Prohibitin regulates TGF-beta induced apoptosis as a downstream effector of Smad-dependent and -independent signaling.
BACKGROUND
Prohibitin (PHB), a protein located on the inner mitochondrial membrane and nuclei, is an intracellular effector of transforming growth factor-beta (TGF-beta) signaling in prostate cancer cells. This study investigated the involvement of PHB in the apoptosis and survival outcomes of human prostate cancer cell to TGF-beta. shRNA PHB loss of function in prostate cancer cells led to enhanced apoptotic response to TGF-beta via Smad-dependent mechanism.


METHODS
TGF-beta activation of Raf-Erk intracellular signaling, led to PHB phosphorylation, decreased inner mitochondrial permeability, and increased cell survival. Calcein-based immunofluorescence studies revealed the functional involvement of PHB in maintaining inner mitochondrial membrane permeability as an integral component of TGF-beta induced apoptosis in prostate cancer cells.


RESULTS
These finding indicates that induction of TGF-beta apoptosis is mediated by Smad-dependent and Smad-independent signaling (MAPK) converging at PHB as a downstream effector regulating inner mitochondrial permeability. Putative PHB associated proteins were identified by subjecting TGF-beta treated cells to immunoprecipitation with anti-PHB, and mass spectrometry. A screen for the kinase specific phosphorylation sites of PHB revealed three protein kinase (PKC) binding sites.


CONCLUSIONS
Our results demonstrate that TGF-beta led to upregulation of the PKC inhibitor 14-3-3 protein and promoted its association with PHB, while PHB association with PKC-delta, was inhibited by the MEK1 inhibitor, documenting a critical interdependence between the MEK-ERK signaling and prohibitin phosphorylation. These findings suggest a dual role for PHB as a downstream determinant of the cellular response to TGF-beta via Smad-dependent pathway (apoptosis) and MAPK intracellular signaling (survival).
Zhu
Brian
B
Department of Surgery, Division of Urology, University of Kentucky College of Medicine, Lexington, Kentucky, USA.
Zhai
Jianjun
J
Zhu
Haining
H
Kyprianou
Natasha
N
D000595:Amino Acid Sequence; D017209:Apoptosis / Q000502:physiology*; D045744:Cell Line, Tumor; D015536:Down-Regulation / Q000502:physiology*; D006801:Humans; D020935:MAP Kinase Signaling System / Q000502:physiology*; D008297:Male; D051336:Mitochondrial Membranes / Q000502:physiology; D008969:Molecular Sequence Data; D010539:Permeability; D000091402:Prohibitins; D012097:Repressor Proteins / Q000502:physiology*; D051785:Smad Proteins / Q000502:physiology*; D016212:Transforming Growth Factor beta / Q000378:metabolism / Q000502:physiology*
C000717328:PHB protein, human; D000091402:Prohibitins; D012097:Repressor Proteins; D051785:Smad Proteins; D016212:Transforming Growth Factor beta