Abstract
The calcium-responsive transactivator (CREST) is required for the normal development of neuronal dendritic trees. Here we report that CREST is localized to sub-nuclear structures in the rat neuroendocrine pheochromocytoma PC12 cells. A yellow fluorescence protein-CREST fusion protein was expressed in HEK 293 and PC12 cells and the recombinant protein was exclusively targeted to nuclear bodies. A similar result was obtained with a Flag-tagged CREST. Deleting the N-terminal 148 or the C-terminal 79 amino acid sequences had no effect on targeting, whereas removing 164 amino acid residues from the C-terminus abolished nuclear body localization. We found that CREST did not co-localize with promyelocytic leukaemia oncoprotein (PML) body and was not targeted to PML bodies. Overexpression of CREST markedly increased the number of nuclear bodies positive for the histone acetyltransferase CREB binding protein (CBP). Double immunofluorescence staining of Flag-CREST and CBP suggested that CREST and CBP had a high degree of co-localization within the nuclear bodies. Deletion of the CBP binding domain of CREST inhibited the recruitment of CBP to CREST nuclear bodies. These results suggest that CBP recruitment to nuclear bodies by CREST may play an important role in CREST-mediated calcium-responsive transactivation, and CREST nuclear body may function as an assembly site for activators/co-activators in gene transcription control.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Alcohol Oxidoreductases
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Animals
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Bacterial Proteins / metabolism
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Blotting, Western / methods
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CREB-Binding Protein
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Calcium / metabolism
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Carrier Proteins
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Cell Count / methods
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Cell Line / cytology*
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Cell Line / metabolism
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Cell Nucleus / genetics
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Cell Nucleus / metabolism*
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DNA-Binding Proteins
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Fluorescent Antibody Technique / methods
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Humans
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Luminescent Proteins / metabolism
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Mutagenesis / physiology
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Neoplasm Proteins / metabolism
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Phosphoproteins
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Promyelocytic Leukemia Protein
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RNA, Messenger / biosynthesis
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Rats
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Recombinant Proteins / metabolism
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Reverse Transcriptase Polymerase Chain Reaction / methods
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Trans-Activators / genetics
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Trans-Activators / metabolism*
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Trans-Activators / physiology*
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Transcription Factors / metabolism
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Transcription, Genetic / physiology
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Transcriptional Activation / physiology*
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Transfection / methods
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Tumor Suppressor Proteins
Substances
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Bacterial Proteins
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Carrier Proteins
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DNA-Binding Proteins
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Luminescent Proteins
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Neoplasm Proteins
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Nuclear Proteins
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Phosphoproteins
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Promyelocytic Leukemia Protein
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RNA, Messenger
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Recombinant Proteins
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Ss18l1 protein, rat
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Trans-Activators
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Transcription Factors
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Tumor Suppressor Proteins
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citrate-binding transport protein
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yellow fluorescent protein, Bacteria
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PML protein, human
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Alcohol Oxidoreductases
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C-terminal binding protein
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CREB-Binding Protein
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CREBBP protein, human
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Crebbp protein, rat
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Calcium