New Phosphospecific Antibody Reveals Isoform-Specific Phosphorylation of CPEB3 Protein.

TitleNew Phosphospecific Antibody Reveals Isoform-Specific Phosphorylation of CPEB3 Protein.
Publication TypeJournal Article
Year of Publication2016
AuthorsKaczmarczyk L, Labrie-Dion É, Sehgal K, Sylvester M, Skubal M, Josten M, Steinhäuser C, De Koninck P, Theis M
JournalPLoS One
Date Published2016

Cytoplasmic Polyadenylation Element Binding proteins (CPEBs) are a family of polyadenylation factors interacting with 3'UTRs of mRNA and thereby regulating gene expression. Various functions of CPEBs in development, synaptic plasticity, and cellular senescence have been reported. Four CPEB family members of partially overlapping functions have been described to date, each containing a distinct alternatively spliced region. This region is highly conserved between CPEBs-2-4 and contains a putative phosphorylation consensus, overlapping with the exon seven of CPEB3. We previously found CPEBs-2-4 splice isoforms containing exon seven to be predominantly present in neurons, and the isoform expression pattern to be cell type-specific. Here, focusing on the alternatively spliced region of CPEB3, we determined that putative neuronal isoforms of CPEB3 are phosphorylated. Using a new phosphospecific antibody directed to the phosphorylation consensus we found Protein Kinase A and Calcium/Calmodulin-dependent Protein Kinase II to robustly phosphorylate CPEB3 in vitro and in primary hippocampal neurons. Interestingly, status epilepticus induced by systemic kainate injection in mice led to specific upregulation of the CPEB3 isoforms containing exon seven. Extensive analysis of CPEB3 phosphorylation in vitro revealed two other phosphorylation sites. In addition, we found plethora of potential kinases that might be targeting the alternatively spliced kinase consensus site of CPEB3. As this site is highly conserved between the CPEB family members, we suggest the existence of a splicing-based regulatory mechanism of CPEB function, and describe a robust phosphospecific antibody to study it in future.

Alternate JournalPLoS ONE
PubMed ID26915047
PubMed Central IDPMC4767366


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Canadian Institutes of Health Research - Instituts de recherche en santé du Canada Fonds de recherche du Québec – Nature et technologies (FRQNT)Fonds de la recherche en santé du Québec   Natural Sciences and Engineering Research Council of Canada (NSERC) - Conseil de recherche en sciences naturelles et en génie du Canada (CRSNG)innovation.caHuman Frontier Science ProgramCanada First Research Excellence FundSentinelle Nord