Temperature-sensitive mRNA degradation is an early event in hepatocyte de-differentiation
- 15 December 1997
- journal article
- Published by Portland Press Ltd. in Biochemical Journal
- Vol. 328 (3) , 937-944
- https://doi.org/10.1042/bj3280937
Abstract
The isolation and culture of metabolically active hepatocytes by proteolytic digestion of the extracellular matrix of the liver results in the transcriptional silencing of liver-specific genes encoding cytochromes P-450 (CYP) and albumin together with an induction of cellular RNase activity. The levels of albumin mRNA are maintained in cultured hepatocytes at similar levels to that present in the intact liver for at least 24 h, whereas the major constitutively expressed CYP2C11 mRNA is rapidly degraded. Hepatocytes heat-shocked at 40 °C during the isolation procedure (which results in an induction of heat-shock protein mRNA species) blocks the increase in RNase activity and abrogates the loss of CYP2C11 mRNA for at least 4 h. Cycloheximide-dependent inhibition of protein synthesis blocks the temperature-dependent induction of heat-shock proteins without affecting the protection afforded to CYP2C11 mRNA, indicating that CYP2C11 mRNA levels are not directly dependent on heat-shock protein induction and suggesting that the induction of RNase activity might be responsible for the specific loss of CYP2C11 mRNA in hepatocytes isolated at 37 °C. Differential rates of degradation of CYP2C11 transcribed in vitro and of albumin mRNA are observed in the presence of cellular extracts from cultured hepatocytes isolated at 37 °C (which have maximally induced levels of cellular RNase activity) but not in comparable extracts from cultured hepatocytes isolated at 40 °C, supporting the hypothesis that an RNase activity is induced in culture that specifically degrades CYP2C11 mRNA but not albumin mRNA. These results suggest that an early event in hepatocyte de-differentiation involves the induction of RNase activity in addition to transcriptional silencing of liver-specific genes and that the induced RNase activity demonstrates specificity within liver-specific gene products.Keywords
This publication has 25 references indexed in Scilit:
- P450 superfamily: update on new sequences, gene mapping, accession numbers and nomenclaturePharmacogenetics, 1996
- Review : Heterogeneity of cytochrome P450 and its toxicological significanceHuman & Experimental Toxicology, 1995
- Alteration of Transcription Factor mRNAs during the Isolation and Culture of Rat Hepatocytes Suggests the Activation of a Proliferative Mode Underlies Their DedifferentiationBiochemical and Biophysical Research Communications, 1993
- Evidence that the loss of rat liver cytochrome P450 in vitro is not solely associated with the use of collagenase, the loss of cell-cell contacts and/or the absence of an extracellular matrixBiochemical Pharmacology, 1992
- The maintenance of cytochrome P-450 in rat hepatocyte culture: Some applications of liver cell cultures to the study of drug metabolism, toxicity and the induction of the P-450 systemChemico-Biological Interactions, 1990
- Molecular genetics of the P-450 superfamilyPharmacology & Therapeutics, 1990
- Ethanol-, fasting-, and aceton-inducible cytochromes P-450 in rat liver: regulation and characteristics of enzymes belonging to the IIB and IIE gene subfamiliesBiochemistry, 1988
- Regulation of rat hepatic cytochrome P-450: age-dependent expression, hormonal imprinting, and xenobiotic inducibility of sex-specific isoenzymesBiochemistry, 1985
- Culture shockExperimental Cell Research, 1984
- Attachment of rat hepatocytes to plastic substrata in the absence of serum requires protein synthesisBiochemical and Biophysical Research Communications, 1979