Document Detail


C-terminal cleavage of DeltaNp63alpha is associated with TSA-induced apoptosis in immortalized corneal epithelial cells.
MedLine Citation:
PMID:  20375332     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
PURPOSE: In the central human corneal epithelium, loss of DeltaNp63 occurs in all surface epithelial cells preparing to undergo desquamation, suggesting a potential role for DeltaNp63 isoforms in mediating surface cell apoptotic shedding. In this study, the authors investigated a role for DeltaNp63 isoforms in caspase-mediated apoptosis in a telomerase-immortalized corneal epithelial cell line.
METHODS: For in vitro studies, hTCEpi cells were cultured in KGM-2 serum-free culture media containing 0.15 mM calcium. To assess dynamic protein interactions among individual DeltaNp63 isoforms, DeltaNp63-EGFP expression plasmids were transiently expressed in hTCEpi cells and evaluated by FRAP. Trichostatin-A (TSA; 3.31 muM) was used to induce cell death as measured by caspase activity. Cleavage and loss of endogenous DeltaNp63alpha, DeltaNp63-EGFP expression plasmids, and p53 were assessed after treatment with TSA and siRNA.
RESULTS: Transient expression of DeltaNp63-EGFP alpha and beta isoforms resulted in the formation of a smaller isoform similar in size to DeltaNp63gamma-EGFP. FRAP demonstrated that DeltaNp63alpha-EGFP has greater immobile fraction than beta or gamma. TSA induced caspase-mediated apoptotic pathways; caspase induction was accompanied by a decrease in endogenous DeltaNp63alpha and p53. TSA upregulated DeltaNp63-EGFP plasmid expression; this was accompanied by a selective increase in cleavage of DeltaNp63alpha-EGFP. siRNA knockdown of DeltaNp63alpha correlated with a reduction in p53 independently of TSA.
CONCLUSIONS: DeltaNp63alpha is the dominant active isoform in corneal epithelial cell nuclei. Loss of DeltaNp63alpha occurs during apoptotic signaling by cleavage at the C terminus. The corresponding loss of p53 suggests that a significant relationship appears to exist between these two regulatory proteins.
Authors:
Danielle M Robertson; Su-Inn Ho; H Dwight Cavanagh
Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't     Date:  2010-04-07
Journal Detail:
Title:  Investigative ophthalmology & visual science     Volume:  51     ISSN:  1552-5783     ISO Abbreviation:  Invest. Ophthalmol. Vis. Sci.     Publication Date:  2010 Aug 
Date Detail:
Created Date:  2010-07-23     Completed Date:  2010-08-20     Revised Date:  2014-09-21    
Medline Journal Info:
Nlm Unique ID:  7703701     Medline TA:  Invest Ophthalmol Vis Sci     Country:  United States    
Other Details:
Languages:  eng     Pagination:  3977-85     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Apoptosis / drug effects*
Blotting, Western
Caspase 3 / metabolism
Cell Culture Techniques
Epithelium, Corneal / metabolism,  pathology*
Fluorescence Recovery After Photobleaching
Gene Silencing
Green Fluorescent Proteins / genetics,  metabolism
Humans
Hydroxamic Acids / pharmacology*
Plasmids
Protein Isoforms
Protein Synthesis Inhibitors / pharmacology*
RNA, Small Interfering / pharmacology
Recombinant Fusion Proteins
Trans-Activators / physiology*
Transcription Factors
Transfection
Tumor Suppressor Protein p53 / metabolism
Tumor Suppressor Proteins / physiology*
Up-Regulation
Grant Support
ID/Acronym/Agency:
EY016664/EY/NEI NIH HHS; R01 EY018219/EY/NEI NIH HHS; R24 EY016664/EY/NEI NIH HHS; R24 EY016664-05/EY/NEI NIH HHS
Chemical
Reg. No./Substance:
0/Hydroxamic Acids; 0/Protein Isoforms; 0/Protein Synthesis Inhibitors; 0/RNA, Small Interfering; 0/Recombinant Fusion Proteins; 0/TP63 protein, human; 0/Trans-Activators; 0/Transcription Factors; 0/Tumor Suppressor Protein p53; 0/Tumor Suppressor Proteins; 0/enhanced green fluorescent protein; 147336-22-9/Green Fluorescent Proteins; 3X2S926L3Z/trichostatin A; EC 3.4.22.-/Caspase 3
Comments/Corrections

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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