The oncogenic effect of sulfatase 2 in human hepatocellular carcinoma is mediated in part by glypican 3-dependent Wnt activation.
Heparan sulfate proteoglycans (HSPGs) act as coreceptors or storage sites for growth factors and cytokines such as fibroblast growth factor and Wnts. Glypican 3 (GPC3) is the most highly expressed HSPG in hepatocellular carcinoma (HCC). Sulfatase 2 (SULF2), an enzyme with 6-O-desulfatase activity on HSPGs, is up-regulated in 60% of primary HCCs and is associated with a worse prognosis. We have previously shown that the oncogenic effect of SULF2 in HCC may be mediated in part through up-regulation of GPC3. Here we demonstrate that GPC3 stimulates the Wnt/β-catenin pathway and mediates the oncogenic function of SULF2 in HCC. Wnt signaling in vitro and in vivo was assessed in SULF2-negative Hep3B HCC cells transfected with SULF2 and in SULF2-expressing Huh7 cells transfected with short hairpin RNA targeting SULF2. The interaction between GPC3, SULF2, and Wnt3a was assessed by coimmunoprecipitation and flow cytometry. β-catenin-dependent transcriptional activity was assessed with the TOPFLASH (T cell factor reporter plasmid) luciferase assay. In HCC cells, SULF2 increased cell surface GPC3 and Wnt3a expression, stabilized β-catenin, and activated T cell factor transcription factor activity and expression of the Wnt/β-catenin target gene cyclin D1. Opposite effects were observed in SULF2-knockdown models. In vivo, nude mouse xenografts established from SULF2-transfected Hep3B cells showed enhanced GPC3, Wnt3a, and β-catenin levels.


CONCLUSIONS
Together, these findings identify a novel mechanism mediating the oncogenic function of SULF2 in HCC that includes GPC3-mediated activation of Wnt signaling via the Wnt3a/glycogen synthase kinase 3 beta axis.
Lai
Jin-Ping
JP
Miles and Shirley Fiterman Center for Digestive Diseases, Mayo Clinic, Rochester, MN, USA.
Oseini
Abdul M
AM
Moser
Catherine D
CD
Yu
Chunrong
C
Elsawa
Sherine F
SF
Hu
Chunling
C
Nakamura
Ikuo
I
Han
Tao
T
Aderca
Ileana
I
Isomoto
Hajime
H
Garrity-Park
Megan M
MM
Shire
Abdirashid M
AM
Li
Jia
J
Sanderson
Schuyler O
SO
Adjei
Alex A
AA
Fernandez-Zapico
Martin E
ME
Roberts
Lewis R
LR
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000097:blood / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D053673:Glypicans / Q000097:blood / Q000235:genetics; D006801:Humans; D019394:Ki-67 Antigen / Q000235:genetics; D008113:Liver Neoplasms / Q000097:blood / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D010957:Plasmids / Q000235:genetics; D013429:Sulfatases; D015238:Sulfotransferases / Q000097:blood*; D014162:Transfection; D051153:Wnt Proteins / Q000235:genetics; D060508:Wnt3 Protein; D060509:Wnt3A Protein
C105937:GPC3 protein, human; D053673:Glypicans; D019394:Ki-67 Antigen; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; C555511:Wnt3a protein, mouse; D008156:Luciferases; D015238:Sulfotransferases; C470792:SULF2 protein, human; D013429:Sulfatases; D053148:Caspase 3