Expression of tumor-associated aldehyde dehydrogenase gene in rat hepatoma cell lines.
Significant changes in aldehyde dehydrogenase (ALDH) activity occur during rat hepatocarcinogenesis in vivo. To compare the structure and expression of the tumor aldehyde dehydrogenase gene in rat hepatoma cell lines and normal rat liver, several rat hepatoma cell lines, including HTC, H4-II-EC3, JM2, McA-RH7777, and four lines established in this laboratory have been examined for T-ALDH gene expression using a tumor ALDH complementary DNA. Northern blot analysis of polyadenylate-containing RNA from log-phase cells and normal rat liver with T-ALDH complementary DNA indicates production of a single major 1.7-kilobase transcript in the high activity lines HTC, JM2, RLT-2M, RLT-3C, RLT-9F, and intermediate activity line RLT-5G. There is a direct correlation between expression of T-ALDH enzyme activity and the amount of 1.7-kilobase transcript. S1 nuclease protection experiments confirm that there is only one major T-ALDH transcript in the high activity lines. Thus, cell line differences in T-ALDH activity are reflected in the level of a single T-ALDH transcript. Southern analysis was used to identify the T-ALDH gene in genomic DNA. The results indicate that no significant amplification or rearrangement of the T-ALDH gene has occurred in these hepatoma cells. DNA methylation has been proposed to play an important role in gene expression. Genomic DNA from HTC, JM2, McA-RH7777, H4-II-EC3, RLT-2M, RLT-9F, RLT-3C, RLT-5G, rat embryo and normal rat liver were digested with MspI and HpaII to examine methylation patterns. A digestion pattern consistent with hypomethylation was detected only in DNA from the high T-ALDH activity cell lines HTC, JM2, RLT-2M, and RLT-9F. This suggests that constitutive expression of T-ALDH in the hepatoma cells is related to changes in DNA methylation patterns.
Lin
K H
KH
Department of Biology, University of Alabama, Tuscaloosa 35487.
Brennan
M D
MD
Lindahl
R
R
D000444:Aldehyde Dehydrogenase / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D004247:DNA / Q000032:analysis; D015247:Deoxyribonuclease HindIII / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D005786:Gene Expression Regulation; D008114:Liver Neoplasms, Experimental / Q000201:enzymology* / Q000235:genetics; D051381:Rats; D011919:Rats, Inbred Strains
D004247:DNA; D000444:Aldehyde Dehydrogenase; D015247:Deoxyribonuclease HindIII; C024388:CTGCAG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific