Ethylenediaminetetraacetic acid affects subcellular expression of clusterin protein in human colon adenocarcinoma COLO 205 cell line.
The aim of our study was to determine the expression of various isoforms of clusterin and to evaluate how etoposide or calcium chelators [ethylenediaminetetraacetic acid and (2-aminoethoxyethane)-N,N,N',N'-tetraacetic acid] affect the subcellular expressions of the 50-kDa isoform of clusterin protein in colon adenocarcinoma COLO 205 cells. We then determined how the cytoplasmic vs. nuclear expression of the 50-kDa isoform of clusterin correlates with the viability of COLO 205 cells. To identify the clusterin isoforms, and its nuclear and cytoplasmic expression in COLO 205 cells, Western bloting was used. Cell viability was determined by the 3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyltetrazolium bromide assay. Etoposide decreased the viability of COLO 205 cells with a concomitant increase in the 50-kDa clusterin concentration in the cell nucleus. Chelation of the extracellular calcium ions by (2-aminoethoxyethane)-N,N,N',N'-tetraacetic acid did not modulate the subcellular distribution of clusterin. The use of ethylenediaminetetraacetic acid, which reduces the intracellular and extracellular calcium levels, stimulated nuclear expression of clusterin protein and was accompanied by extensive cell death. Intracellular calcium determines cytoplasmic expression and antiapoptotic activity of the intracellular protein clusterin. The depletion of intracellular calcium leads to increased nuclear expression of the 50-kDa clusterin protein, which is accompanied by cell death. We concluded that there is at least one cell death-promoting pathway in COLO 205 cells that is dependent on intracellular calcium and nuclear localization of 50-kDa clusterin.
Pajak
Beata
B
Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw Agricultural University, Warsaw, Poland.
Orzechowski
Arkadiusz
A
D000230:Adenocarcinoma / Q000188:drug therapy / Q000378:metabolism*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects*; D002614:Chelating Agents / Q000494:pharmacology*; D051152:Clusterin / Q000494:pharmacology*; D015179:Colorectal Neoplasms / Q000188:drug therapy / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology*; D005047:Etoposide / Q000494:pharmacology*; D006706:Homeostasis / Q000187:drug effects; D006801:Humans; D008967:Molecular Biology; D020033:Protein Isoforms
D000972:Antineoplastic Agents, Phytogenic; D002614:Chelating Agents; D051152:Clusterin; D020033:Protein Isoforms; D005047:Etoposide; D004492:Edetic Acid; D002118:Calcium