Colorectal carcinoma (CRC) is the third most common malignancy worldwide, and displays a lower incidence in women than in men (1). The protective role of oestrogen in CRC is associated with oestrogen receptor (ER)β expression and the loss of ERβ correlates with worse prognosis (2). It is not known whether oestrogen can promote tumour progression following the loss of ERβ. microRNAs (miRNAs) have the ability to repress gene expression and are believed to play an important role in development, differentiation, proliferation, survival, and oncogenesis. Pre-miRNA precursor transcript and mature miRNA can be modulated within minutes by transcription factors and represent a novel class of molecules rapidly activated by steroid hormones. miRNA expression is modulated by oestrogen, therefore, ERα and/or G-protein coupled ER (GPER)-regulated miRNAs may be detrimental in CRC. We have investigated the ER isoform abundance and 17β-oestradiol (E2)-responsive miRNAs, and their target mRNAs, in colon carcinoma cells. Real-time quantitative PCR analysis revealed both ERα and ERβ mRNA were absent in HT29 cells (n=3). GPER mRNA was, however, abundantly expressed in T84, HT29, RKO and DLD-1 colon carcinoma cells with a decrease in GPER expression correlating with a more aggressive cell phenotype (n=3). GPER was down-regulated in colon carcinoma tumour specimens compared to normal colonic tissue (R2 analysis). Microarray analysis of 750 miRNAs and 24,000 mRNAs revealed novel E2-modulated miRNA and mRNA transcripts in HT29 cells. Eleven E2-sensitive miRNAs were significantly differentially expressed (>2-fold, n=3, p<0.05). By in silico analysis, E2-modulated miRNAs were predicted to target significantly differentially expressed mRNAs (>2-fold, n=3, p<0.05). In particular, tyrosine kinase substrate 4 (SH3PXD2B), a recently documented regulator of colon carcinoma cell invasion (3), was significantly up-regulated (>2.4-fold, n=3, p=0.003) after E2 (10nM) treatment and showed significant enrichment for E2-repressed miRNA target sites (p=0.012). This study shows, for the first time, that GPER mRNA is expressed in colon carcinoma cells and, by meta-analysis, correlates with colon carcinoma tumour progression. E2 modulates miRNA expression in colon carcinoma cells. Novel E2-responsive mRNAs, including known regulators of tumourigenesis, were also identified and are predicted targets for E2-modulated miRNAs. We hypothesise that miRNA induction through GPER may contribute to tumour promotion following loss of ERβ expression in colon carcinoma. This work will direct future research of oestrogen-sensitive miRNAs to establish their potential value as early diagnostic and therapeutic targets in colon carcinoma.