title: C-terminal binding proteins are essential pro-survival factors that undergo caspase-dependent downregulation during neuronal apoptosis.
abstract: C-terminal binding proteins (CtBPs) are transcriptional co-repressors that are subject to proteasome-dependent downregulation during apoptosis. Alternative mechanisms that regulate CtBP expression are currently under investigation and the role of CtBPs in neuronal survival is largely unexplored. Here, we show that CtBPs are downregulated in cerebellar granule neurons (CGNs) induced to undergo apoptosis by a variety of stressors. Moreover, antisense-mediated downregulation of CtBP1 is sufficient to cause CGN apoptosis. Similarly, the CtBP inhibitor, 4-methylthio-2-oxobutyric acid, induces expression of the CtBP target Noxa and causes actinomycin-sensitive CGN apoptosis. Unexpectedly, we found that the mechanism of CtBP downregulation in CGNs undergoing apoptosis varies in a stimulus-specific manner involving either the proteasome or caspases. In the case of CGNs deprived of depolarizing potassium (5K apoptotic condition), caspases appear to play a dominant role in CtBP downregulation. However, incubation in 5K does not enhance the kinetics of CtBP1 degradation and recombinant CtBP1 is not cleaved in vitro by caspase-3. In addition, 5K has no significant effect on CtBP transcript expression. Finally, mouse embryonic stem cells display caspase-dependent downregulation of CtBP1 following exposure to staurosporine, an effect that is not observed in DGCR8 knockout cells which are deficient in miRNA processing. These data identify caspase-dependent downregulation of CtBPs as an alternative mechanism to the proteasome for regulation of these transcriptional co-repressors in neurons undergoing apoptosis. Moreover, caspases appear to regulate CtBP expression indirectly, at a post-transcriptional level, and via a mechanism that is dependent upon miRNA processing. We conclude that CtBPs are essential pro-survival proteins in neurons and their downregulation contributes significantly to neuronal apoptosis via the de-repression of pro-apoptotic genes.
authors.affiliation: Research Service, Veterans Affairs Medical Center, Denver, Colorado, 80220, USA.|Department of Biological Sciences and Eleanor Roosevelt Institute, University of Denver, Denver, Colorado, 80208, USA.
authors.forename: Trisha R
authors.initials: TR
authors.lastname: Stankiewicz
authors.affiliation: Research Service, Veterans Affairs Medical Center, Denver, Colorado, 80220, USA.
authors.forename: Emily K
authors.initials: EK
authors.lastname: Schroeder
authors.affiliation: Department of Biological Sciences and Eleanor Roosevelt Institute, University of Denver, Denver, Colorado, 80208, USA.
authors.forename: Natalie A
authors.initials: NA
authors.lastname: Kelsey
authors.affiliation: Research Service, Veterans Affairs Medical Center, Denver, Colorado, 80220, USA.
authors.forename: Ron J
authors.initials: RJ
authors.lastname: Bouchard
authors.affiliation: Research Service, Veterans Affairs Medical Center, Denver, Colorado, 80220, USA.|Department of Biological Sciences and Eleanor Roosevelt Institute, University of Denver, Denver, Colorado, 80208, USA.|Division of Clinical Pharmacology and Toxicology, Department of Medicine and Neuroscience Program, University of Colorado School of Medicine, Aurora, Colorado, 80045, USA.
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Linseman
mesh_terms: D000818:Animals; D017209:Apoptosis; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D053148:Caspase 3 / Q000378:metabolism; D015536:Down-Regulation; D005260:Female; D008297:Male; D008715:Methionine / Q000031:analogs & derivatives / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D011188:Potassium / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D019311:Staurosporine / Q000494:pharmacology; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; C119945:Ctbp1 protein, rat; C539472:Pmaip1 protein, rat; D019253:Proto-Oncogene Proteins c-bcl-2; D014157:Transcription Factors; C014632:2-keto-4-methylthiobutyric acid; D008715:Methionine; D053148:Caspase 3; D019311:Staurosporine; D011188:Potassium
keywords: 4,6-diamidino-2-phenylidole; 4-methylthio-2-oxobutyric; 6-OHDA; 6-hydroxydopamine; AD; Anti; B-cell lymphoma-3; BKLF; Bcl-3; C-terminal binding protein; C-terminal binding proteins (CtBPs); CCD; CGNs; Caspase-3; Con; CtBP; DAPI; DGCR8; DiGeorge critical region 8; EP; G protein-coupled receptor kinase-interacting protein-z-short; GITz-short; Inv; KO; LTox; MEFs; MPP(+); MTOB; Neuronal apoptosis; PARP; PBS; PCR; RNA silencing suppressor; RSS; SNP; STS; Tox B; Toxin B; WT; actinomycin D; basic Krüppel-like factor; cerebellar granule neurons; charge-coupled device; control; dCtBP; drosophila CtBP; endoporter; inverse; knockout; lethal toxin; mESCs; methyl-4-phenylpyridinium; miRNA; micro RNA; morpholino-antisense oligonucleotides; mouse embryonic stem cells; murine embryonic fibroblasts; phosphate buffer saline; poly-(ADP-ribose) polymerase; polymerase chain reaction; rCasp3; recombinant caspase-3; sodium nitroprusside; staurosporine; wild type