A SMYD3/ITGB6/TGFβ1 Positive Feedback Loop Promotes the Invasion and Adhesion of Ovarian Cancer Spheroids

Abstract

BackgroundImplantation metastasis is the main means of dissemination in ovarian cancer. Our previous studies showed that SET and MYND domain-containing protein 3 (SMYD3) expression was higher in ovarian cancer spheroids than in monolayers. SMYD3 enhancement of spheroid invasion and adhesion is mediated by the downstream effectors ITGB6 and ITGAM. However, the potential mechanisms underlying the SMYD3/integrin-mediated invasion and adhesion of spheroids still need to be explored.MethodsWestern blotting was used to examine the expression of SMYD3, ITGB6 and downstream molecules under different treatments. Immunofluorescence was used to detect the expression of F-actin, E-cadherin and N-cadherin. Anti-ITGB6 antibody-based inhibition and dual-luciferase reporter assays were used to confirm the binding between ITGB6 and latent TGFβ1. Transwell invasion, adherence and 3D tumor spheroid invasion assays were employed to test the effects of TGFβ1 on the invasion and adhesion of ovarian cancer spheroids. ELISA was performed to assess the release of latent TGFβ1 from ovarian cancer spheroids.ResultsSMYD3 and ITGB6 activated the TGFβ1/Smad3 pathway and then induced the upregulation of Snail, Vimentin and N-cadherin and the downregulation of E-cadherin in 3D-cultured ovarian cancer spheroids. In this process, latent TGFβ1 could bind to ITGB6 and become activated to stimulate the Smad3 pathway. Moreover, SMYD3 and ITGB6 could facilitate the release of latent TGFβ1 from 3D-cultured ovarian cancer spheroids. Interestingly, TGFβ1 could promote the expression of SMYD3 and ITGB6 via feedback. This positive feedback loop could further amplify the biological effect and promote the invasion and adhesion of ovarian cancer spheroids.ConclusionOur results demonstrated that the SMYD3/ITGB6/TGFβ1-Smad3 positive feedback loop could promote the invasion and adhesion of ovarian cancer spheroids by upregulating the expression of N-cadherin, Snail, and Vimentin and downregulating the expression of E-cadherin. Thus, our study unmasked the mechanism of SMYD3- and ITGB6-induced ovarian cancer metastasis and provides new ideas for targeted ovarian cancer treatment.

Overview of the signaling pathways involved in metastasis: An intriguing story-tale of the metastatic journey of ovarian cancer cells

“…SMYD3 and ITGB6 promoted the release and activation of latent TGFβ1 and increased the phosphorylation of SMAD3 in ovarian cancer spheroids. TGFβ1 promoted the expression of ITGB6 and SMYD3 (15).…”

“…SMYD3 (SET and MYND domain-containing protein 3) is a histone methyltransferase (15). SMYD3 has been shown to promote ovarian cancer.…”

SMYD3 promotes endometrial cancer through epigenetic regulation of LIG4/XRCC4/XLF complex in non-homologous end joining repair

“…of tumors, such as colorectal cancer, hepatocellular cancer, breast cancer and ovarian cancer (26)(27)(28), suggesting its important role in tumor initiation and progression. SMYD3 was also reported to methylate the histone H2A.Z.1 at lysine 101 at the promoter of cyclin A1, promoting cyclin A1's transcription in breast cancer (29).…”

Mechanisms of Cell Adhesion Molecules in Endocrine-Related Cancers: A Concise Outlook

“…Integrin αv activates latent TGF-β and drives EMT in PDAC cells ( 149 ). Similarly, integrin α6 induced EMT with the upregulation of N-cadherin and downregulation of CDH1 in ovarian cancer spheroids via the TGF-β1/Smad3 pathway ( 186 ). Interestingly, TGF-β1 promotes the expression of SMYD3 and ITGB6 as feedback mechanisms ( 186 ).…”

“…Similarly, integrin α6 induced EMT with the upregulation of N-cadherin and downregulation of CDH1 in ovarian cancer spheroids via the TGF-β1/Smad3 pathway ( 186 ). Interestingly, TGF-β1 promotes the expression of SMYD3 and ITGB6 as feedback mechanisms ( 186 ). Integrin α6 is enriched and enhances stem cell phenotypes in breast CSCs, especially in TNBC cells ( 187 , 188 ).…”