DNA damaging agents increase the stability of interleukin-1 alpha, interleukin-1 beta, and interleukin-6 transcripts and the production of the relative proteins.

M Mallardo,V Giordano,E Dragonetti,G Scala,I Quinto

doi:10.1016/s0021-9258(17)36550-x

M Mallardo, V Giordano + Show 3 more

Open Access

https://doi.org/10.1016/s0021-9258(17)36550-x

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Abstract

In this study, we addressed the question of whether carcinogens affected the expression of interleukin-1 alpha (IL1 alpha), interleukin-1 beta (IL1 beta), and interleukin-6 (IL6) genes and the production of the relative proteins. Primary cultures of human monocytes were exposed to the alkylating agents mitomycin C (Mit C), methylmethane sulfonate (MMS), and ethylmethane sulfonate (EMS) and tested for the production of IL1 alpha, IL1 beta, and IL6 proteins, as well as for the expression of IL1 alpha, IL1 beta, and IL6 transcripts. The production of IL1 beta and IL6 was significantly augmented by all the three chemicals after 24-48 h of treatment. IL1 alpha was also increased by Mit C and MMS. By Northern blotting analysis, the increased expression of IL1 alpha, IL1 beta, and IL6 genes was shown to occur at 30 min of Mit C and MMS treatment and to decline after 8 h. Similarly, EMS up-regulated the expression of IL1 beta and IL6 genes. The mutagen-mediated increase in interleukin transcripts did not require de novo protein synthesis, and it was due to the enhanced half-life of IL1 alpha, IL1 beta, and IL6 mRNAs rather than to the increased rate of gene transcription. These results suggest that carcinogens, in addition to causing DNA mutations and rearrangements, may also affect cell growth and differentiation by enhancing the expression of cytokine genes.

Highlights

Pression ofILla, ILlp, andIL6 genes was shown to occurDNA damaging agents arechemical or physical agents that at 30 min of Mit C and MMS treatment and to decline cause gene mutations and rearrangements andpromote carciafter 8 h
These results suggest that induce a tumorigenic phenotypeby deregulating theexpression carcinogens, in addition to causingDNA mutations and of cellular genesinvolved in cell growth and differentiation
Pellets were resuspended in 10 ml of lysis buffer without Nonidet P-40, layered onto 5 ml of isolation buffer (15 m Tris-HC1, pH 7.4, 60mM potassium chloride, 15 M M S (mM) sodium chloride,15 mM spermine, 0.5 mM spermidine, 15 mM p-mercaptoethanol, 30% saccharose), and centrifugeda t 500 x g for 15 min at 4 "C

Summary

Gene Regulation in Genotoxic Distress

(density = 1077)on Ficoll Hypaque (Flow,Thame, United Kingdom) and which were treated with PMMit C, 0.5 nm MMS, 5 mM EMS, 100 pl Percoll (Pharmacia, Uppsala, Sweden) centrifugatioans described [23]. Cells were culturedin RPMI 1640(Life collected by centrifugation and tested for the presence of ILla, ILIp, and IL6 molecules by ELISA. Onocytes were resuspended at 5 x 106/mlin RPMI 1640 with 10% fetal calf serum and divided in aliquots to be treated or left untreated. 87p~ Mit C, 0.25-3 mM MM6S2, 0.25-10 mM EMS). Mit C238 of chemical-treated or untreated monocyte cultures were collected at MMS differenttimesandtested for the presence of ILla, ILlp, and IL6. E1M34S molecules by enzyme-linked immunosorbent assay (R & D Systems) according tothe manufacturer's instructionsA. Mega).Thehybridizationwasperformedwith the 32P-labeledprobe (0.8-1 x lo cpdpg ) i n 1M sodium chloride, 10% dextran sulfate, 1%

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RESULTS

DISCUSSION