MKP-1 inhibits high NaCl-induced activation of p38 but does not inhibit the activation of TonEBP/OREBP: Opposite roles of p38α and p38δ

Abstract

High NaCl rapidly activates p38 MAPK by phosphorylating it, the phosphorylation presumably being regulated by a balance of kinases and phosphatases. Kinases are known, but the phosphatases are uncertain. Our initial purpose was to identify the phosphatases. We find that in HEK293 cells transient overexpression of MAPK phosphatase-1 (MKP-1), a dual-specificity phosphatase, inhibits high NaCl-induced phosphorylation of p38, and that overexpression of a dominant negative mutant of MKP-1 does the opposite. High NaCl lowers MKP-1 activity by increasing reactive oxygen species, which directly inhibit MKP-1, and by reducing binding of MKP-1 to p38. Because inhibition of p38 is reported to reduce hypertonicity-induced activation of the osmoprotective transcription factor, TonEBP/OREBP, we anticipated that MKP-1 expression might also. However, overexpression of MKP-1 has no significant effect on Ton EBP/OREBP activity. This paradox is explained by opposing effects of p38␣ and p38␦, both of which are activated by high NaCl and inhibited by MKP-1. Thus, we find that overexpression of p38␣ increases high NaCl-induced TonEBP/ OREBP activity, but overexpression of p38␦ reduces it. Also, siRNAmediated knockdown of p38␦ enhances the activation of TonEBP/ OREBP. We conclude that high NaCl inhibits MKP-1, which contributes to the activation of p38. However, opposing actions of p38␣ and p38␦ negate any effect on TonEBP/OREBP activity. Thus, activation of p38 isoforms by hypertonicity does not contribute to activation of TonEBP/OREBP because of opposing effects of p38␣ and p38␦, and effects of inhibitors of p38 depend on which isoform is affected, which can be misleading.hypertonicity ͉ reactive oxygen species ͉ transcription ͉ HEK293 cells ͉ mIMCD3 cells

Rac1/osmosensing scaffold for MEKK3 contributes via phospholipase C-γ1 to activation of the osmoprotective transcription factor NFAT5

“…p38α contributes to hypertonicity-induced activation of TonEBP/OREBP (9), but full understanding of this interaction has been complicated by opposite effects of p38α and p38δ; p38α increases TonEBP/OREBP activity and p38δ decreases it, but the phosphospecific antibodies commonly used to measure p38 activity do not directly distinguish between p38α activity and p38δ activity (6). In addition, some of the ways of inhibiting p38 inhibit both p38α and p38δ, resulting in no net effect on TonEBP/OREBP activity.…”

“…Because MKK3 activates both p38α and p38δ (7), inhibition of MKK3 reduces their opposing effects, resulting in no net change of TonEBP/OREBP activity. Another example is MAPK phosphatase 1 (MKP-1), which also inhibits both p38α and p38δ without any effect on TonEBP/OREBP activity (6). SB203580 is an often-used inhibitor of p38.…”

“…In this article, we will continue to refer to "p38" unless p38α and p38δ are distinguished. Hypertonicity activates p38α via the upstream kinase MKK3 (MAP2K3) or MKK6 (MAP2K6) (7), and p38α and MEKK3 (MAP3K3) contribute to hypertonicity-induced activation of TonEBP/OREBP (6,8,9). Thus, p38α contributes to TonEBP/OREBP activity.…”

“…The stress-activated MAP kinase p38 has been extensively studied in this regard, but understanding its role has been complicated because hypertonicity activates two different p38s, namely p38α (MAPK14) and p38δ (MAPK13), which have opposite effects on TonEBP/OREBP activity: p38α increases TonEBP/OREBP activity and p38δ decreases it (6). Furthermore, the phosphospecific antibodies commonly used to measure p38 activity do not directly distinguish between p38α activity and p38δ activity, nor do some forms of inhibition that have been used (6). In this article, we will continue to refer to "p38" unless p38α and p38δ are distinguished.…”

p38δ mitogen-activated protein kinase regulates skin homeostasis and tumorigenesis

“…siRNA-mediated knockdown of p38δ enhances hypertonicity-induced transcriptional activation of the osmoprotective transcription factor TonEBP/OREBP, while overexpresson of p38δ reduces it. 22 Notably, p38α is shown to have an opposite effect on the activity of this transcription factor. The physiological implications of the differential effects of p38δ and p38α isoforms on TonEBP/OREBP are presently being investigated.…”