TY - JOUR
T1 - A novel FOXO1-mediated dedifferentiation blocking role for DKK3 in adrenocortical carcinogenesis
AU - Cheng, Joyce Y.
AU - Brown, Taylor C.
AU - Murtha, Timothy D.
AU - Stenman, Adam
AU - Juhlin, C. Christofer
AU - Larsson, Catharina
AU - Healy, James M.
AU - Prasad, Manju L.
AU - Knoefel, Wolfram T.
AU - Krieg, Andreas
AU - Scholl, Ute I.
AU - Korah, Reju
AU - Carling, Tobias
N1 - Funding Information:
This work was supported by the Damon Runyon Cancer Research Foundation (T.C.), the Ohse Research Foundation (T.C.B.), and a Medical Student Research Fellowship from the NIH – NHLBI (J.C.). None of the funding bodies played a role in data collection, analysis, or interpretation of data, the writing of the manuscript, or the decision to submit the manuscript for publication.
Publisher Copyright:
© 2017 The Author(s).
PY - 2017/3/1
Y1 - 2017/3/1
N2 - Background: Dysregulated WNT signaling dominates adrenocortical malignancies. This study investigates whether silencing of the WNT negative regulator DKK3 (Dickkopf-related protein 3), an implicated adrenocortical differentiation marker and an established tumor suppressor in multiple cancers, allows dedifferentiation of the adrenal cortex. Methods: We analyzed the expression and regulation of DKK3 in human adrenocortical carcinoma (ACC) by qRT-PCR, immunofluorescence, promoter methylation assay, and copy number analysis. We also conducted functional studies on ACC cell lines, NCI-H295R and SW-13, using siRNAs and enforced DKK3 expression to test DKK3's role in blocking dedifferentiation of adrenal cortex. Results: While robust expression was observed in normal adrenal cortex, DKK3 was down-regulated in the majority (>75%) of adrenocortical carcinomas (ACC) tested. Both genetic (gene copy loss) and epigenetic (promoter methylation) events were found to play significant roles in DKK3 down-regulation in ACCs. While NCI-H295R cells harboring β-catenin activating mutations failed to respond to DKK3 silencing, SW-13 cells showed increased motility and reduced clonal growth. Conversely, exogenously added DKK3 also increased motility of SW-13 cells without influencing their growth. Enforced over-expression of DKK3 in SW-13 cells resulted in slower cell growth by an extension of G1 phase, promoted survival of microcolonies, and resulted in significant impairment of migratory and invasive behaviors, largely attributable to modified cell adhesions and adhesion kinetics. DKK3-over-expressing cells also showed increased expression of Forkhead Box Protein O1 (FOXO1) transcription factor, RNAi silencing of which partially restored the migratory proficiency of cells without interfering with their viability. Conclusions: DKK3 suppression observed in ACCs and the effects of manipulation of DKK3 expression in ACC cell lines suggest a FOXO1-mediated differentiation-promoting role for DKK3 in the adrenal cortex, silencing of which may allow adrenocortical dedifferentiation and malignancy.
AB - Background: Dysregulated WNT signaling dominates adrenocortical malignancies. This study investigates whether silencing of the WNT negative regulator DKK3 (Dickkopf-related protein 3), an implicated adrenocortical differentiation marker and an established tumor suppressor in multiple cancers, allows dedifferentiation of the adrenal cortex. Methods: We analyzed the expression and regulation of DKK3 in human adrenocortical carcinoma (ACC) by qRT-PCR, immunofluorescence, promoter methylation assay, and copy number analysis. We also conducted functional studies on ACC cell lines, NCI-H295R and SW-13, using siRNAs and enforced DKK3 expression to test DKK3's role in blocking dedifferentiation of adrenal cortex. Results: While robust expression was observed in normal adrenal cortex, DKK3 was down-regulated in the majority (>75%) of adrenocortical carcinomas (ACC) tested. Both genetic (gene copy loss) and epigenetic (promoter methylation) events were found to play significant roles in DKK3 down-regulation in ACCs. While NCI-H295R cells harboring β-catenin activating mutations failed to respond to DKK3 silencing, SW-13 cells showed increased motility and reduced clonal growth. Conversely, exogenously added DKK3 also increased motility of SW-13 cells without influencing their growth. Enforced over-expression of DKK3 in SW-13 cells resulted in slower cell growth by an extension of G1 phase, promoted survival of microcolonies, and resulted in significant impairment of migratory and invasive behaviors, largely attributable to modified cell adhesions and adhesion kinetics. DKK3-over-expressing cells also showed increased expression of Forkhead Box Protein O1 (FOXO1) transcription factor, RNAi silencing of which partially restored the migratory proficiency of cells without interfering with their viability. Conclusions: DKK3 suppression observed in ACCs and the effects of manipulation of DKK3 expression in ACC cell lines suggest a FOXO1-mediated differentiation-promoting role for DKK3 in the adrenal cortex, silencing of which may allow adrenocortical dedifferentiation and malignancy.
KW - Adrenocortical carcinogenesis
KW - DKK3
KW - FOXO1
UR - http://www.scopus.com/inward/record.url?scp=85014305210&partnerID=8YFLogxK
U2 - 10.1186/s12885-017-3152-5
DO - 10.1186/s12885-017-3152-5
M3 - Article
C2 - 28249601
AN - SCOPUS:85014305210
SN - 1471-2407
VL - 17
JO - BMC Cancer
JF - BMC Cancer
IS - 1
M1 - 164
ER -