Sp1-mediated up-regulation of lnc00152 promotes invasion and metastasis of retinoblastoma cells via the miR-30d/SOX9/ZEB2 pathway

Cell Oncol (Dordr). 2021 Feb;44(1):61-76. doi: 10.1007/s13402-020-00522-8. Epub 2020 Jun 1.

Abstract

Purpose: Previously, we found that long non-coding RNA (lncRNA) MEG3 may act as a tumour suppressor in retinoblastoma. Overall, however, little is known about the role of lncRNAs in retinoblastoma. Here, we aimed to determine the expression and clinical significance of lnc00152 in retinoblastoma.

Methods: Lnc00152 and its downstream targets were selected using GEO datasets. The level of lnc00152 in primary patient samples was determined using RT-qPCR. Odds ratios of invasion and metastasis were calculated using logistic regression analysis. Recurrence-free survival was assessed using Cox regression analysis. Scratch wound healing, transwell and tumorigenesis assays were used to determine migration and invasion abilities of retinoblastoma cells in vitro and in vivo. Levels of EMT-related proteins were measured using Western blotting. Binding sites between lnc00152 and its targets were validated using dual-luciferase reporter and RNA pull-down assays. Lnc00152 activating transcription factors were determined using ChIP assays.

Results: We found that Lnc00152 was significantly up-regulated in retinoblastoma tumour tissues, and was a risk factor for tumour invasion, metastasis and recurrence. Lnc00152 overexpressing retinoblastoma cells exhibited a tendency to transform into mesenchymal cells, with significantly increased migration and invasion capacities, significantly decreased E-cadherin expression levels, and significantly increased N-cadherin, SOX9 and ZEB2 expression levels. In addition, we found that lnc00152, which was activated by Sp1, could inhibit miR-30d as an endogenous miRNA 'sponge', thereby regulating the expression of SOX9 and ZEB2.

Conclusions: Our data indicate that Lnc00152 may be associated with retinoblastoma invasion, metastasis and prognosis. In addition, we conclude that Lnc00152, which can be activated by Sp1, can induce EMT via the miR-30d/SOX9/ZEB2 pathway and, by doing so, promote the invasion and metastasis of retinoblastoma cells.

Keywords: EMT; Retinoblastoma; Sp1 protein; lnc00152; lncRNA; miR-30d.

MeSH terms

  • Animals
  • Base Sequence
  • Binding, Competitive
  • Cell Line, Tumor
  • Cell Movement / genetics
  • Cell Shape
  • Child, Preschool
  • Databases, Genetic
  • Disease-Free Survival
  • Epithelial-Mesenchymal Transition / genetics
  • Female
  • Gene Expression Profiling
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Logistic Models
  • Male
  • Mice
  • Mice, Inbred NOD
  • Mice, SCID
  • MicroRNAs / genetics
  • MicroRNAs / metabolism
  • Multivariate Analysis
  • Neoplasm Invasiveness
  • Neoplasm Metastasis
  • Neoplasm Proteins / metabolism
  • Prognosis
  • Proportional Hazards Models
  • RNA, Long Noncoding / genetics*
  • RNA, Long Noncoding / metabolism
  • Retinoblastoma / genetics
  • Retinoblastoma / pathology*
  • SOX9 Transcription Factor / metabolism*
  • Signal Transduction
  • Sp1 Transcription Factor / metabolism*
  • Up-Regulation / genetics*
  • Zinc Finger E-box Binding Homeobox 2 / metabolism*

Substances

  • MIRN30a microRNA, human
  • MicroRNAs
  • Neoplasm Proteins
  • RNA, Long Noncoding
  • SOX9 Transcription Factor
  • SOX9 protein, human
  • Sp1 Transcription Factor
  • ZEB2 protein, human
  • Zinc Finger E-box Binding Homeobox 2
  • long non-coding RNA Linc00152, human