An important mechanism in the up-regulation of cytochrome P-450 2A5 (CYP2A5, coumarin hydroxylase, Coh) is the stabilization of the corresponding mRNA; some evidence suggests that proteins binding to CYP2A5 mRNA may be involved in this stabilization. Here we report that pyrazole, a well known inducer of CYP2A5 and stabilizer of its message, enhances the binding of a set of proteins to 32P-labelled 3´-untranslated region (3´UTR) of CYP2A5 to give 32P-labelled bands of apparent molecular mass 37/39, 45/48 and 70/72 kDa after UV cross-linking/RNase cleavage; in addition, we found different proteins binding to other parts of CYP2A5 mRNA. The 70/72 kDa bands are also formed with the 3´UTR of c-jun. The inducible proteins are found in different cellular subfractions at different concentrations, with a maximum of five-fold induction of binding activity in microsomes. When a gel-mobility-shift assay was combined with UV cross-linking to resolve different pyrazole-inducible RNA–protein complexes into single RNA-binding protein bands, the smallest complex contained a double band of 37/39 kDa, 45/48 kDa bands, 70/72 kDa bands, and additional weaker bands at higher molecular masses (around 120 kDa). This composition was found also for all other complexes detected by gel-mobility-shift assay; occasionally, bands at higher molecular masses were also observed. The proteins of the smallest complex might therefore represent a core with which other proteins interact to build up larger complexes. Binding of proteins 37/39 kDa and 70/72 kDa was located to a 20-base loop and adjacent sequences in a 70 nt AU-rich region of the 3´UTR of the CYP2A5. Based on our previous evidence, this 70-nt sequence may play an important role in the stabilization and processing of the message.
Pyrazole-inducible proteins in DBA/2 mouse liver bind with high affinity to the 3′-untranslated regions of the mRNAs of coumarin hydroxylase (CYP2A5) and c-jun
- Views Icon Views
- PDF LinkPDF
- Share Icon Share
- Cite Icon Cite
Monika THULKE-GROSS, Manfred HERGENHAHN, Anne TILLOY-ELLUL, Matti LANG, Helmut BARTSCH; Pyrazole-inducible proteins in DBA/2 mouse liver bind with high affinity to the 3′-untranslated regions of the mRNAs of coumarin hydroxylase (CYP2A5) and c-jun. Biochem J 15 April 1998; 331 (2): 473–481. doi: https://doi.org/10.1042/bj3310473
Download citation file: