MicroRNA-34 mediates AR-dependent p53-induced apoptosis in prostate cancer.
We investigated whether knocking down AR expression effects apoptosis after treatment with different apoptosis-inducing agents. We found that siRNA AR (si-AR) significantly decreased apoptosis induced by topoisomerase inhibitors doxorubicin (DOX) and camptothecin (Campt). It is known that DNA double-strand break inducing agents leads to activation (phosphorylation) of p53 that in turn regulates the expression of a variety of apoptosis-related genes including microRNA(miR)-34a and 34b/c. We found that DOX induced five phosphorylation sites of p53 (Ser15, 20, 37, 46 and 392); all of these sites were inhibited by si-AR. Subsequently we identified three kinases, SPAK, MDC1 and CaMKII that are under AR control and two of them, MDC1 and CaMKII, apparently participate in p53 upstream events that resulted in p53 inhibition. Using qPCR we showed that the level of miR-34a increased by 3-fold after DOX, but no increase was found with si-AR. MiR-34c expression increased 27 fold after DOX and only by 2.7 times with si-AR. It appears that AR-dependent inhibition of p53 resulted in suppression of miR-34a and -34c expression. Importantly, DOX did not induce miR-34 in LNCaP grown in an androgen free medium or in AR-negative prostate cancer cell lines, DU145 and PC3. To directly investigate the role of miR-34 in DOX-mediated apoptosis, we transfected cells with anti-miR-34 oligonucleotides or with miR-34. We found that inhibition of individual miR-34, either 34a or 34c, or forced overexpression of miR-34a or miR-34c did not modulate DOX-mediated apoptosis. Only simultaneous inhibition or forced overexpression of both miR-34 resulted in modulation of DOX-mediated apoptosis. Taken together, our data indicate that cooperation between miR-34a and 34c plays an important role in AR-dependent p53-mediated apoptosis in prostate cancer.
Rokhlin
Oskar W
OW
Department of Pathology, The University of Iowa, Iowa City, Iowa 52242, USA. oskar-rokhlin@uiowa.edu
Scheinker
Vladimir S
VS
Taghiyev
Agshin F
AF
Bumcrot
David
D
Glover
Rebecca A
RA
Cohen
Michael B
MB
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000494:pharmacology; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D011944:Receptors, Androgen / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology
D000903:Antibiotics, Antineoplastic; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D034741:RNA, Small Interfering; D011944:Receptors, Androgen; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin