A multifunctional domain of the calcium-responsive transactivator (CREST) that inhibits dendritic growth in cultured neurons.
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.
Pradhan
Anuradha
A
Department of Pathology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73190, USA.
Liu
Yuechueng
Y
D000818:Animals; D001426:Bacterial Proteins; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D003712:Dendrites / Q000378:metabolism*; D019281:Dimerization; D017353:Gene Deletion; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D008164:Luminescent Proteins; D008957:Models, Genetic; D009154:Mutation; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D016716:PC12 Cells; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D013997:Time Factors; D015534:Trans-Activators / Q000737:chemistry*; D015533:Transcriptional Activation
D001426:Bacterial Proteins; D008164:Luminescent Proteins; C502810:SS18L1 protein, human; C481041:Ss18l1 protein, rat; D015534:Trans-Activators; C054562:yellow fluorescent protein, Bacteria; D008156:Luciferases