FIG. 3.

CPEB family proteins are nucleus-cytoplasm shuttling proteins. (A) DIV16 hippocampal neurons were treated with 0.1% ethanol alone or with 10 nM LMB for 40 min before fixation and then immunostained for endogenous CPEB4. (B) DIV16 neurons infected with lentivirus expressing HA-CPEB1, myc-CPEB3, or myc-CPEB4 for 2 days were treated with 10 nM LMB for 1 h and then immunostained with HA or myc antibodies. Other infected cells were stimulated with NMDA for 1 h prior to immunostaining. (C) Quantification of CPEB1, CPEB3, or CPEB4 in the nucleus or cytoplasm following the treatments described for panel B. The asterisks refer to statistically significant differences (P < 0.01, Student's t test) between the results for the indicated samples. C4RBD, CPEB4 RNA binding domain. (D) HeLa cells were treated with digitonin, and the permeabilized cells were incubated with an ATP-regenerating system, purified recombinant GST-CPEB4 RBD fusion protein, with or without HeLa cell cytosol. After 40 min, the cells were fixed and stained with GST antibody. Size bar = 10 μm.