rise to two sperm cells and one vegetative cell. Morover, 'Grk bijeli' pollen has acolporate morphology, which prevents germination and contributes to reduced reproduction. Furthermore, fertilisation after pollination with other varieties results in various degrees of ovule abortion depending on the pollinator, revealing Croatian cultivars 'Plavac mali crni' and 'Pošip bijeli' as favourable varieties. Although this study describes a highly valuable cultivar of local importance, it also contributes to fundamental knowledge of grapevine reproductive biology and offers a strategy for improvement of wine production and oenological performance of semi-fertile varieties in general

Reduced protein secretion and glycosylation induced by ammonium stress inhibits somatic embryo development in pumpkin

Extracellular proteins and glycoproteins secreted by ammonium- or auxin-induced somatic embryogenic cultures of pumpkin were analyzed. Despite an overall similarity in developmental characteristics between these embryogenic cultures, distinct expression patterns of extracellular proteins and glycoproteins were observed. Ammonium, when supplied as the sole source of nitrogen, caused acidification of the culture medium and significantly reduced protein secretion. Buffering pH in the ammonium-containing medium restored extracellular protein secretion and glycosylation and an enhanced cell aggregation but not the development of later embryo stages. As revealed by Concavalin A (Con A) immunodetection, extracellular glycoproteins containing α-D-mannose and α-D-glucose were most abundant in proembryogenic cultures grown in a buffered ammonium-containing medium and in a medium supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D). We assume that extracellular proteins (Mr 28, 31, and 44 kDa) and Con Abinding glycoproteins (Mr 26, 30, 40, 53, and 100 kDa) found in both proembryogenic cultures may have a role during somatic embryogenesis induction. The glycan components of proteins were further characterized by affinity blotting with different lectins. Binding patterns of mannose-specific lectin from Galanthus nivalis partially correlated with those detected with Con A, whereas no signal was observed with lectins from Datura stramonium and Arachis hypogea regardless of the treatment applied. Results indicate that complex N- or O-glycans are not typical for early phases of pumpkin embryo development. The accumulation of extracellular glycoproteins with high-mannose-type glycans from 30 to 34 kDa, observed after the transfer from the ammonium- or 2,4-D-containing media into a maturation medium, appeared to be associated with development of later embryo stages. This study also revealed the presence of EP-3-like endochitinases in pumpkin embryogenic cultures, particularly in cultures grown in the buffered ammonium-containing medium, however, these proteins should be examined further

Protection of Phaseolus vulgaris L. from Herbicide 2,4-D Results from Exposing Seeds to Humic Acid

Bean (Phaseolus vulgarisL.) is the world's most significant and basic legume crop for people diet. The intense use of the herbicides such as 2,4-dichlorophenoxyacetic acid (2,4-D) in bean planting areas can lead to a number of toxicological issues. To prevent such damages, humic acid (HA) may be used to increase plant development by improving nutrient uptake and to play a protecting role against stresses by regulating the antioxidative system of plants. Hence, the aim of the current study was to investigate the impacts of HA on DNA damage levels and DNA methylation changes against 2,4-D stress in the bean. HA treatments were applied to bean seedlings, and the 2,4-D was sprayed on the three-to-four-leaf stages at 2,4-D dose. We used random amplified polymorphic DNA (RAPD) for determining the changes in DNA damage and coupled restriction enzyme digestion-random amplification (CRED-RA) for DNA methylation changes. According to results, while the genomic template stability (GTS) decreased in the 2,4-D (5, 10, 20 and 40 mg/L) treatments, this value increased comparatively in the 2,4-D applied together with HA (0, 2, 4, 6, 8 and 10 mg/L) treatments. In CRED-RA patterns, as the HA doses were increased, there was generally a decrease in polymorphism rates (DNA methylation changes) caused by 2,4-D applications. Our results have clearly demonstrated that HA has a curative effect up to a level against genotoxic and DNA methylation changes caused by 2,4-D

De novo zygotic transcription in wheat (Triticum aestivum L.) includes genes encoding small putative secreted peptides and a protein involved in proteasomal degradation

Wheat is one of the world's most important crops, and increasing grain yield is a major challenge for the future. Still, our knowledge about the molecular machineries responsible for early post-fertilization events such as zygotic reprogramming, the initial cell-specification events during embryogenesis, and the intercellular communication between the early embryo and the developing endosperm is very limited. Here, we describe the identification of de novo transcribed genes in the wheat zygote. We used wheat ovaries of defined post-fertilization stages to isolate zygotes and early embryos, and identified genes that are specifically induced in these particular stages. Importantly, we observed that some of the zygotic-induced genes encode proteins with similarity to secreted signaling peptides such as TAPETUM DETERMINANT 1 and EGG APPARATUS 1, and to MATH-BTB proteins which are known substrate-binding adaptors for the Cullin-3-based ubiquitin E3 ligase. This suggests that both cell-cell signaling and targeted proteasomal degradation may be important molecular events during zygote formation and the progression of early embryogenesis

Molecular memory of Flavescence dorée phytoplasma in recovering grapevines

Flavescence dorée (FD) is a destructive phytoplasma disease of European grapevines. Spontaneous and cultivar-dependent recovery (REC) may occur in the field in FD-infected vines starting the year following the first symptoms. However, the biological underpinnings of this process are still largely unexplored. In this study, transcriptome sequencing (RNAseq), whole-genome bisulphite sequencing (WGBS) and metabolite analysis were combined to dissect molecular and metabolic changes associated to FD and REC in leaf veins collected in the field from healthy (H), FD and REC plants of the highly susceptible Vitis vinifera 'Barbera'. Genes involved in flavonoid biosynthesis, carbohydrate metabolism and stress responses were overexpressed in FD conditions, whereas transcripts linked to hormone and stilbene metabolisms were upregulated in REC vines. Accumulation patterns of abscisic acid and stilbenoid compounds analysed in the same samples confirmed the RNAseq data. In recovery conditions, we also observed the persistence of some FD-induced expression changes concerning inhibition of photosynthetic processes and stress responses. Several differentially expressed genes tied to those pathways also underwent post-transcriptional regulation by microRNAs, as outlined by merging our transcriptomic data set with a previously conducted smallRNAseq analysis. Investigations by WGBS analysis also revealed different DNA methylation marks between REC and H leaves, occurring within the promoters of genes tied to photosynthesis and secondary metabolism. The results allowed us to advance the existence of a "molecular memory" of FDp infection, involving alterations in the DNA methylation status of REC plants potentially related to transcriptional reprogramming events, in turn triggering changes in hormonal and secondary metabolite profiles