ERK and p38 pathways regulate amino acid signalling

Abstract

The ribosomal protein S6 kinase 1 (S6K1) is emerging as a common downstream target of signalling by hormones and nutrients such as insulin and amino acids. Here, we have investigated how amino acids signal through the S6K1 pathway. First, we found that a commercial anti-phospho-Thr389-S6K1 antibody detects an 80-90 kDa protein that is rapidly phosphorylated in response to amino acids. Unexpectedly, this phosphorylation was insensitive to both mTOR and PI-3 kinase inhibitors, and knockdown experiments showed that this protein was not S6K1. Looking for candidate targets of this phosphorylation, we found that amino acids stimulated phosphorylation of RSK and MSK kinases at residues that are homologous to Thr389 in S6K1. In turn, these phosphorylations required the activity of either p38 or ERK MAP kinases, which could compensate for each other. Moreover, we show that these MAP kinases are also needed for the amino acid-induced phosphorylation of S6K1 at Thr421/Ser424, as well as for that of S6K1 substrate, the S6 ribosomal protein. Consistent with these results, concomitant inhibition of p38 and ERK pathways also antagonised the well-known effects of amino acids on the process of autophagy. Altogether, these findings demonstrate a previously unknown role for MAP kinases in amino acid signalling.

Akt-dependent Activation of the Heart 6-Phosphofructo-2-kinase/Fructose-2,6-bisphosphatase (PFKFB2) Isoenzyme by Amino Acids

“…HEK293 cells were transfected using the calcium phosphate transfection system. Cell treatments such as serum starvation, amino acid deprivation, and insulin or amino acid stimulation were performed as reported previously (6,11). For experiments with insulin or the phorbol ester PMA, cells were deprived of serum overnight and then incubated with 200 nM insulin or 10 nM PMA for 30 min.…”

Transfer-RNA-mediated enhancement of ribosomal proteins S6 kinases signaling for cell proliferation

“…24 , 30 Initially, we assumed that the p90 protein that we were monitoring was the p85 S6K; however, we had to rule out p85 S6K due to its reactivity to mTOR, unlike the p90 protein of our interest. While searching for other candidates, we came upon a report on RSK and MSK.…”

“…4A). 24 They also investigated the induction of phosphorylation of RSK and MSK under amino acid supplementation, although they did not consider the effect of tRNA overexpression.

10.1080/15476286.2017.1356563-f0004Figure 4.RSK1 and MSK2 were the 90-kDa RPS6K phosphorylated by tRNA Leu CAG overexpression.

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“…4A). 24 , 39 We transfected p85 S6K, 4 RSKs, and 2 MSKs into HEK 293T cells, along with tRNA Leu , and monitored their levels of phosphorylation. We found that p85 S6K, RSK1, and MSK2 were ideal candidates for the p90 protein; the tRNA Leu -mediated activation of these proteins was confirmed by detection with specific antibodies (Fig.…”

“…22 RPS6K shares a hydrophobic motif and representative phosphorylation site among the family members and it comprises 4 RSKs (ribosomal S6 kinases, RSK1–4), 2 MSKs (mitogen-and stress-activated protein kinases, MSK1–2), S6K1 and S6K2 proteins. 23 , 24 Among them, p70 S6K and p85 S6K are two splice variants of S6K1, which have been well studied due to the roles under mTOR (mammalian target of rapamycin) signaling. 25 …”