Objectives: 

Oestrogen receptor beta (b) expression in colon carcinoma is associated with patient survival, however the precise role of oestrogen and its receptors is controversial. MicroRNAs are regulated by oestrogen in other carcinomas. In this study we investigated whether tumour progression is modulated by the loss of ERb through ERa and/or G-protein coupled ER (GPER)-regulated microRNAs and their mRNA targets.

Materials: 

ER isoform abundance in commonly used colon carcinoma cell lines (T84, HT29, RKO and DLD-1) was analysed by RT-qPCR. 17b-oestradiol (E2)-responsive microRNAs, and their target mRNAs, in HT29 cells were investigated by microarray analysis of 750 microRNAs and 24,000 mRNAs, respectively. The microRNA target prediction tool Target Scan (www.targetscan.org) was used to predict microRNA targets of interest. Cell line gene expression was correlated with human colonic tumour specimens using the R2 microarray analysis platform (http://r2.amc.nl). Statistical significance was determined by Students t-test.

Results: 

Both ERa and ERb were absent in HT29 cells. GPER was however expressed in T84, HT29, RKO and DLD-1 cells, and down-regulation correlated with an aggressive tumour cell phenotype. GPER expression was also decreased in colon carcinoma patient tumour specimens. Eleven novel E2-sensitive microRNAs were significantly differentially expressed (>2-fold, p<0.05) and were predicted to target differentially expressed E2-responsive mRNAs (>2-fold, p<0.05), including known regulators of tumourigenesis. In particular, tyrosine kinase substrate 4 (SH3PXD2B), a regulator of colon carcinoma cell invasion, was significantly up-regulated (>2.4-fold, p=0.003) after E2 (10nM) treatment and showed significant enrichment for E2-repressed microRNA target sites (p=0.012).

Conclusions: 

We conclude that microRNA induction through GPER may contribute to tumour promotion following loss of ERb in colon carcinoma and may have potential as early diagnostic and therapeutic targets in colon carcinoma.