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Regulation of two JunD isoforms by Jun N-terminal kinases
1Department of Cell Biology and Biochemistry and the Southwest Cancer Center at University Medical Center, Texas Tech University Health Sciences Center, Lubbock, Texas 79430, USA.
Abstract:
The JunD transcription factor is one member of the Jun family of proteins that also includes c-Jun and JunB. Although c-Jun can function to promote cell proliferation and can cooperate with other oncogenes to transform cells, JunD slows proliferation of fibroblasts and antagonizes transformation by activated ras. Two isoforms of JunD, a full-length isoform containing 341 amino acids (JunD-FL) and a truncated isoform lacking 48 amino acids at the N terminus (Delta JunD), are generated through utilization of two translation start sites within a single mRNA. Here we show that both isoforms of JunD are phosphorylated by Jun N-terminal kinases (JNKs) at three identical residues and that both contain a docking domain that specifically binds JNKs. The JunD-FL isoform binds to and is phosphorylated by JNK more efficiently than Delta JunD in vitro; correspondingly, JunD-FL is a more potent transcriptional activator than Delta JunD. Although increased JNK signaling can activate both JunD isoforms, mutating either the JNK docking domain or the target JNK phosphorylation sites blocks this activation. These results identify two distinct isoforms of JunD with differential responses to JNK signaling pathways.
Insights
JunD transcription factor isoforms, JunD-FL and Delta JunD, exhibit differential responses to Jun N-terminal kinases (JNKs). JunD-FL shows greater JNK binding, phosphorylation, and transcriptional activation, highlighting isoform-specific JNK signaling regulation.
Area of Science:
- Molecular Biology
- Cell Signaling
- Transcription Factors
Background:
- JunD is a transcription factor in the Jun family, contrasting with c-Jun's role in cell proliferation.
- JunD antagonizes oncogene-induced transformation, suggesting a tumor-suppressive function.
- Two JunD isoforms, JunD-FL and Delta JunD, arise from a single mRNA via alternative translation start sites.
Purpose of the Study:
- To investigate the differential regulation of JunD isoforms by Jun N-terminal kinases (JNKs).
- To elucidate the roles of specific phosphorylation sites and a JNK docking domain in JunD isoform activation.
Main Methods:
- In vitro phosphorylation assays using Jun N-terminal kinases (JNKs) on JunD-FL and Delta JunD.
- Analysis of JNK binding affinity and transcriptional activation potential of both JunD isoforms.
- Site-directed mutagenesis of JNK phosphorylation sites and the JNK docking domain.
Main Results:
- Both JunD isoforms are phosphorylated by JNKs at three identical residues and possess a JNK docking domain.
- JunD-FL binds and is phosphorylated by JNK more efficiently than Delta JunD.
- JunD-FL demonstrates more potent transcriptional activation than Delta JunD, with activation dependent on the docking domain and phosphorylation sites.
Conclusions:
- JunD isoforms display distinct responses to JNK signaling pathways.
- Differential JNK interaction and phosphorylation contribute to the functional divergence of JunD isoforms.
- These findings offer insights into the nuanced regulation of gene expression by JunD isoforms in cellular processes.