Elsevier

Genomics

Volume 83, Issue 4, April 2004, Pages 600-614
Genomics

Ethanol-response genes and their regulation analyzed by a microarray and comparative genomic approach in the nematode Caenorhabditis elegans

https://doi.org/10.1016/j.ygeno.2003.10.008Get rights and content

Abstract

The nematode shows responses to acute ethanol exposure that are similar to those observed in humans, mice, and Drosophila, namely hyperactivity followed by uncoordination and sedation. We used in this report the nematode Caenorhabditis elegans as a model system to identify and characterize the genes that are affected by ethanol exposure and to link those genes functionally into an ethanol-induced gene network. By analyzing the expression profiles of all C. elegans ORFs using microarrays, we identified 230 genes affected by ethanol. While the ethanol response of some of the identified genes was significant at early time points, that of the majority was at late time points, indicating that the genes in the latter case might represent the physiological consequence of the ethanol exposure. We further characterized the early response genes that may represent those involved directly in the ethanol response. These genes included many heat shock protein genes, indicating that high concentration of ethanol acts as a strong stress to the animal. Interestingly, we identified two non-heat-shock protein genes that were specifically responsive to ethanol. glr-2 was the only glutamate receptor gene to be induced by ethanol. T28C12.4, which encodes a protein with limited homology to human neuroligin, was also specific to ethanol stress. Finally, by analyzing the promoter regions of the early response genes, we identified a regulatory element, TCTGCGTCTCT, that was necessary for the expression of subsets of ethanol response genes.

Section snippets

Effects of ethanol on C. elegans behavior

We first confirmed the relevance of ethanol action in C. elegans to that in higher organisms, as previously reported [24]. When treated with 7% ethanol, the nematode responded to this stress by showing higher motility and then started to slow its movement and eventually stopped moving (Fig. 1A). This series of action occurred within 10 min. When retrieved from ethanol, the animals began to recover from ataxia within 3 min, fully recovering by 10 min, indicating that the ethanol effect was fully

Strains and culture

C. elegans Bristol strain N2 was used as wild type. C. briggsae was obtained from the Caenorhabditis Genome Center. Worms were maintained by standard methods at 20°C, as previously described [30].

Ethanol microarray experiments of worms

To extract mRNA for use as probes for microarray analysis, wild-type worms were grown on roughly 80 NGM-Lite plates with 100 mm diameter, which were seeded with OP50-1, a streptomycin-resistant strain of Escherichia coli. The worms were harvested with S basal buffer, divided equally, and treated in

Acknowledgements

We thank S.H. Im and S.K. Jeon for technical support. We also thank Dr. Byung Soo Kim and his research group at the Department of Applied Statistics, Yonsei University, for their helpful discussion. We also thank Dr. A. Fire for the vectors and Caenorhabditis Genome Center for the nematode strain. This project was supported by a National Research Laboratory grant (M1-0318-00-0053) from the Ministry of Science and Technology, Korea.

References (52)

  • J.E. Mellem et al.

    Decoding of polymodal sensory stimuli by postsynaptic glutamate receptors in C. elegans

    Neuron

    (2002)
  • V. Costa et al.

    Mitochondrial superoxide dismutase is essential for ethanol tolerance of Saccharomyces cerevisiae in the post-diauxic phase

    Microbiology

    (1997)
  • M. Covarrubias et al.

    Ethanol selectively blocks a noninactivating K+ current expressed in Xenopus oocytes

    Proc. Natl. Acad. Sci. USA

    (1993)
  • K. Minami et al.

    Effects of ethanol and anesthetics on type 1 and 5 metabolic glutamate receptors expressed in Xenopus laevis oocytes

    Mol. Pharmacol.

    (1998)
  • D. Belelli et al.

    The interaction of the general anesthetic etomidate with the γ-aminobutyric acid type A receptor is influenced by a single amino acid

    Proc. Natl. Acad. Sci. USA

    (1997)
  • T. Miyakawa et al.

    Fyn-kinase as a determinant of ethanol sensitivity: relation to NMDA-receptor function

    Science

    (1997)
  • D.M. Lovinger et al.

    Ethanol inhibits NMDA-activated ion current in hippocampal neurons

    Science

    (1989)
  • I. Diamond et al.

    Cellular and molecular neuroscience of alcoholism

    Physiol. Rev.

    (1997)
  • S.J. Mihic

    Sites of alcohol and volatile anaesthetic action on GABA(A) and glycine receptors

    Nature

    (1997)
  • G.E. Homanics

    Gene knockout of the alpha6 subunit of the gamma-aminobutyric acid type A receptor: lack of effect on responses to ethanol, pentobarbital, and general anesthetics

    Mol. Pharmacol.

    (1997)
  • G.E. Homanics et al.

    Alcohol and anesthetic mechanisms in genetically engineered mice

    Front. Biosci.

    (1998)
  • J.C. Crabbe et al.

    Elevated alcohol consumption in null mutant mice lacking 5-HT1B serotonin receptors

    Nat. Genet.

    (1996)
  • R.A. Harris et al.

    Mutant mice lacking the gamma isoform of protein kinase C show decreased behavioral actions of ethanol and altered function of gamma-aminobutyrate type A receptors

    Proc. Natl. Acad. Sci. USA

    (1995)
  • L. Hilakivi-Clarke et al.

    Gonadal hormones and aggression-maintaining effect of alcohol in male transgenic transforming growth factor-alpha mice

    Alcohol Clin. Exp. Res.

    (1995)
  • T.J. Phillips et al.

    Alcohol preference and sensitivity are markedly reduced in mice lacking dopamine D2 receptors

    Nat. Neurosci.

    (1998)
  • Y. Ruan et al.

    Ethanol hypersensitivity and olfactory discrimination defect in mice lacking a homolog of Drosophila neuralized

    Proc. Natl. Acad. Sci. USA

    (2001)
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    Supplementary data associated with this article can be found at doi:10.1016/S0888-7543(03)00335-5.

    1

    These two authors equally contributed to this work.

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