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. 2017 May 22;7:2229. doi: 10.1038/s41598-017-02494-6

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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HMGA2 directly targets Twist1 to promote VM in GC. (A) HMGA2 interacted with Twist1 in MGC803 and MKN74 cell line with HMGA2 overexpression. (B) ChIP was performed on MKN74 cells transfected with pc DNA-HMGA2. The precipitated chromatin was PCR-amplified with the use of specific primers in the Twist1 promoter (Supplementary Table S3) as indicated by black dots. Bar graphs show fold enrichment of HMGA2 binding of Twist1 promoter. Relative enrichment compared to irrelative antibody control is shown. The mean ± SD of three determinations is shown. (C) Luciferase activity assays in 293T cells showed the enhancement of Twist1 promoter activity by HMGA2 (*P < 0.05). (D) Western blot showed the protein expression of HMGA2, Twist1, VE-cadherin, vimentin, E-cadherin, N-cadherin when Twist1 knockdown in MKN74 cells with HMGA2 overexpression (*P < 0.05). 3D culture (E) and migration (F) showed Twist1-konckdown decreased VM formation and migration in MKN74 cells with HMGA2 overexpression. Scale bar represents 100 um, *P < 0.05. (G) IHC staining of Twist1 and VE-cadherin in GC samples (×200, bar 50 um); (H) Kaplan-Meier survival analysis of GC patients categorized by a combination of Twist1 and VE-cadherin (*P < 0.05). (I) A positive correlation between Twist1 and VE-cadherin using the Pearson correlation (r = 0.4826 *P < 0.001).