Characterization of the AGR2-NPM3 axis uncovers the AGR2 involvement in PD-L1 regulation in colorectal cancer.

Despite extensive research, the molecular role of AGR2 in the progression and metastasis of colorectal cancer (CRC) has not been fully characterized. We used quantitative mass spectrometry (SWATH MS) to identify differentially expressed proteins in paired CRC cell models of the SW480 and SW620 cell lines in response to AGR2 protein level manipulation. Relying on the results from SWATH MS and subsequent immunochemical validation, we selected NMP3 as the top candidate protein associated with AGR2 in CRC tumour cells in our screen. RT‒qPCR and immunochemical analysis confirmed the involvement of AGR2-mediated regulation of NPM3 at the transcriptional and posttranscriptional levels. Since PD-L1 is a constituent of the NPM3 regulatory axis, we aimed to correlate the changes in PD-L1 to the differential expression of AGR2 in our cell models. We found that AGR2 positively regulates PD-L1 levels in both SW480 and SW620 cell lines; additionally, several different CRC patient transcriptome cohorts confirmed the association of AGR2 with PD-L1. Our work reveals a new AGR2-NPM3 regulatory axis and the involvement of AGR2 in the regulation of PD-L1, which paves the way for the association of AGR2 with immune evasion in CRC cells.

Eight hydroxyproline-O-galactosyltransferases play essential roles in female reproductive development.

In angiosperms, ovules give rise to seeds upon fertilization. Thus, seed formation is dependent on both successful ovule development and tightly controlled communication between female and male gametophytes. During establishment of these interactions, cell walls play a pivotal role, especially arabinogalactan-proteins (AGPs). AGPs are highly glycosylated proteins decorated by arabinogalactan side chains, representing 90 % of the AGP molecule. AGP glycosylation is initiated by a reaction catalysed by hydroxyproline-O-galactosyltransferases (Hyp-GALTs), specifically eight of them (GALT2-9), which add the first galactose to Hyp residues. Five Hyp-GALTs (GALT2, 5, 7, 8 and 9) were previously described as essential for AGP functions in pollen and ovule development, pollen-pistil interactions, and seed morphology. In the present work, a higher order Hyp-GALT mutant (23456789) was studied, with a high degree of under-glycosylated AGPs, to gain deeper insight into the crucial roles of these eight enzymes in female reproductive tissues. Notably, the 23456789 mutant demonstrated a high quantity of unfertilized ovules, displaying abnormal callose accumulation both at the micropylar region and, sometimes, throughout the entire embryo sac. Additionally, this mutant displayed ovules with abnormal embryo sacs, had a disrupted spatiotemporal distribution of AGPs in female reproductive tissues, and showed abnormal seed and embryo development, concomitant with a reduction in AGP-GlcA levels. This study revealed that at least three more enzymes exhibit Hyp-O-GALT activity in Arabidopsis (GALT3, 4 and 6), and reinforces the crucial importance of AGP carbohydrates in carrying out the biological functions of AGPs during plant reproduction.

AGR2 facilitates teratoma progression by regulating glycolysis via the AnXA2/EGFR axis.

Anterior gradient-2 (AGR2) is highly expressed in several tumors and plays an important role in tumor development. However, the biological function of AGR2 in teratomas has not yet been thoroughly studied. In this study, AGR2 was found to be upregulated in teratoma tissues and in human testicular teratoma cell lines by Western blotting and qRT-PCR assays. A DNA Methylation-Specific PCR assay demonstrated that AGR2 upregulation resulted from hypomethylated AGR2 in teratoma cells. NCC-IT and NT2-D1 cells were transfected with pcDNA-AGR2 or sh-AGR2 to obtain AGR2-overexpressed or -silenced cells, and cell proliferation, invasion and glycolysis were determined using CCK-8, 5-ethynyl-2'-deoxyuridine (EdU), Transwell assays, and commercial kits. The results revealed that overexpression of AGR2 promoted teratoma cell proliferation and invasion and elevated glycolysis levels evidencing by the increase in lactate secretion, glucose consumption, ATP levels and the expression of glycolysis-related proteins, while knockdown of AGR2 showed the opposite results. The interactions between AGR2 and annexin A2 (AnXA2), as well as between AnXA2 and epidermal growth factor receptor (EGFR) were verified by co-immunoprecipitation assay. Mechanistic studies revealed that AGR2 interacts with AnXA2 and increases the level of AnXA2 to recruit more AnXA2 to EGFR, there by promoting EGFR expression. A series of rescue experiments showed that knockdown of AnXA2 or EGFR weakened the promotional effects of AGR2 overexpression on the proliferation, invasion, and glycolysis of teratoma cells. Finally, tumorigenicity assays were performed using NT2-D1 cells stably transfected with either LV-NC-shRNA or LV-shAGR2. The results showed that AGR2 knockdown significantly inhibited teratoma tumor growth in vivo. In conclusion, our data suggested that AGR2 facilitates glycolysis in teratomas through promoting EGFR expression by interacting with AnXA2, thereby promoting teratoma cells proliferation and invasion.

Organoleptic modulation functions and physiochemical characteristics of mannoproteins: Possible correlations and precise applications in modulating color evolution and orthonasal perception of wines.

Mannoproteins have traditionally been recognized as effective wine organoleptic modulators, however, ambiguous understanding of the relationship between their organoleptic functions and physiochemical characteristics often lead to inappropriate application in winemaking. To reveal the possible role the physiochemical characteristics of mannoproteins play in modulating wine color and aroma properties, three water-soluble mannoproteins (MP1, MP2, MP3) with different physiochemical characteristics have been prepared, and accelerated red wine aging, malvidin pigments formation experiments, accelerated aroma release experiments have been designed to observe their organoleptic modulating functions in this research. Results suggest that the phenolic/chromatic stability of red wines could be enhanced by MP3, probably due to its low steric hindrance potential, high reactivity, and good hydro-alcoholic stability conferred by its high Mannan/Glucan ratio (8.68), abundant hydrophobic/hydrophilic amino acids (65.29 % of total protein), and low/medium molecular weight level (30.71-57.77 kDa), respectively, which protected the phenolic compounds and promoted the formation of pyranoanthocyanins. Mannoproteins could modulate the volatility of aroma compounds by expelling or retention effects, which depended on the duration of mannoprotein application (the expelling effect was firstly observed possibly because of the significant adsorption of free H2O by MPs) and the types of mannoproteins. MP1 and MP2 were prone to retain and expel aroma compounds, respectively, probably due to their medium/high molecular weight levels (60.48-135.39 kDa) that conferred abundant interacting sites, and the high proportion of hydrophobic and hydrophilic components in MP1 (97.71 % polysaccharides of total mannoprotein, 34.58 % hydrophobic amino acids of total protein) and MP2 (97.96 % polysaccharides of total mannoprotein, 28.36 % hydrophobic amino acids of total protein) guaranteed a relatively higher interacting frequency with aroma compounds and free H2O molecules, respectively.

Systems genetic analysis of lignin biosynthesis in Populus tremula.

The genetic control underlying natural variation in lignin content and composition in trees is not fully understood. We performed a systems genetic analysis to uncover the genetic regulation of lignin biosynthesis in a natural 'SwAsp' population of aspen (Populus tremula) trees. We analyzed gene expression by RNA sequencing (RNA-seq) in differentiating xylem tissues, and lignin content and composition using Pyrolysis-GC-MS in mature wood of 268 trees from 99 genotypes. Abundant variation was observed for lignin content and composition, and genome-wide association study identified proteins in the pentose phosphate pathway and arabinogalactan protein glycosylation among the top-ranked genes that are associated with these traits. Variation in gene expression and the associated genetic polymorphism was revealed through the identification of 312 705 local and 292 003 distant expression quantitative trait loci (eQTL). A co-expression network analysis suggested modularization of lignin biosynthesis and novel functions for the lignin-biosynthetic CINNAMYL ALCOHOL DEHYDROGENASE 2 and CAFFEOYL-CoA O-METHYLTRANSFERASE 3. PHENYLALANINE AMMONIA LYASE 3 was co-expressed with HOMEOBOX PROTEIN 5 (HB5), and the role of HB5 in stimulating lignification was demonstrated in transgenic trees. The systems genetic approach allowed linking natural variation in lignin biosynthesis to trees´ responses to external cues such as mechanical stimulus and nutrient availability.

AGR2: The Covert Driver and New Dawn of Hepatobiliary and Pancreatic Cancer Treatment.

The anterior gradient protein 2 (AGR2) plays a crucial role in facilitating the formation of protein disulfide bonds within the endoplasmic reticulum (ER). Research suggests that AGR2 can function as an oncogene, with its heightened expression linked to the advancement of hepatobiliary and pancreatic cancers through invasion and metastasis. Notably, AGR2 not only serves as a pro-oncogenic agent but also as a downstream targeting protein, indirectly fostering cancer progression. This comprehensive review delves into the established functions and expression patterns of AGR2, emphasizing its pivotal role in cancer progression, particularly in hepatobiliary and pancreatic malignancies. Furthermore, AGR2 emerges as a potential cancer prognostic marker and a promising target for immunotherapy, offering novel avenues for the treatment of hepatobiliary and pancreatic cancers and enhancing patient outcomes.

miR-200c-3p regulates α4 integrin-mediated T cell adhesion and migration.

A microRNA miR-200c-3p is a regulator of epithelial-mesenchymal transition to control adhesion and migration of epithelial and mesenchymal cells. However, little is known about whether miR-200c-3p affects lymphocyte adhesion and migration mediated by integrins. Using TK-1 (a T lymphoblast cell) as a model of T cell, here we show that repressed expression of miR-200c-3p upregulated α4 integrin-mediated adhesion to and migration across mucosal addressin cell adhesion molecule-1 (MAdCAM-1). Conversely, overexpression of miR-200c-3p downregulated α4 integrin-mediated adhesion and migration. Unlike in epithelial cells, miR-200c-3p did not target talin, a conformation activator of integrin, but, targeted E26-transformation-specific sequence 1 (ETS1), a transcriptional activator of α4 integrin, in T cells. Treatment of the miR-200c-3p-low-expressing TK-1 cells that possessed elevated α4 integrin with ETS1 small interfering RNA (siRNA) resulted in the reversion of the α4 integrin expression, supporting that ETS1 is a target of miR-200c-3p. A potential proinflammatory immune-modulator retinoic acid (RA) treatment of TK-1 cells elicited a significant reduction of miR-200c-3p and simultaneously a marked increase in ETS1 and α4 integrin expression. An anti-inflammatory cytokine TGF-ß1 treatment elevated miR-200c-3p, thereby downregulating ETS1 and α4 integrin expression. These results suggest that miR-200c-3p is an important regulator of α4 integrin expression and functions and may be controlled by RA and TGF-ß1 in an opposite way. Overexpression of miR-200c-3p could be a novel therapeutic option for treatment of gut inflammation through suppressing α4 integrin-mediated T cell migration.

Do Arabinogalactan Proteins Occur in the Transfer Cells of Utricularia dichotoma?

Species in the genus Utricularia are carnivorous plants that prey on invertebrates using traps of leaf origin. The traps are equipped with numerous different glandular trichomes. Trichomes (quadrifids) produce digestive enzymes and absorb the products of prey digestion. The main aim of this study was to determine whether arabinogalactan proteins (AGPs) occur in the cell wall ingrowths in the quadrifid cells. Antibodies (JIM8, JIM13, JIM14, MAC207, and JIM4) that act against various groups of AGPs were used. AGP localization was determined using immunohistochemistry techniques and immunogold labeling. AGPs localized with the JIM13, JIM8, and JIM14 epitopes occurred in wall ingrowths of the pedestal cell, which may be related to the fact that AGPs regulate the formation of wall ingrowths but also, due to the patterning of the cell wall structure, affect symplastic transport. The presence of AGPs in the cell wall of terminal cells may be related to the presence of wall ingrowths, but processes also involve vesicle trafficking and membrane recycling, in which these proteins participate.

Effects of steaming on physicochemical and emulsification properties of gum arabic.

Gum arabic finds extensive application and typically undergoes sterilization prior to utilization in the food industry. This study explored the impact of steam sterilization temperature and duration on the physicochemical and emulsification characteristics of gum arabic, accompanied by proposed mechanisms elucidating observed effects. The results showed that when gum arabic was treated with high temperature sterilization (110 °C âˆ¼ 140 °C), the emulsion prepared turned unstable. The interfacial tension decreased from 8.26 mN/m to 6.77 mN/m after sterilization, while the elastic modulus decreased from 23.65 mN/m to 16.16 mN/m. Moreover, the circular dichroic chromatographic results indicated that the arabinogalactan protein (AGP) structure of gum arabic was more relaxed after high temperature treatment with ß-sheets content decreased from 36.2 % to 29.8 % and random coil content increased from 41.3 % to 51.8 %. Quartz crystal microbalance with dissipation (QCM-D) results demonstrated that emulsion surface film thickness and toughness decreased after sterilization treatment of gum arabic. The study indicates that high temperature sterilization may change protein structure in gum arabic and reduce the stability of prepared emulsions.

Elucidating Korean meadowsweet (Filipendula glaberrima Nakai)-derived arabinogalactan protein-induced macrophage activation and its associated mechanism of action.

This study aimed to confirm macrophage-stimulatory component from Korean meadowsweet (Filipendula glaberrima; FG) and characterize its compositional and structural properties. FG-CWH, prepared via cool-water extraction and ethanol precipitation, induced the highest secretion of NO (6.0-8.0 µM), TNF-α (8.7-9.5 ng/mL), and IL-6 (1.0-5.7 ng/mL) compared to other samples at 0.4-10 µg/mL in RAW 264.7 cells. Analytical results revealed that FG-CWH is a high-molecular-weight component with an average molecular weight of 220 kDa, constituting a polysaccharide-protein mixture. Chemical and enzymatic treatment of FG-CWH indicated its primary composition as arabinogalactan protein (AGP)-rich glycoprotein, with activity likely associated with the chemical and structural characteristics of AGP. FG-CWH treatment resulted in significant and concentration-dependent increases in iNOS (20.0-29.6 folds), TNFα (10.6-18.6 folds) and IL6 (10.9-155.6 folds) gene expression, as well as the secretion of NO (5.3-6.3 µM), TNF-α (35.4-44.3 ng/mL), and IL-6 (4.1-8.4 ng/mL) secretion, even at a reduced concentration range of 125-500 ng/mL, compared to the negative control group. Immunoblotting analysis indicated FG-CWH-induced macrophage stimulation significantly associated with the activation of MAPK (ERK, JNK, and p38) and NF-κB (p65 and IκBα). These findings can serve as valuable groundwork for developing FG-derived AGP as novel functional ingredients to enhance human immunity.

Cell Wall Microdomains in the External Glands of Utricularia dichotoma Traps.

The genus Utricularia (bladderworts) species are carnivorous plants that prey on invertebrates using traps with a high-speed suction mechanism. The outer trap surface is lined by dome-shaped glands responsible for secreting water in active traps. In terminal cells of these glands, the outer wall is differentiated into several layers, and even cell wall ingrowths are covered by new cell wall layers. Due to changes in the cell wall, these glands are excellent models for studying the specialization of cell walls (microdomains). The main aim of this study was to check if different cell wall layers have a different composition. Antibodies against arabinogalactan proteins (AGPs) were used, including JIM8, JIM13, JIM14, MAC207, and JIM4. The localization of the examined compounds was determined using immunohistochemistry techniques and immunogold labeling. Differences in composition were found between the primary cell wall and the cell secondary wall in terminal gland cells. The outermost layer of the cell wall of the terminal cell, which was cuticularized, was devoid of AGPs (JIM8, JIM14). In contrast, the secondary cell wall in terminal cells was rich in AGPs. AGPs localized with the JIM13, JIM8, and JIM14 epitopes occurred in wall ingrowths of pedestal cells. Our research supports the hypothesis of water secretion by the external glands.

Low oxygen environment effect on the tomato cell wall composition during the fruit ripening process.

BACKGROUND: Oxygen concentration is a key characteristic of the fruit storage environment determining shelf life and fruit quality. The aim of the work was to identify cell wall components that are related to the response to low oxygen conditions in fruit and to determine the effects of such conditions on the ripening process. Tomato (Solanum lycopersicum) fruits at different stages of the ripening process were stored in an anoxic and hypoxic environment, at 0% and 5% oxygen concentrations, respectively. We used comprehensive and comparative methods: from microscopic immunolabelling and estimation of enzymatic activities to detailed molecular approaches. Changes in the composition of extensin, arabinogalactan proteins, rhamnogalacturonan-I, low methyl-esterified homogalacturonan, and high methyl-esterified homogalacturonan were analysed. RESULTS: In-depth molecular analyses showed that low oxygen stress affected the cell wall composition, i.e. changes in protein content, a significantly modified in situ distribution of low methyl-esterified homogalacturonan, appearance of callose deposits, disturbed native activities of ß-1,3-glucanase, endo-ß-1,4-glucanase, and guaiacol peroxidase (GPX), and disruptions in molecular parameters of single cell wall components. Taken together, the data obtained indicate that less significant changes were observed in fruit in the breaker stage than in the case of the red ripe stage. The first symptoms of changes were noted after 24 h, but only after 72 h, more crucial deviations were visible. The 5% oxygen concentration slows down the ripening process and 0% oxygen accelerates the changes taking place during ripening. CONCLUSIONS: The observed molecular reset occurring in tomato cell walls in hypoxic and anoxic conditions seems to be a result of regulatory and protective mechanisms modulating ripening processes.

Selective Targeting of α4ß7/MAdCAM-1 Axis Suppresses Fibrosis Progression by Reducing Proinflammatory T Cell Recruitment to the Liver.

Integrin α4ß7+ T cells perpetuate tissue injury in chronic inflammatory diseases, yet their role in hepatic fibrosis progression remains poorly understood. Here, we report increased accumulation of α4ß7+ T cells in the liver of people with cirrhosis relative to disease controls. Similarly, hepatic fibrosis in the established mouse model of CCl4-induced liver fibrosis was associated with enrichment of intrahepatic α4ß7+ CD4 and CD8 T cells. Monoclonal antibody (mAb)-mediated blockade of α4ß7 or its ligand mucosal addressin cell adhesion molecule (MAdCAM)-1 attenuated hepatic inflammation and prevented fibrosis progression in CCl4-treated mice. Improvement in liver fibrosis was associated with a significant decrease in the infiltration of α4ß7+ CD4 and CD8 T cells, suggesting that α4ß7/MAdCAM-1 axis regulates both CD4 and CD8 T cell recruitment to the fibrotic liver, and α4ß7+ T cells promote hepatic fibrosis progression. Analysis of hepatic α4ß7+ and α4ß7- CD4 T cells revealed that α4ß7+ CD4 T cells were enriched for markers of activation and proliferation, demonstrating an effector phenotype. The findings suggest that α4ß7+ T cells play a critical role in promoting hepatic fibrosis progression, and mAb-mediated blockade of α4ß7 or MAdCAM-1 represents a promising therapeutic strategy for slowing hepatic fibrosis progression in chronic liver diseases.

MAdCAM-1 co-stimulation combined with retinoic acid and TGF-ß induces blood CD8+ T cells to adopt a gut CD101+ TRM phenotype.

Resident memory T cells (TRMs) help control local immune homeostasis and contribute to tissue-protective immune responses. The local cues that guide their differentiation and localization are poorly defined. We demonstrate that mucosal vascular addressin cell adhesion molecule 1, a ligand for the gut-homing receptor α4ß7 integrin, in the presence of retinoic acid and transforming growth factor-ß (TGF-ß) provides a co-stimulatory signal that induces blood cluster of differentiation (CD8+ T cells to adopt a TRM-like phenotype. These cells express CD103 (integrin αE) and CD69, the two major TRM cell-surface markers, along with CD101. They also express C-C motif chemokine receptors 5 (CCR5) , C-C motif chemokine receptors 9 (CCR9), and α4ß7, three receptors associated with gut homing. A subset also expresses E-cadherin, a ligand for αEß7. Fluorescent lifetime imaging indicated an αEß7 and E-cadherin cis interaction on the plasma membrane. This report advances our understanding of the signals that drive the differentiation of CD8+ T cells into resident memory T cells and provides a means to expand these cells in vitro, thereby affording an avenue to generate more effective tissue-specific immunotherapies.

AGR2-mediated unconventional secretion of 14-3-3ε and α-actinin-4, responsive to ER stress and autophagy, drives chemotaxis in canine mammary tumor cells.

BACKGROUND: Canine mammary tumors (CMTs) in intact female dogs provide a natural model for investigating metastatic human cancers. Our prior research identified elevated expression of Anterior Gradient 2 (AGR2), a protein disulfide isomerase (PDI) primarily found in the endoplasmic reticulum (ER), in CMT tissues, highly associated with CMT progression. We further demonstrated that increased AGR2 expression actively influences the extracellular microenvironment, promoting chemotaxis in CMT cells. Unraveling the underlying mechanisms is crucial for assessing the potential of therapeutically targeting AGR2 as a strategy to inhibit a pro-metastatic microenvironment and impede tumor metastasis. METHODS: To identify the AGR2-modulated secretome, we employed proteomics analysis of the conditioned media (CM) from two CMT cell lines ectopically expressing AGR2, compared with corresponding vector-expressing controls. AGR2-regulated release of 14-3-3ε (gene: YWHAE) and α-actinin 4 (gene: ACTN4) was validated through ectopic expression, knockdown, and knockout of the AGR2 gene in CMT cells. Extracellular vesicles derived from CMT cells were isolated using either differential ultracentrifugation or size exclusion chromatography. The roles of 14-3-3ε and α-actinin 4 in the chemotaxis driven by the AGR2-modulated CM were investigated through gene knockdown, antibody-mediated interference, and recombinant protein supplement. Furthermore, the clinical relevance of the release of 14-3-3ε and α-actinin 4 was assessed using CMT tissue-immersed saline and sera from CMT-afflicted dogs. RESULTS: Proteomics analysis of the AGR2-modulated secretome revealed increased abundance in 14-3-3ε and α-actinin 4. Ectopic expression of AGR2 significantly increased the release of 14-3-3ε and α-actinin 4 in the CM. Conversely, knockdown or knockout of AGR2 expression remarkably reduced their release. Silencing 14-3-3ε or α-actinin 4 expression diminished the chemotaxis driven by AGR2-modulated CM. Furthermore, AGR2 controls the release of 14-3-3ε and α-actinin 4 primarily via non-vesicular routes, responding to the endoplasmic reticulum (ER) stress and autophagy activation. Knockout of AGR2 resulted in increased α-actinin 4 accumulation and impaired 14-3-3ε translocation in autophagosomes. Depletion of extracellular 14-3-3ε or α-actinin 4 reduced the chemotaxis driven by AGR2-modulated CM, whereas supplement with recombinant 14-3-3ε in the CM enhanced the CM-driven chemotaxis. Notably, elevated levels of 14-3-3ε or α-actinin 4 were observed in CMT tissue-immersed saline compared with paired non-tumor samples and in the sera of CMT dogs compared with healthy dogs. CONCLUSION: This study elucidates AGR2's pivotal role in orchestrating unconventional secretion of 14-3-3ε and α-actinin 4 from CMT cells, thereby contributing to paracrine-mediated chemotaxis. The insight into the intricate interplay between AGR2-involved ER stress, autophagy, and unconventional secretion provides a foundation for refining strategies aimed at impeding metastasis in both canine mammary tumors and potentially human cancers.

Intestinal MAdCAM-1 imaging as biomarker for prognostic in murine models of multiple sclerosis.

INTRODUCTION: Multiple sclerosis (MS) is an autoimmune disease of the central nervous system. Recent evidence suggests that lymphocyte trafficking in the intestines could play a key role in its etiology. Nevertheless, it is not clear how intestinal tissue is involved in the disease onset nor its evolution. In the present study, we aimed to evaluate intestinal inflammation dynamic throughout the disease course and its potential impact on disease progression. METHODS: We used tissue immunophenotyping (immunohistofluorescence and flow cytometry) and a recently described molecular magnetic resonance imaging (MRI) method targeting mucosal addressin cell adhesion molecule-1 (MAdCAM-1) to assess intestinal inflammation in vivo in two distinct animal models of MS (Experimental Autoimmune Encephalomyelitis - EAE) at several time points of disease progression. RESULTS: We report a positive correlation between disease severity and MAdCAM-1 MRI signal in two EAE models. Moreover, high MAdCAM-1 MRI signal during the asymptomatic phase is associated with a delayed disease onset in progressive EAE and to a lower risk of conversion to a secondary-progressive form in relapsing-remitting EAE. During disease evolution, in line with a bi-directional immune communication between the gut and the central nervous system, we observed a decrease in T-CD4+ and B lymphocytes in the ileum concomitantly with their increase in the spinal cord. CONCLUSION: Altogether, these data unveil a crosstalk between intestinal and central inflammation in EAE and support the use of molecular MRI of intestinal MAdCAM-1 as a new biomarker for prognostic in MS patients.

Functional analysis of fasciclin-like arabinogalactan in carotenoid synthesis during tomato fruit ripening.

Carotenoids are important pigmented nutrients synthesized by tomato fruits during ripening. To reveal the molecular mechanism underlying carotenoid synthesis during tomato fruit ripening, we analyzed carotenoid metabolites and transcriptomes in six development stages of tomato fruits. A total of thirty different carotenoids were detected and quantified in tomato fruits from 10 to 60 DPA. Based on differential gene expression profiles and WGCNA, we explored several genes that were highly significant and negatively correlated with lycopene, all of which encode fasciclin-like arabinogalactan proteins (FLAs). The FLAs are involved in plant signal transduction, however the functional role of these proteins has not been studied in tomato. Genome-wide analysis revealed that cultivated and wild tomato species contained 18 to 22 FLA family members, clustered into four groups, and mainly evolved by means of segmental duplication. The functional characterization of FLAs showed that silencing of SlFLA1, 5, and 13 were found to contribute to the early coloration of tomato fruits, and the expression of carotenoid synthesis-related genes was up-regulated in fruits that changed phenotypically, especially in SlFLA13-silenced plants. Furthermore, the content of multiple carotenoids (including (E/Z)-phytoene, lycopene, γ-carotene, and α-carotene) was significantly increased in SlFLA13-silenced fruits, suggesting that SlFLA13 has a potential inhibitory function in regulating carotenoid synthesis in tomato fruits. The results of the present study broaden the idea of analyzing the biological functions of tomato FLAs and preliminary evidence for the inhibitory role of SlFLA13 in carotenoid synthesis in fruit, providing the theoretical basis and a candidate for improving tomato fruit quality.

Potential toxicity of graphene (oxide) quantum dots via directly covering the active site of anterior gradient homolog 2 protein.

Graphene quantum dots (GQDs) have attracted significant attention in biomedicine, while extensive investigations have revealed a reverse regarding the potential biotoxicity of GQDs. In order to supplementing the understanding of the toxicity profile of GQDs, this study employs a molecular dynamics (MD) simulation approach to systematically investigate the potential toxicity of both GQDs and Graphene Oxide Quantum Dots (GOQDs) on the Anterior Gradient Homolog 2 (AGR2) protein, a key protein capable of protecting the intestine. We construct two typical simulation systems, in which an AGR2 protein is encircled by either GQDs or GOQDs. The MD results demonstrate that both GQDs and GOQDs can directly make contact with and even cover the active site (specifically, the Cys81 amino acid) of the AGR2 protein. This suggests that GQDs and GOQDs have the capability to inhibit or interfere with the normal biological interaction of the AGR2 active site with its target protein. Thus, GQDs and GOQDs exhibit potential detrimental effects on the AGR2 protein. Detailed analyses reveal that GQDs adhere to the Cys81 residue due to van der Waals (vdW) interaction forces, whereas GOQDs attach to the Cys81 residue through a combination of vdW (primary) and Coulomb (secondary) interactions. Furthermore, GQDs aggregation typically adsorb onto the AGR2 active site, while GOQDs adsorb to the active site of AGR2 one by one. Consequently, these findings shed new light on the potential adverse impact of GQDs and GOQDs on the AGR2 protein via directly covering the active site of AGR2, providing valuable molecular insights for the toxicity profile of GQD nanomaterials.

Implications of cell wall immunocytochemical profiles on the structural and functional traits of root and stem galls induced by Eriosoma lanigerum on Malus domestica.

Alterations in cell wall composition imply in new structural and functional traits in gall developmental sites, even when the inducer is a sucking exophytophagous insect with strict feeding sites as the aphid associated to Malus domestica Borkh. This host plant is an economically important, fruit-bearing species, susceptible to gall induction by the sucking aphid Eriosoma lanigerum Hausmann, 1802. Herein, the immunocytochemical detection of arabinogalactan-proteins (AGPs), pectins, and hemicelluloses using monoclonal antibodies was performed in samples of non-galled roots and stems, and of root and stem galls on M. domestica. The dynamics of these cell wall components was discussed under the structural and functional traits of the galls proximal, median, and distal regions, according to the proximity of E. lanigerum colony feeding site. In the proximal region, the epitopes of AGPs and homogalacturonans (HGs) are related to cell growth and divisions, which result in the overproduction of parenchyma cells both in root and stem galls. In the proximal and median regions, the co-occurrence of HGs and arabinans in the cell walls of parenchyma and secondary tissues favors the nutrient flow and water-holding capacity, while the xylogalacturonans and hemicelluloses may function as additional carbohydrate resources to E. lanigerum. The immunocytochemical profile of the cell walls support the feeding activity of E. lanigerum mainly in the gall proximal region. The similarity of the cell wall components of the gall distal region and the non-galled portions, both in roots and stems, relates to the decrease of the cecidogenetic field the more distant the E. lanigerum colony is.

Structure of KOH-soluble polysaccharides from сoniferous greens of Norway spruce (Picea abies): The pectin-xylan-AGPs complex. Part 1.

Using 7 % KOH, the polysaccharide PAK has been isolated from the coniferous greens of Norway spruce. PAK was found to contain predominantly arabinoglucuronoxylan, xyloglucan and arabinan, but also pectic polysaccharides, glucomannan and arabinogalactan proteins (AGPs), as determined by 1D/2D NMR analysis. It was found that fractionation of PAK on DEAE-cellulose resulted in simultaneous elution of pectins, arabinoglucuronoxylans and AGPs. It was evident that the content of 4-OMe-α-D-GlcpA and xylose, 1,4-ß-D-GlcpA, and T-ß-D-GlcpA increased with an increase in NaCl concentration. However, 1,4-α-D-GalpA content was almost independent of NaCl concentration, indicating unchanged pectic polysaccharide concentration. Interestingly, pectins extracted with 0.1-0.3 M NaCl solutions were richer in rhamnogalacturonan-I (RG-I) than those extracted with water and 0.01 M NaCl. Conclusion: The content of RG-I, AGPs and arabinoglucuronoxylan rises with rising NaCl concentration. An intense signal indicating an intermolecular linkage between the xylan and RG-I domains, i.e. that part of the arabinoglucuronoxylan is covalently bound to RG-I, is observed in the HMBC spectra of the polysaccharides obtained. The discovery here of a new relationship between rhamnogalacturonan I and xylan contradicts the prevailing cell wall model.