Gene expression profiling in wild-type and metallothionein mutant fibroblast cell lines

Abstract

The role of metallothioneins (MT) in copper homeostasis is of great interest, as it appears to be partially responsible for the regulation of intracellular copper levels during adaptation to extracellular excess of the metal. To further investigate a possible role of MTs in copper metabolism, a genomics approach was utilized to evaluate the role of MT on gene expression. Microarray analysis was used to examine the effects of copper overload in fibroblast cells from normal and MT I and II double knock-out mice (MT-/-). As a first step, we compared genes that were significantly upregulated in wild-type and MT-/-cells exposed to copper. Even though wild-type and mutant cells are undistinguishable in terms of their morphological features and rates of growth, our results show that MT-/-cells do not respond with induction of typical markers of cellular stress under copper excess conditions, as observed in the wild-type cell line, suggesting that the transcription initiation rate or the mRNA stability of stress genes is affected when there is an alteration in the copper store capacity. The functional classification of other up-regulated genes in both cell lines indicates that a large proportion (>80%) belong to two major categories: 1) metabolism; and 2) cellular physiological processes, suggesting that at the transcriptional level copper overload induces the expression of genes associated with diverse molecular functions. These results open the possibility to understand how copper homeostasis is being coordinated with other metabolic pathways.

Physiological and toxicological transcriptome changes in HepG2 cells exposed to copper

“…The cellular and molecular mechanisms underlying copperregulated gene expression and toxicity have been investigated in yeast, mouse fibroblasts, and rodent strains with mutations in Atp7b (5,28,33,69). Atp7b Ϫ/Ϫ mice demonstrate intracellular copper accumulation, low serum oxidase activity, and increased copper excretion in the urine and liver pathology, similar to Wilson's disease patients (12,32).…”

“…There were also enriched GO categories that mapped to specific treatment conditions: oxygen and ROS metabolism and IGF binding (400 M, 8 h); ubiquitin-protein ligase activity (600 M, 8 h); endosome organization and (2) Lipid transport (8)* Cholesterol binding (2) Lipoprotein metabolism (6)* Cholesterol metabolism (4) Acyl-CoA binding (3) Hemoglobin binding (2) Alcohol metabolism (19) Platelet activation (2) Amine metabolism (26) Regulation of body fluids (4) Chromatin assembly (6) Serine-type endopeptidase inhibitor activity (5) Creatine biosynthesis (2) Wound healing (4) Deoxyribonucleotide biosynthesis ( (2) B cell-mediated immunity (7) Hemoglobin binding (2) Complement activation (7) Lipid binding (9) Creatine biosynthesis (2) Cytolysis (4) Hormone metabolism (10) Lipid transporter activity (13) Nickel ion binding (2) Oxidoreductase activity (52) Positive regulation of immune response (7) Serine-type peptidase activity (14) Xenobiotic metabolism (5) Shown are Gene Ontology (GO) categories with P Յ 0.005. Responsive genes were 2-fold downregulated with FDR P Յ 0.05.…”

Environmental epitranscriptomics

“…Another potentially fruitful avenue of future research is in the effects of copper exposure. Independent in vitro studies from two groups using human liver and lung cell lines have similarly revealed increased expression of the m 6 A writer METTL14 (Song et al, 2009), while elsewhere the reader HNRNPA2B1 has been shown to be similarly upregulated (Armendariz et al, 2006). The latter is supported by in vivo findings of increased Hnrnpa2b1 expression in mouse liver (Burkhead et al, 2011).…”

Molecular Mechanisms Underlying the Nephrotoxicity of Cisplatin, Lead Acetate and Cyclosporine: Key Roles of Myc and Smad4

“…During apoptosis Hnrpu is cleaved in a caspase-dependent way. Interestingly, Li et al found that Hnrpu up-regulation is followed by cooper overload and vancomycin nephrotoxicity [36,37]. By analysis of PPI networks, we present Hnrpu as a promising biomarker for Lead acetate nephrotoxicity.…”