Expression of PTRF in PC-3 Cells modulates cholesterol dynamics and the actin cytoskeleton impacting secretion pathways.
Expression of caveolin-1 is up-regulated in prostate cancer metastasis and is associated with aggressive recurrence of the disease. Intriguingly, caveolin-1 is also secreted from prostate cancer cell lines and has been identified in secreted prostasomes. Caveolin-1 is the major structural component of the plasma membrane invaginations called caveolae. Co-expression of the coat protein Polymerase I and transcript release factor (PTRF) is required for caveolae formation. We recently found that expression of caveolin-1 in the aggressive prostate cancer cell line PC-3 is not accompanied by PTRF, leading to noncaveolar caveolin-1 lipid rafts. Moreover, ectopic expression of PTRF in PC-3 cells sequesters caveolin-1 into caveolae. Here we quantitatively analyzed the effect of PTRF expression on the PC-3 proteome using stable isotope labeling by amino acids in culture and subcellular proteomics. We show that PTRF reduced the secretion of a subset of proteins including secreted proteases, cytokines, and growth regulatory proteins, partly via a reduction in prostasome secretion. To determine the cellular mechanism accounting for the observed reduction in secreted proteins we analyzed total membrane and the detergent-resistant membrane fractions. Our data show that PTRF expression selectively impaired the recruitment of actin cytoskeletal proteins to the detergent-resistant membrane, which correlated with altered cholesterol distribution in PC-3 cells expressing PTRF. Consistent with this, modulating cellular cholesterol altered the actin cytoskeleton and protein secretion in PC-3 cells. Intriguingly, several proteins that function in ER to Golgi trafficking were reduced by PTRF expression. Taken together, these results suggest that the noncaveolar caveolin-1 found in prostate cancer cells generates a lipid raft microenvironment that accentuates secretion pathways, possibly at the step of ER sorting/exit. Importantly, these effects could be modulated by PTRF expression.
The University of Queensland Diamantina Institute, The University of Queensland, Brisbane, Queensland 4102, Australia.
Kerry L
KL
Inder
Yu Zi
YZ
Zheng
Melissa J
MJ
Davis
Hyeongsun
H
Moon
Dorothy
D
Loo
Hien
H
Nguyen
Judith A
JA
Clements
Robert G
RG
Parton
Leonard J
LJ
Foster
Michelle M
MM
Hill
D008841:Actin Cytoskeleton / Q000378:metabolism*; D015153:Blotting, Western; D021941:Caveolae / Q000378:metabolism; D051242:Caveolin 1 / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D002853:Chromatography, Liquid; D004705:Endocytosis; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D008297:Male; D021962:Membrane Microdomains / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism*; D021381:Protein Transport; D020543:Proteome / Q000032:analysis*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D055571:Secretory Pathway; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014407:Tumor Cells, Cultured
C497835:CAVIN1 protein, human; D051242:Caveolin 1; D008565:Membrane Proteins; D020543:Proteome; D016601:RNA-Binding Proteins; D002784:Cholesterol