Modification of Ser and Thr residues by attachment of O-linked N-acetylglucosamine [Ser(Thr)-O-GlcNAcylation] to eukaryotic nuclear and cytosolic proteins is as dynamic and possibly as abundant as Ser(Thr) phosphorylation. Known O-GlcNAcylated proteins include cytoskeletal proteins and their regulatory proteins; viral proteins; nuclear-pore, heat-shock, tumor-suppressor, and nuclear-oncogene proteins; RNA polymerase II catalytic subunit; and a multitude of transcription factors. Although functionally diverse, all of these proteins are also phosphoproteins. Most O-GlcNAcylated proteins form highly regulated multimeric associations that are dependent upon their posttranslational modifications. Evidence is mounting that O-GlcNAcylation is an important regulatory modification that may have a reciprocal relationship with O-phosphorylation and may modulate many biological processes in eukaryotes.

Dynamic O-Linked Glycosylation of Nuclear and Cytoskeletal Proteins

Twenty human colon carcinoma cell lines were studied for their ability to develop some of the characteristics of the normal intestinal epithelium, e.g., epithelial polarity, presence of the actin-binding protein villin, or the occurrence of an enterocytic differentiation either when cultured under standard conditions, as for Caco-2 cells, or when grown in a glucose-free medium, as for HT-29 cells. Except for the regular presence of villin, which can be considered a marker of the colonic origin of the cells, the cell lines of this study could be classified into four types with regard to their differentiation characteristics. In type 1 (only one cell line, i.e., Caco-2) the cells undergo spontaneously an enterocytic differentiation characterized by a polarization of the cell layer with the formation of domes and the presence of an apical brush border the membrane of which is endowed with hydrolases such as sucrase-isomaltase, lactase, amino-peptidase N, dipeptidylpeptidase IV and alkaline phosphatase. In type 2 (three cell lines: HT-29, HCT-EB, and HCT-GEO) the cells are undifferentiated when grown in the presence of glucose but undergo an enterocytic differentiation when grown in the absence of glucose. In type 3 (eight cell lines: HCT-GLY, HCT-FET, HCT-FRI, HCT-CBS, HCT-ALA, Co-115, HRT-18, and SW-1116) the cells are organized into a polarized monolayer with the formation of domes but without any enterocytic differentiation characteristics, whatever the culture conditions. In type 4 (eight cell lines: HCT-116a, HCT-R, HCT-RCA, HCT-Moser, HCT-8R, SW-480, LS-174T, and Vaco-9P) the cells are organized into a multilayer without any feature of epithelial polarity or enterocytic differentiation, whatever the culture conditions.

https://cancerres.aacrjournals.org/content/canres/48/7/1936.full.pdf

Epithelial polarity, villin expression, and enterocytic differentiation of cultured human colon carcinoma cells: A survey of twenty cell lines

The human colon carcinoma cell lines HT-29 and Caco-2: Two in vitro models for the study of intestinal differentiation

The purpose of this work was to investigate whether the phenomenon of metabolic adaptation of HT-29 cells to glucose deprivation and subsequent emergence of differentiated subpopulations (A. Zweibaum et al., J. Cell. Physiol., 122: 21–29, 1985) also applies to anticancer drugs that act at a metabolic level like methotrexate (MTX). Stepwise adaptation of exponentially growing HT-29 cells to increasing concentrations of MTX (10-7, 10-6, and 10-5 mol) results, after a phase of high mortality, in the emergence of subpopulations with stable growth rates and curves close to those of untreated control cells. In contrast to control cells which are heterogenous and contain, after confluency, only a small proportion of differentiated cell types (less than 4%), postconfluent cultures of MTX-adapted cells are totally differentiated. Cells adapted to 10-7m MTX form a mixed population of columnar absorptive and mucus cells; at higher concentrations cells are almost exclusively of the mucus-secreting type. All cells, whether mucus-secreting or not, develop an apical brush border which strongly expresses dipeptidylpeptidase IV, carcinoembryonic antigen, and villin. These differentiation features, which resemble those of fetal colon, are associated with decreased rates of glucose consumption and lactic acid production. Both differentiation characteristics and metabolic changes are stably maintained when the cells are subcultured in the absence of the drug. Like the original population, MTX-adapted cells are tumorigenic in nude mice. We propose that cells which are able to differentiate and which are the origin of the small proportion of differentiated cell types found in postconfluent cultures of the original cell line possess an advantage which allows them to be adaptable to “metabolic stress” conditions.

/pdf/growth-adaptation-to-methotrexate-of-ht-29-human-colon-4c2t7pn65w.pdf

Growth Adaptation to Methotrexate of HT-29 Human Colon Carcinoma Cells Is Associated with Their Ability to Differentiate into Columnar Absorptive and Mucus-secreting Cells

http://www.reboundhealth.com/cms/images/pdf/Glucosamine/pmid%2015238246%20o-glcnac%20id%2014823.pdf

O-GlcNAc a sensor of cellular state: the role of nucleocytoplasmic glycosylation in modulating cellular function in response to nutrition and stress.

http://www.jbc.org/content/260/1/139.full.pdf

B. M. Wice

Papers

The intracellular accumulation of UDP-N-acetylhexosamines is concomitant with the inability of human colon cancer cells to differentiate.