Hydrocortisone reveals its action by activation of transcription of specific genes that is achieved by binding of hormones nuclear receptors (NRs) to cognate sites on DNA and recruiting coactivators of basal transcription machinery. NRs are able to overcome restricting conformation of chromatin by altering chromatin-remodeling machinery (chromatin-remodeling protein complexes, histone and nonhistone modifying enzymes). However, a possibility arise that genomic DNA accessibility to NRs as well as chromatin packaging pattern in nuclei implicates complex processes which involve regulation of DNA modifying enzymes activities.
Whether hydrocortisone induced decondensation of chromatin alter DNA accessibility in nuclei upon exogenously applied DNAse I and cause eventual changes in endogenous Ca-Mg endonuclease-depended fragmentation become a focus of our research.
Experiments were conducted in vivo. Hydrocortisone 5 mg/100g body wt was administrated by intraperitonial injection. Hormone treated and control animals were sacrificed under light ether anesthesia by decapitation .
Nuclei were isolated from liver according to Blobel&Potter (1) and by Hewish & Burgoyne (2), chromatin by Umansky (3) . The DNA was prepared from isolated nuclei by successive treatments with 0.5 mg/ml RNase A and 0.5 mg/ml proteinase K. The resulting DNA preparation was subjected to electrophoresis in 1,6% agarose gels. DNA fragmentation, visualized by ethidium bromide staining, was examined in photographs taken under UV-illumination.
The data obtained upon using the system of isolated nuclei as substrate revealed that hydrocortisone cause suppression of Ca-Mg endonuclease activity in nuclei as well as significantly hindered activity of exogenously applied DNAase 1. An assumption can be derived that hydrocortisone treatment enhanced liver nuclei resistance upon endogenous and exogenous nucleases.
1. Blobel G., Potter V.R.//Science 1966,v.154,p.76-79
2. Hewish D.R.,Bugoyne L.A.// Biochem.Biophys.Acta 1973,v.52,N2,p.475-481
3. Umansky S.R., Kovalev Y.,Tokarskaya V.Y.//Biochem.Biophys.Acta 1975,v.383, N3, p.242-248
03 - 05 Nov 2003
Society for Endocrinology