Role of deadenylation and AUF1 binding in the pH-responsive stabilization of glutaminase mRNA.

Abstract

During chronic metabolic acidosis, increased expression of renal glutaminase (GA) results from selective stabilization of the GA mRNA. This response is mediated by a direct repeat of an 8-base adenylate-uridylate (AU) sequence that binds zeta-crystallin and functions as a pH response element (pH-RE). A tetracycline-responsive promoter system was developed in LLC-PK(1)-F(+) cells to perform pulse-chase analysis of the turnover of a chimeric beta-globin (betaG) mRNA that contains 960 bp of the 3'-UTR of GA mRNA including the pH-RE. The betaG-GA mRNA exhibits a 14-fold increase in half-life when the LLC-PK(1)-F(+) cells are transferred to acidic medium. RNase H cleavage and Northern blot analysis of the 3'-ends established that rapid deadenylation occurred concomitantly with the rapid decay of the betaG-GA mRNA in cells grown in normal medium. Stabilization of the betaG-GA mRNA in acidic medium is associated with a pronounced decrease in the rate of deadenylation. Mutation of the pH-RE within the betaG-GA mRNA blocked the pH-responsive stabilization, but not the rapid decay, whereas insertion of only a 29-bp segment containing the pH-RE was sufficient to produce both a rapid decay and a pH-responsive stabilization. Various kidney cells express multiple isoforms of AUF1, an AU-binding protein that enhances mRNA turnover. RNA gel-shift assays demonstrated that the recombinant p40 isoform of AUF1 binds to the pH-RE with high affinity and specificity. Thus AUF1 may mediate the rapid turnover of the GA mRNA, whereas increased binding of zeta-crystallin during acidosis may inhibit degradation and result in selective stabilization.

• Authors: Jill M Schroeder, Hend Ibrahim, Lynn Taylor, Norman P Curthoys
• Journal: American journal of physiology. Renal physiology (Am J Physiol Renal Physiol) Vol. 290 Issue 3 Pg. F733-40 (Mar 2006) ISSN: 1931-857X [Print] United States
• PMID: 16219914 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)

Chemical References

• Gentamicins
• Heterogeneous Nuclear Ribonucleoprotein D0
• Heterogeneous-Nuclear Ribonucleoprotein D
• Hnrnpd protein, rat
• RNA, Messenger
• beta-Crystallins
• Hygromycin B
• Adenosine Monophosphate
• antibiotic G 418
• Ribonuclease H
• Glutaminase

Topics

• Adenosine Monophosphate (metabolism)
• Animals
• Cell Line
• Cytosol (physiology)
• Gentamicins (pharmacology)
• Glutaminase (genetics)
• Heterogeneous Nuclear Ribonucleoprotein D0
• Heterogeneous-Nuclear Ribonucleoprotein D (metabolism)
• Hydrogen-Ion Concentration
• Hygromycin B (pharmacology)
• Kidney Cortex (physiology)
• Male
• Mutagenesis
• RNA, Messenger (genetics)
• Rats
• Rats, Sprague-Dawley
• Ribonuclease H (metabolism)
• beta-Crystallins (metabolism)