MIK2 is a candidate gene of the S-locus for sporophytic self-incompatibility in chicory (Cichorium intybus, Asteraceae)

The Cichorium genus offers a unique opportunity to study the sporophytic self-incompatibility (SSI) system, being composed of species characterized by highly efficient self-incompatibility (e.g., C. intybus) and complete self-compatibility (e.g., C. endivia). To this end, the chicory genome was used to map seven previously identified SSI locus-associated markers. The region containing the S-locus was therefore restricted to an similar to 4 M bp window on chromosome 5. Among the genes predicted in this region, MDIS1 INTERACTING RECEPTOR LIKE KINASE 2 (ciMIK2) was particularly promising as a candidate for SSI. Its ortholog in Arabidopsis (atMIK2) is involved in pollen-stigma recognition reactions, and its protein structure is similar to that of S-receptor kinase (SRK), a key component of the SSI system in the Brassica genus. The amplification and sequencing of MIK2 in chicory and endive accessions revealed two contrasting scenarios. In C. endivia, MIK2 was fully conserved even when comparing different botanical varieties (i.e., smooth and curly endive). In C. intybus, 387 polymorphic positions and 3 INDELs were identified when comparing accessions of different biotypes all belonging to the same botanical variety (i.e., radicchio). The polymorphism distribution throughout the gene was uneven, with hypervariable domains preferentially localized in the LRR-rich extracellular region, putatively identified as the receptor domain. The gene was hypothesized to be under positive selection, as the nonsynonymous mutations were more than double the synonymous ones (dN/dS = 2.17). An analogous situation was observed when analyzing the first 500 bp of the MIK2 promoter: no SNPs were observed among the endive samples, whereas 44 SNPs and 6 INDELs were detected among the chicory samples. Further analyses are needed to confirm the role of MIK2 in SSI and to demonstrate whether the 23 species-specific nonsynonymous SNPs in the CDS and/or the species-specific 10 bp-INDEL found in a CCAAT box region of the promoter are responsible for the contrasting sexual behaviors of chicory and endive

Not only editing: a cas-cade of CRISPR/cas-based tools for functional genomics in plants and animals

The advent of CRISPR/Cas9 technology has revolutionized genome editing, enabling the attainment of once-unimaginable goals. CRISPR/Cas’s groundbreaking attributes lie in its simplicity, versatility, universality, and independence from customized DNA-protein systems, erasing the need for specialized expertise and broadening its scope of applications. It is therefore more and more used for genome modification including the generation of mutants. Beyond such editing scopes, the recent development of novel or modified Cas-based systems has spawned an array of additional biotechnological tools, empowering both fundamental and applied research. Precisely targeting DNA or RNA sequences, the CRISPR/Cas system has been harnessed in fields as diverse as gene regulation, deepening insights into gene expression, epigenetic changes, genome spatial organization, and chromatin dynamics. Furthermore, it aids in genome imaging and sequencing, as well as effective identification and countering of viral pathogens in plants and animals. All in all, the non-editing aspect of CRISPR/Cas exhibits tremendous potential across diverse domains, including diagnostics, biotechnology, and fundamental research. This article reviews and critically evaluates the primary CRISPR/Cas-based tools developed for plants and animals, underlining their transformative impac

Condensation properties of stress granules and processing bodies are compromised in Myotonic Dystrophy Type 1

RNA regulation in mammalian cells requires complex physical compartmentalisation, using structures thought to be formed by liquid-liquid phase separation. Disruption of these structures is implicated in numerous degenerative diseases. Myotonic dystrophy type 1 (DM1) is a multi-systemic trinucleotide repeat disorder resulting from an expansion of nucleotides CTG (CTGexp) in the DNA encoding DM1 protein kinase (DMPK). The cellular hallmark of DM1 is the formation of nuclear foci that contain expanded DMPK RNA (CUGexp) (with thymine instead of uracil). We report here the deregulation of stress granules (SGs) and processing bodies (P-bodies), two cytoplasmic structures key for mRNA regulation, in cell culture models of DM1. Alterations to the rates of formation and dispersal of SGs suggest an altered ability of cells to respond to stress associated with DM1, while changes to the structure and dynamics of SGs and P-bodies suggest that a widespread alteration to the biophysical properties of cellular structures is a consequence of the presence of CUGexp RNA.</p

Clinical Applicability of Visible Light-Mediated Cross-linking for Structural Soft Tissue Reconstruction

Abstract Visible light‐mediated cross‐linking has utility for enhancing the structural capacity and shape fidelity of laboratory‐based polymers. With increased light penetration and cross‐linking speed, there is opportunity to extend future applications into clinical spheres. This study evaluated the utility of a ruthenium/sodium persulfate photocross‐linking system for increasing structural control in heterogeneous living tissues as an example, focusing on unmodified patient‐derived lipoaspirate for soft tissue reconstruction. Freshly‐isolated tissue is photocross‐linked, then the molar abundance of dityrosine bonds is measured using liquid chromatography tandem mass spectrometry and the resulting structural integrity assessed. The cell function and tissue survival of photocross‐linked grafts is evaluated ex vivo and in vivo, with tissue integration and vascularization assessed using histology and microcomputed tomography. The photocross‐linking strategy is tailorable, allowing progressive increases in the structural fidelity of lipoaspirate, as measured by a stepwise reduction in fiber diameter, increased graft porosity and reduced variation in graft resorption. There is an increase in dityrosine bond formation with increasing photoinitiator concentration, and tissue homeostasis is achieved ex vivo, with vascular cell infiltration and vessel formation in vivo. These data demonstrate the capability and applicability of photocrosslinking strategies for improving structural control in clinically‐relevant settings, potentially achieving more desirable patient outcomes using minimal manipulation in surgical procedures

The transcriptional landscape of berry skin in red and white PIWI (“Pilzwiderstandsfähig”) grapevines possessing QTLs for partial resistance to downy and powdery mildews

PIWI, from the German word Pilzwiderstandsfähig, meaning “fungus-resistant”, refers to grapevine cultivars bred for resistance to fungal pathogens such as Erysiphe necator (the causal agent of powdery mildew) and Plasmopara viticola (the causal agent of downy mildew), two major diseases in viticulture. These varieties are typically developed through traditional breeding, often crossbreeding European Vitis vinifera with American or Asian species that carry natural disease resistance. This study investigates the transcriptional profiles of exocarp tissues in mature berries from four PIWI grapevine varieties compared to their elite parental counterparts using RNA-seq analysis. We performed RNA-seq on four PIWI varieties (two red and two white) and their noble parents to identify differential gene expression patterns. Comprehensive analyses, including Differential Gene Expression (DEGs), Gene Set Enrichment Analysis (GSEA), Weighted Gene Co-expression Network Analysis (WGCNA), and tau analysis, revealed distinct gene clusters and individual genes characterizing the transcriptional landscape of PIWI varieties. Differentially expressed genes indicated significant changes in pathways related to organic acid metabolism and membrane transport, potentially contributing to enhanced resilience. WGCNA and k-means clustering highlighted co-expression modules linked to PIWI genotypes and their unique tolerance profiles. Tau analysis identified genes uniquely expressed in specific genotypes, with several already known for their defense roles. These findings offer insights into the molecular mechanisms underlying grapevine resistance and suggest promising avenues for breeding strategies to enhance disease resistance and overall grape quality in viticultur

Exploring the gene regulatory networks of WRKY family in grapevine (Vitis vinifera L.) using DAP-Seq

The recent development of regulatory genomics has raised increasing interest in plant research since transcriptional regulation of genes plays a pivotal role in many biological processes. By shedding light on the target genes of the various transcription factors (TFs), it is therefore possible to infer the influence they exert on the different molecular mechanisms. In this regard, the attention was focused on WRKYs, a family of TFs almost exclusively found in plant species. In grapevine, WRKYs are involved in several biological processes, playing a key role in berry development, hormonal balance and signalling, biotic and abiotic stresses responses, and secondary metabolites biosynthesis. In this study, DAP-Seq was used to investigate the regulatory activity of the whole WRKY family in gDNA from young leaves of Cabernet Franc. This approach allowed the definition of the WRKYs cistrome (i.e., the set of bound genes). 46 out of 59 WRKYs gave results, outlining a total number of 674,407 binding events along whole grapevine genome, of which 459,791 (68%) are localized in the perigenic region, according to its intense regulatory activity. Cistrome maps were integrated with gene centred co-expression networks based on a large transcriptomics dataset, and with the results of an ATAC-Seq. This allowed to isolate some High Confidence Targets, characterized by high degree of co-expression with the related TF and laying down in genomic regions of open chromatin. The networks generated can be used to provide a complete regulatory map of WRKY family, shedding light on its biological role in grapevine

Inativação do crescimento micelial in vitro e erradicação de Cylindrocarpon destructans por tratamento térmico.

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