Lithium Can Relieve Translational Repression of TOP mRNAs Elicited by Various Blocks along the Cell Cycle in a Glycogen Synthase Kinase-3- and S6-Kinase-independent Manner

Miri Stolovich,Tal Lerer,Yoav Bolkier,Hannah Cohen,Oded Meyuhas

doi:10.1074/jbc.m412434200

Abstract

TOP mRNAs are translationally controlled by mitogenic, growth, and nutritional stimuli through a 5'-terminal oligopyrimidine tract. Here we show that LiCl can alleviate the translational repression of these mRNAs when progression through the cell cycle is blocked at G(0), G(1)/S, or G(2)/M phases in different cell lines and by various physiological and chemical means. This derepressive effect of LiCl does not involve resumption of cell division. Unlike its efficient derepressive effect in mitotically arrested cells, LiCl alleviates inefficiently the repression of TOP mRNAs in amino acid-deprived cells and has no effect in lymphoblastoids whose TOP mRNAs are constitutively repressed even when they are proliferating. LiCl is widely used as a relatively selective inhibitor of glycogen synthase kinase-3. However, inhibition per se of this enzyme by more specific drugs failed to derepress the translation of TOP mRNAs, implying that relief of the translational repression of TOP mRNAs by LiCl is carried out in a glycogen synthase kinase-3-independent manner. Moreover, this effect is apparent, at least in some cell lines, in the absence of S6-kinase 1 activation and ribosomal protein S6 phosphorylation, thus further supporting the notion that translational control of TOP mRNAs does not rely on either of these variables.

Highlights

Their mass) or when amino acid-starved cells are refed [4, 5]
We show that LiCl can alleviate the translational repression of these mRNAs when progression through the cell cycle is blocked at G0, G1/S, or G2/M phases in different cell lines and by various physiological and chemical means
Our results clearly show that translational activation of TOP mRNAs by LiCl cannot be attributed to the inhibition of GSK-3, regardless of the nature of the examined cell line, the means used for mitotic arrest, or the assay applied for monitoring GSK-3 activity

Summary

EXPERIMENTAL PROCEDURES

Cell Culture and DNA Transfection—HEK 293 cells were grown and transfected as well as starved for serum or amino acids as described previously [4, 5, 21]. P1798.C7 mouse lymphosarcoma cells were grown as suspension cultures and mitotically arrested by treatment with 0.1 ␮M dexamethasone (Sigma) for 24 h and hormonally withdrawn as described previously [7]. NIH 3T3 mouse fibroblasts were grown as monolayer [7] and arrested by either 24 h treatment with 10 mM hydroxyurea (Sigma) or by splitting a confluent culture 1:2 and maintaining the cells for 5 days without further splitting (contact inhibition). HeLa 229 cells were grown as monolayer as described previously [6] and arrested by a 24-h treatment with 15 ␮M nocodazole (Sigma). Polyclonal anti-rpS6 antibody was raised against a peptide corresponding to non-phosphorylatable residues 185– 205 of mouse rpS6 (LQHKRRRIALKKQRTKKNKEE) and was kindly provided by Cell Signaling Technology. Tau-5 is a phosphorylationindependent tau antibody (Ref. 26 and references therein)

RESULTS

Lithium Can Relieve Translational Repression of TOP mRNAs

DISCUSSION