Rapid induction of rat liver S14 gene transcription by thyroid hormone

D B Jump

doi:10.1016/s0021-9258(18)83799-1

Abstract

The mRNA coding for the rat liver S14 protein (Mr 17,000; pI 4.9) is rapidly induced by triiodothyronine (T3) (Jump, D. B., Narayan, P., Towle, H. C., and Oppenheimer, J. H. (1984) J. Biol. Chem. 259, 2789-2797). In an effort to define the molecular basis for the rapid increase in mRNAS14, the in vitro run-on activity and chromatin structure of the S14 gene was examined. Following injection of hypothyroid rats with a receptor-saturating dose of T3, a 5-min lag period preceded a rapid increase in S14 gene transcription. S14 transcriptional activity was induced 3.8-fold within 15 min and reached nearly 70% of the maximal 9-fold induction within 60 min of T3 administration. Hepatic mRNAS14 levels were induced 2.4-, 19-, and 24-fold within 15 min, 4 h, and 24 h, respectively. Thus, the rapid T3-mediated induction of mRNAS14 was due, in large part, to a rapid and apparent direct effect of T3 on S14 transcriptional activity. Analysis of S14 chromatin structure showed that the Hss-3 DNase I hypersensitive site located 3.3 kilobases (kb) upstream from the hepatic S14 cap site was present in both euthyroid and hyperthyroid states, but either absent or present at diminished DNase I sensitivity in the hypothyroid state. However, within 5 min of T3 administration to hypothyroid rats, the Hss-3 DNase I hypersensitive site was significantly induced. The induction of this structure preceded T3 induction of S14 gene transcription. Other DNase I hypersensitive sites located adjacent to the S14 cap site at -65 to -265 base pairs (Hss-1) or upstream at -1.3 kb (Hss-2), -2.1 kb (Hss-3'), -5.3 kb (Hss-4), and -6.2 kb (Hss-5) remained unaffected by changes in S14 gene transcription. The rapid T3 effect on the Hss-3 DNase I hypersensitive site may reflect the presence of T3 receptors in the vicinity of this chromatin locus. Modification of chromatin structure in the vicinity of the Hss-3 site may be an important antecedent event for T3-mediated induction of S14 gene transcription.

Highlights

The mRNA coding for the rat liver 514 protein
One DNase I hypersensitive site located adjacent to theS14 cap site (Hss-1,65 to -265 bp)’ forms prior to initiation of gene transcription tion. 514 transcriptional activity waisnduced 3.8-fold while a second site located 3.3 kb upstream from the S14 cap within 15min and reached nearly 70%of the maximal site (Hss-3) forms at weaning when S14 gene transcription
Chromatin structure showed that the Hss-3 DNase I The presence of DNase I hypersensitive sites within chrohypersensitive sitelocated 3.3 kilobases upstream matin reflects a local disruption of nucleosomal periodicity fromthehepatic 514 cap site was present in both due the interaction of DNA-binding proteins euthyroid and hyperthyroid states, but either absent or present at diminished DNase I sensitivity in the hypothyroid state

Summary

To whom correspondence should be addressed

Dept. of Physi- HEPES, 4-(2-hydroxyethyl)-l-piperazineethanesulfonaicid; EGTA, ology, 214 Giltner Hall, Michigan State University, East Lansing, MI [ethylenebis(oxyethylenenitrilo)]tetraaceticacid; SDS, sodium dode-. This at 37 "C) followedby extraction with pheno1:chloroform (1:l)and washing procedure reduces background on the Southern blots. In Nuclear Storage Buffer (40% glycerol; 75 mM HEPES, pH 7.5; 60 The Southern blots were hybridized under the conditions described mM KCl; 15 mM NaCl; 0.15 mM spermidine; 0.5 mM spermine; 0.5 previously (7). Blots were to a transcription reaction in which the final reactant concentrations washed a t room temperature in 2 X SSC, 0.1% SDS for 30 min were: 25% glycerol; 75mM HEPES, pH 7.5; 3 mM MgC12; 0.1 M KC1; followed by four cycles of washing at 60 "C in 0.1 X SSC, 0.1% SDS.

RESULTS

HYPOTHYROID HYPERTHYROID III abcd abcd abcd

Findings

DISCUSSION