Estrogen Down-regulation of the Scx Gene Is Mediated by the Opposing Strand-overlapping Gene Bop1

Yukitomo Arao,Karen Carpenter,Sylvia Hewitt,Kenneth S Korach

doi:10.1074/jbc.m109.036681

Abstract

Recent genome-wide transcriptome studies suggest the presence of numerous bidirectional overlapping coding gene pairs in mammalian genomes. Various antisense RNAs are reported as non-coding RNAs that regulate the expression of sense RNA. However, it is still unclear whether the expression of bidirectional overlapping coding genes are regulated by the opposite strand gene transcript acting as a non-coding RNA. Bop1 and Scx are a pair of bidirectional overlapping coding genes related to cellular proliferation and differentiation, respectively. Scx gene is localized in the intron 3 region of the Bop1 gene. The expression of these genes is reciprocally regulated by estrogen (E2) in the mouse uterus. In situ hybridization indicated that both genes are expressed in the uterine endometrial epithelial cells and that the antisense RNA of Scx (Bop1 intronic RNA) accumulates as a stable RNA in these cells. The existence of Bop1 intronic RNA was confirmed by reverse transcription-PCR and was increased after E2 treatment, coinciding with a decrease in Scx mRNA. Murine myoblasts expressing doxycycline-inducible endogenous Bop1 gene showed an increase in Bop1 intronic RNA and a simultaneous decrease in Scx mRNA. Murine fibroblasts expressing Scx mRNA from an exogenous Scx mini-gene indicated that the accumulation of Bop1 intronic RNA impairs the Scx gene expression in a trans-acting manner, which resulted in the reduction of the Scx mRNA level. This study demonstrates a novel example of hormone-stimulated intronic non-coding RNA down-regulating the expression of an opposing strand-overlapping coding gene.

Highlights

Genome-wide transcriptome studies suggest that about 15–25% of mammalian genes are arranged such that they overlap one another on opposite DNA strands, giving rise to pairs of sense and antisense RNAs [1,2,3,4,5,6]
Murine fibroblasts expressing Scx mRNA from an exogenous Scx mini-gene indicated that the accumulation of Bop1 intronic RNA impairs the Scx gene expression in a trans-acting manner, which resulted in the reduction of the Scx mRNA level
Tation; category 3, exonic regions overlap in divergent orientation; category 4, one transcript unit is completely overlapped within an intron of the transcription unit on the opposite strand; category 5, transcription units overlap in convergent orientation, but exonic regions do not overlap; category 6, transcription units overlap in divergent orientation, but exonic regions do not overlap

Summary

The abbreviations used are

E2, estrogen; ER, estrogen receptor; ICI, ICI182,780; OVX, ovariectomized; 18 S, 18 S ribosomal RNA; ISH, in situ hybridization; Tet, tetracycline; DMEM, Dulbecco’s modified Eagle’s medium; FBS, fetal bovine serum; CMV, cytomegalovirus; RT-PCR, reverse transcription-PCR; ϩRT, with reverse transcriptase; ϪRT, without reverse transcriptase. We reported that E2 induces the expression of cell proliferation-related genes similar to different growth factors in the ovariectomized (OVX) mouse uterus [29]. We demonstrate a mechanism of reciprocal regulation of Bop and Scx gene expression

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION