Altered glycosylation of several metastasis-associated glycoproteins with terminal GalNAc defines the highly invasive cancer cell phenotype

Publisher Copyright: © 2022 Khosrowabadi et al.Several distinct metastasis-associated glycosylation changes have been shown to promote cancer cell invasion and metastasis, the main cause of death of cancer patients. However, it is unclear whether their presence reflects cell- or tissue-specific variations for metastasis, or species needed to drive different phases of the metastatic cascade. To address this issue from a different perspective, we investigated here whether different cancer cell lines share any glycotopes that are common and important for their invasive phenotype. By using lectin microarray glycan profiling and an established myoma tissue-based 3D invasion assay, we identified a single glycotope recognized by Helix Pomatia agglutinin (HPA), whose expression level in different cancer cells correlated significantly with their invasive potential. Lectin pull-down assay and LC-MS/MS analysis in highly- (A431 and SW-48) and poorly invasive (HepG2 and RCC4) cancer cells revealed ~85 glycoproteins of which several metastasis-promoting members of the integrin family of cell adhesion receptors, the epidermal growth factor receptor (EGFR) and the matrix metalloproteinase-14 (MMP-14) were among the abundant ones. Moreover, we showed that the level of the GalNAc glycotope in MMP-14, EGFR, αV-, β1- and β4 integrin in highly and poorly invasive cancer cells correlated positively with their invasive potential. Collectively, our findings suggest that altered glycosylation of several metastasis-associated glycoproteins with terminal GalNAc drives the highly invasive cancer cell phenotype.Peer reviewe

Screening and characterization of several 2,2-dicholoropropionic acid-degrading bacteria isolated from marine sediment of danga bay and east coast of Singapore Island

Industries and agriculture activities extensively utilize halogenated compounds. These compounds were found to be toxic and pollute the environment. Thus, many studies have been done on microbial degradation of these chemicals. In this study, an attempt was made to isolate bacterial strains EK1-EK5 from marine sediments collected at Danga Bay and east coast of Singapore island. The 16S rRNA analysis suggested that the isolated bacteria had more than 96% sequence identity to the sequence in the database; therefore, they were designated as Bacillus sp., Rhodococcus sp., Lysinibacillus sp., Microbacterium sp., and Aminobacter sp. The results of molecular analysis were supported by biochemical and microscopic examinations. Bacterial isolates were able to grow slowly in minimal medium containing only 2,2-dichloropropionate as the sole carbon source. The cellular doubling times were 39.60 ± 0.1, 36.60 ± 0.2, 30.71 ± 0.1, 41.23 ± 0.1, and 36.70 ± 0.3 h for EK1, EK2, EK3, EK4, and EK5, respectively. In the future, it will be important to further investigate the presence of the dehalogenase gene in their genomic DNA for further characterization

N-glycan biosynthesis:basic principles and factors affecting its outcome

Abstract Carbohydrate chains are the most abundant and diverse of nature’s biopolymers and represent one of the four fundamental macromolecular building blocks of life together with proteins, nucleic acids, and lipids. Indicative of their essential roles in cells and in multicellular organisms, genes encoding proteins associated with glycosylation account for approximately 2% of the human genome. It has been estimated that 50–80% of all human proteins carry carbohydrate chains—glycans—as part of their structure. Despite cells utilize only nine different monosaccharides for making their glycans, their order and conformational variation in glycan chains together with chain branching differences and frequent post-synthetic modifications can give rise to an enormous repertoire of different glycan structures of which few thousand is estimated to carry important structural or functional information for a cell. Thus, glycans are immensely versatile encoders of multicellular life. Yet, glycans do not represent a random collection of unpredictable structures but rather, a collection of predetermined but still dynamic entities that are present at defined quantities in each glycosylation site of a given protein in a cell, tissue, or organism. In this chapter, we will give an overview of what is currently known about N-glycan synthesis in higher eukaryotes, focusing not only on the processes themselves but also on factors that will affect or can affect the final outcome—the dynamicity and heterogeneity of the N-glycome. We hope that this review will help understand the molecular details underneath this diversity, and in addition, be helpful for those who plan to produce optimally glycosylated antibody-based therapeutics

Altered glycosylation of several metastasis-associated glycoproteins with terminal GalNAc defines the highly invasive cancer cell phenotype

Publisher Copyright: © 2022 Khosrowabadi et al.Several distinct metastasis-associated glycosylation changes have been shown to promote cancer cell invasion and metastasis, the main cause of death of cancer patients. However, it is unclear whether their presence reflects cell- or tissue-specific variations for metastasis, or species needed to drive different phases of the metastatic cascade. To address this issue from a different perspective, we investigated here whether different cancer cell lines share any glycotopes that are common and important for their invasive phenotype. By using lectin microarray glycan profiling and an established myoma tissue-based 3D invasion assay, we identified a single glycotope recognized by Helix Pomatia agglutinin (HPA), whose expression level in different cancer cells correlated significantly with their invasive potential. Lectin pull-down assay and LC-MS/MS analysis in highly- (A431 and SW-48) and poorly invasive (HepG2 and RCC4) cancer cells revealed ~85 glycoproteins of which several metastasis-promoting members of the integrin family of cell adhesion receptors, the epidermal growth factor receptor (EGFR) and the matrix metalloproteinase-14 (MMP-14) were among the abundant ones. Moreover, we showed that the level of the GalNAc glycotope in MMP-14, EGFR, αV-, β1- and β4 integrin in highly and poorly invasive cancer cells correlated positively with their invasive potential. Collectively, our findings suggest that altered glycosylation of several metastasis-associated glycoproteins with terminal GalNAc drives the highly invasive cancer cell phenotype.Peer reviewe

Analytical prediction of liquid-gated graphene nanoscroll biosensor performance

The latest discovery of the graphene nanoscroll has provided enormous new stimuli to carbon nanoresearch. Due to its one-dimensional structure and tunable core size, the graphene nanoscroll is suitable for nanoscale applications such as in nanotransistors, and biosensor devices. DNA sensing is critical in the identification of the genetic risk factors associated with complex human diseases, and continues to have an emerging role in therapeutics and personalized medicine. This paper presents the analytical model of liquid-gated field effect transistors (LGFETs) for zig-zag graphene nanoscrolls (ZGNSs) inspired by carbon nanotube behavior when exposed to DNA molecules. First of all, in order to gain physical insight into GNS-based devices, the conductance of GNSs is analytically modelled. Based on the sensing mechanism of the DNA sensor, GNS controlling elements (ηGNS and εGNS) are proposed and the behavior of LGFETs-based GNS nanomaterial in the presence of DNA molecules is predicted to get a greater insight into the rapid development of DNA sensors and their application. Because of the channel-doping effect due to the adsorption of the DNA molecules, the conductance of the channel is altered. On the other hand, the applied voltage effect in the form of tilted electron energy levels is utilized in the form of normalized Fermi energy variation which is used in the sensor modelling. This study emphasizes the promising nature of carbon nanoscrolls for a number of electronic device application

A dominant negative splice variant of the heparan sulfate biosynthesis enzyme NDST1 reduces heparan sulfate sulfation

NDST1 (glucosaminyl N-deacetylase/N-sulfotransferase) is a key enzyme in heparan sulfate (HS) biosynthesis, where it is responsible for HS N-deacetylation and N-sulfation. In addition to the full length human enzyme of 882 amino acids, here designated NDST1A, a shorter form containing 825 amino acids (NDST1B) is synthesized after alternative splicing of the NDST1 mRNA. NDST1B is mostly expressed at a low level, but increased amounts are seen in several types of cancer where it is associated with shorter survival. In this study, we aimed at characterizing the enzymatic properties of NDST1B and its effect on HS biosynthesis. Purified recombinant NDST1B lacked both N-deacetylase and N-sulfotransferase activities. Interestingly, HEK293 cells overexpressing NDST1B synthesized HS with reduced sulfation and altered domain structure. Fluorescence resonance energy transfer-microscopy demonstrated that both NDST1A and NDST1B had the capacity to interact with the HS copolymerase subunits EXT1 and EXT2 and also to form NDST1A/NDST1B dimers. Since lysates from cells overexpressing NDST1B contained less NDST enzyme activity than control cells, we suggest that NDST1B works in a dominant negative manner, tentatively by replacing the active endogenous NDST1 in the enzyme complexes taking part in biosynthesis

Abnormal golgi pH homeostasis in cancer cells impairs apical targeting of carcinoembryonic antigen by inhibiting its glycosyl-phosphatidylinositol anchor-mediated association with lipid rafts

Abstract Aims: Carcinoembryonic antigen (CEACAM5, CEA) is a known tumor marker for colorectal cancer that localizes in a polarized manner to the apical surface in normal colon epithelial cells whereas in cancer cells it is present at both the apical and basolateral surfaces of the cells. Since the Golgi apparatus sorts and transports most proteins to these cell surface domains, we set out here to investigate whether any of the factors commonly associated with tumorigenesis, including hypoxia, generation of reactive oxygen species (ROS), altered redox homeostasis, or an altered Golgi pH, are responsible for mistargeting of CEA to the basolateral surface in cancer cells. Results: Using polarized nontumorigenic Madin-Darby canine kidney (MDCK) cells and CaCo-2 colorectal cancer cells as targets, we show that apical delivery of CEA is not affected by hypoxia, ROS, nor changes in the Golgi redox state. Instead, we find that an elevated Golgi pH induces basolateral targeting of CEA and increases its TX-100 solubility, indicating impaired association of CEA with lipid rafts. Moreover, disruption of lipid rafts by methyl-β-cyclodextrin induced accumulation of the CEA protein at the basolateral surface in MDCK cells. Experiments with the glycosylphosphatidylinositol (GPI)-anchorless CEA mutant and CEA-specific GPI-anchored enhanced green fluorescent protein (EGFP-GPI) fusion protein revealed that the GPI-anchor was critical for the pH-dependent apical delivery of the CEA in MDCK cells. Innovation and Conclusions: The findings indicate that an abnormal Golgi pH homeostasis in cancer cells is an important factor that causes mistargeting of CEA to the basolateral surface of cancer cells via inhibiting its GPI-anchor-mediated association with lipid rafts

Loss of NDST1 N -sulfotransferase activity is associated with autosomal recessive intellectual disability

International audienceIntellectual Disability (ID) is the major cause of handicap, affecting nearly 3% of the general population, and is highly genetically heterogenous with more than a thousand genes involved. Exome sequencing performed in two independent families identified the same missense variant, p.(Gly611Ser), in the NDST1 (N-deacetylase/N-sulfotransferase member 1) gene. This variant had been previously found in ID patients of two other families but has never been functionally characterized. The NDST1 gene encodes a bifunctional enzyme that catalyzes both N-deacetylation and N-sulfation of N-acetyl-glucosamine residues during heparan sulfate (HS) biosynthesis. This step is essential because it influences the downstream enzymatic modifications and thereby determines the overall structure and sulfation degree of the HS polysaccharide chain. To discriminate between a rare polymorphism and a pathogenic variant, we compared the enzymatic properties of wild-type and mutant NDST1 proteins. We found that the p.(Gly611Ser) variant results in a complete loss of N-sulfotransferase activity while the N-deacetylase activity is retained. NDST1 shows the highest and the most homogeneous expression in the human cerebral structures compared to the other members of the NDST gene family. These results indicate that a loss of NDST1 N-sulfation activity is associated with impaired cognitive functions

SLC4A2 anion exchanger promotes tumour cell malignancy via enhancing net acid efflux across golgi membranes

Abstract Proper functioning of each secretory and endocytic compartment relies on its unique pH micro-environment that is known to be dictated by the rates of V-ATPase-mediated H+ pumping and its leakage back to the cytoplasm via an elusive “H+ leak” pathway. Here, we show that this proton leak across Golgi membranes is mediated by the AE2a (SLC4A2a)-mediated bicarbonate-chloride exchange, as it is strictly dependent on bicarbonate import (in exchange for chloride export) and the expression level of the Golgi-localized AE2a anion exchanger. In the acidic Golgi lumen, imported bicarbonate anions and protons then facilitate a common buffering reaction that yields carbon dioxide and water before their egress back to the cytoplasm via diffusion or water channels. The flattened morphology of the Golgi cisternae helps this process, as their high surface-volume ratio is optimal for water and gas exchange. Interestingly, this net acid efflux pathway is often upregulated in cancers and established cancer cell lines, and responsible for their markedly elevated Golgi resting pH and attenuated glycosylation potential. Accordingly, AE2 knockdown in SW-48 colorectal cancer cells was able to restore these two phenomena, and at the same time, reverse their invasive and anchorage-independent growth phenotype. These findings suggest a possibility to return malignant cells to a benign state by restoring Golgi resting pH

Molecular characterization of monochloroacetate-degrading arthrobacter sp. Strain d2 isolated from Universiti Teknologi Malaysia agricultural area

Arthrobacter sp. strains D2 and D3 and Labrys sp. strain D1 capable of degrading 20 mM monochloroacetic acid (MCA) were isolated from soil contaminated with herbicides and pesticides. All three isolates were able to grow onMCAas the sole source of carbon and energy with concomitant chloride ion release in the growth medium (19 mM). Strains D2 and D3 (cells doubling time 7±0.3 h) grew four times faster than D1 (26±0.1 h). Strain D2 was then further investigated and could also grow in 10 mM of monobromoacetic acid (MBA),2,2-dichloropropionic acid (2,2DCP), D,L-2-chloropropionic acid (D,L2CP), L- 2-chloropropionic acid (L-2CP), D-2-chloropropionic acid (D-2CP), and glycolate as the sole sources of carbon and energy. Dehalogenase gene amplification using group I primers revealed a 410-bp polymerase chain reaction (PCR) product, but there was none using group II primers. The partial amino acid sequence analysis of group I DehD2 dehalogenase showed at least 32% identity to the corresponding regions of DehE, DhlIV, DehI, and D,L-DEX, with key amino acid residues Ser188, Ala187, and Asp189. These amino acid residues were involved in substrate binding and catalysis and were conserved in the partial amino acid sequence