Regulation of dusp4 by ERK‐dependent and ‐independent Mechanisms in Adipocytes
Authors: Guo Wei , Ferguson Bradley S. , Hopkins Robin G. , Morrison Ron F. 
Published: November 2023

Mitogen‐activated protein kinases (MAPKs; ERK, JNK, p38) play a central role in a wide variety of cellular processes including proliferation, differentiation, inflammation, stress response and apoptosis, where the functional diversity has been attributed to magnitude, duration and compartmentalization of MAPK activity. While numerous studies have examined upstream signaling for MAPK activation, few have addressed how MAPK deactivation alters biological outcomes. Evidence now supports potential roles for MAPK‐specific, dual‐specificity phosphatases (DUSPs) in the deactivation of MAPK signaling and modulation of obesity‐associated metabolic dysfunction. We initially examined the expression profile of the 10 known DUSPs in insulin‐responsive tissues of genetic‐and diet‐induced obese mice. We previously observed that dusp4 was significantly more abundant in white adipose tissue relative to liver, skeletal muscle, and heart in lean mice and that this phosphatase was induced during early stages of obesity and repressed during later stages correlating with the onset of obesity‐induced inflammation. The objective of this investigation was to determine regulatory mechanisms of dusp4 expression by ERK, JNK and/or p38 activity in adipocytes. We demonstrated that induction of 3T3‐L1 adipocyte differentiation lead to immediate, robust phosphorylation of ERK on phosphosites that are known to be both necessary and sufficient for ERK activity. This phosphorylation peaked with 10 mins of stimulation and decayed to basal levels within 6 hrs of stimulation where steady state phosphorylation of ERK remained throughout the 8 day process of differentiation. Dusp4 mRNA and protein levels kinetically increased in a feedback manner where expression was inversely proportional to ERK phosphorylation supporting the premise that ERK activation resulted in dusp4 expression that suppressed ERK phosphorylation. Surprisingly, we also observed a second peak of dusp4 expression at 4 days of differentiation that did not correlate with ERK phosphorylation. Pretreating cells with the ERK inhibitor, U0126 abolished early dusp4 expression that correlated with ERK phosphorylation and was without effect on subsequent dusp4 expression that did not correlate with ERK phosphorylation. To further examine regulation of dusp4 in adipocytes, we used phorbol 12‐myristate 13‐acetate (PMA) and cobalt chloride (CoCl2) as potent inducers of MEK/ERK activity. Both PMA and CoCl2 stimulation resulted in robust and sustained ERK phosphorylation up to 8 hours. Following ERK phosphorylation, we report that dusp4 mRNA and protein kinetically increased in a manner similar to that observed during the initial peak of differentiation. Pretreatment with U0126 suppressed ERK phosphorylation and abolished dusp4 expression. Collectively, these results demonstrate ERK‐dependent induction of dusp4 expression which supports a feedback hypothesis limiting magnitude and duration of ERK activity. These data also highlight a unique ERK‐independent mechanism for dusp4 expression during adipocyte differentiation. Loss‐of‐function studies are underway to examine the function of dusp4 in regulating ERK mediated biological outcomes in adipocytes.Support or Funding InformationSupported by NIH‐NIDDK (R15‐DK082799)