A multifunctional domain of the calcium-responsive transactivator (CREST) that inhibits dendritic growth in cultured neurons

Abstract

The calcium-responsive transactivator (CREST) is targeted to nuclear bodies and is required for the normal development of neuronal dendritic trees. Here we report the identification of a multifunctional domain (MFD) of CREST that is involved in transcription transactivation, nuclear body targeting, and dimerization. MFD is located near the C terminus of CREST from amino acid 251 to 322 and is required and sufficient for CREST homodimerization. When fused with a GAL4 DNA-binding domain, MFD was effective in transcription transactivation of a luciferase reporter system. A C-terminal 339-401 amino acid sequence of CREST was shown to contain a nuclear localization signal (NLS), which was able to direct a yellow fluorescence protein (YFP) to nucleus. A CREST deletion mutant containing both the MFD and NLS, which spanned the C-terminal amino acid sequence 251-401, was able to target YFP to the nucleus and nuclear bodies. However, MFD alone failed to target YFP and was largely cytosolic. The addition of a SV40 NLS to MFD domain restored nuclear body targeting. When YFP-MFD was expressed in cultured rat embryonic cortical neurons, it was effective in inhibiting depolarization-induced dendritic growth, suggesting that CREST dimerization may be necessary for its function in neuronal dendritic development. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.