Biology subjects

Soares, E.

Publications and source records attributed to Soares, E..

5 recordsLinked to original sources

bioRxiv2023

Mutation of the VInv 5' UTR regulatory region reduces acrylamide levels in processed potato to reach EU food-safety standards

Protein-coding regions of genes have so far been the preferred targets for trait improvement using CRISPR-Cas9 genome editing in plants. Alteration of the reading frame can result in termination of transcription or translation, hence, loss of function of the encoded protein. 5 UTR sequences also represent a practical target region to alter gene expression and protein abundance. However, editing of 5 UTRs has so far been scarcely used to engineer trait in crop plants. Here, we demonstrate that insertion of a single adenine nucleotide mediated by Cas9 in the 5 UTR region of two alleles from the Vacuolar invertase gene (VInv) of the tetraploid potato variety Lady Rosetta (LaRo) is sufficient to substantially lower the content of reducing sugars in the potato under cold-storage conditions. Moreover, the acrylamide content generated during processing of the edited potato lines into crisps was more than three folds lower than the current EU-regulated maximum level of 750 g/kg in crisps. This gene-editing approach represents a durable strategy to improve food safety of potatoes in varieties widely preferred by the consumers and the industry.

plant biology↗

bioRxiv2021

A single cell transcriptome atlas reveals the heterogeneity of the healthy human cornea and identifies novel markers of the corneal limbus and stroma

The cornea is the clear window that lets light into the eye. It is composed of five layers: epithelium, Bowmans layer, stroma, Descemets membrane and endothelium. The maintenance of its structure and transparency are determined by the functions of the different cell types populating each layer. Attempts to regenerate corneal tissue and understand disease conditions requires knowledge of how cell profiles vary across this heterogeneous tissue. We performed a single cell transcriptomic profiling of 19,472 cells isolated from eight healthy donor corneas. Our analysis delineates the heterogeneity of the corneal layers by identifying cell populations and revealing cell states that contribute in preserving corneal homeostasis. We identified that the expression of CAV1, CXCL14, HOMER3 and CPVL were exclusive to the corneal epithelial limbal stem cell niche, CKS2, STMN1 and UBE2C were exclusively expressed in highly proliferative transit amplifying cells, and NNMT was exclusively expressed by stromal keratocytes. Overall, this research provides a basis to improve current primary cell expansion protocols, for future profiling of corneal disease states, to help guide pluripotent stem cells into different corneal lineages, and to understand how engineered substrates affect corneal cells to improve regenerative therapies.

cell biology↗

bioRxiv2020

Differential DNA methylation in the Vinv promoter region controls Cold Induced Sweetening in potato

Control of potato sprouting is important to ensure constant supply of high-quality potato to the industry. Efficient control of sprouting can be achieved by chemical treatment or cold temperature. Recent bans on anti-sprouting molecules are prompting the use of cold storage in the potato value chain. Unfortunately, storage of potato at low temperatures is associated with cold induced sweetening (CIS) due to the induction of the vacuolar invertase gene under low temperatures. Because CIS is associated with the production of the potentially carcinogenic acrylamide in processed potatoes, concise knowledge on the regulatory mechanisms controlling the CIS-phenotype in potatoes is expected to help pave the way for the production of CIS-resistant potato varieties. Here, we dissect the promoters of the Vacuolar invertase (Vinv) genes from CIS-susceptible and CIS-resistant varieties to investigate their implication in CIS-phenotype determination. Using bisulfite sequencing and CRISPR-dCas9-DRM2-mediated de novo DNA methylation, we show that the CIS-resistant phenotype of Verdi, is in part due to hypermethylation of its Vinv promoter, more specifically in the 1.0-1.7kb region. Those findings open new perspectives to engineer CIS-resistant potatoes by genome and epigenome modifications.

plant biology↗

bioRxiv2020

Asparagine accumulation in chicory storage roots is controlled by translocation and feedback regulation of asparagine biosynthesis in leaves

O_LIThe presence of acrylamide (AA), a potentially carcinogenic and neurotoxic compound, in food has become a major concern for public health. AA in plant-derived food mainly arises from the reaction of the amino acid asparagine (Asn) and reducing sugars during processing of foodstuffs at high temperature. C_LIO_LIUsing a selection of genotypes from the chicory germplasm we performed Asn measurements in storage roots and leaves to identify genotypes contrasting for Asn accumulation. We combined molecular analysis and grafting experiments to show that leaf to root translocation controls asparagine biosynthesis and accumulation in chicory storage roots. C_LIO_LIWe could demonstrate that Asn accumulation in storage roots depends on Asn biosynthesis and transport from the leaf, and that a negative feedback loop by Asn on CiASN1 expression impacts Asn biosynthesis in leaves. C_LIO_LIOur results provide a new model for asparagine biosynthesis in root crop species and highlight the importance of characterizing and manipulating asparagine transport to reduce AA content in processed plant-based foodstuffs. C_LI

plant biology↗

bioRxiv2018

Single-cell RNA-seq identifies a reversible epithelial-mesenchymal transition in abnormally specified epithelia of p63 EEC syndrome

Mutations in transcription factor p63 are associated with developmental disorders that manifest defects in stratified epithelia including the epidermis. The underlying cellular and molecular mechanism is however not yet understood. We established an epidermal commitment model using human induced pluripotent stem cells (iPSCs) and characterized differentiation defects of iPSCs derived from ectrodactyly, ectodermal dysplasia, and cleft lip/palate (EEC) syndrome patients carrying p63 mutations. Transcriptome analyses revealed distinct step-wise cell fate transitions during epidermal commitment; from multipotent simple epithelium to basal stratified epithelia, and ultimately to the mature epidermal fate. Differentiation defects of EEC iPSCs caused by mutant p63 occurred during the specification switch from the simple epithelium to the basal stratified epithelial fate. Single-cell transcriptome and pseudotime analyses identified signatures of embryonic epithelial-mesenchymal transition (EMT) associated with the deviated commitment route of EEC iPSCs. Repressing mesodermal activation reversed the EMT and enhanced epidermal commitment. Our findings demonstrate that p63 is required for specification of stratified epithelia, probably by repressing embryonic EMT during epidermal commitment. This study provides insights into disease mechanisms underlying stratified epithelial defects caused by p63 mutations and suggests potential therapeutic strategies for the disease.\n\nSignificance statementMutations in p63 cause several developmental disorders with defects of epithelial related organs and tissues including the epidermis. Our study is to dissect the unknown cellular and molecular pathomechanism. We utilized human induced pluripotent stem cells (iPSCs) derived from ectrodactyly, ectodermal dysplasia, and cleft lip/palate (EEC) syndrome patients carrying p63 mutations and studied transcriptome changes during differentiation of these cells to epidermal cells. Our analyses showed that the specification of the proper epithelial cell fate was affected by p63 EEC mutations, with an abnormal embryonic epithelial-mesenchymal transition (EMT). Repressing mesodermal activation reversed the EMT and enhanced epidermal commitment. This study provides insights into disease mechanisms associated with p63 mutations and suggests potential therapeutic strategies.

developmental biology↗