mRNA Instability in the Nucleus Due to a Novel Open Reading Frame Element Is a Major Determinant of the Narrow Tissue Specificity of Folate Receptor α

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Abstract
The folate receptor type α (FR-α) is a promising tumor marker and target. Here, we investigate the mechanistic basis for the tumor specificity and vast overexpression of FR-α. Among representative FR-α-positive (HeLa and JAR) and FR-α-negative (MG63, Caki1, and HT3) cell lines, the transcription rates of the endogenous FR-α gene, as well as the FR-α promoter activity, were relatively weak and comparable, but the FR-α transcript was abundant only in total RNA and nuclear RNA from the FR-α-positive cells. Rous sarcoma virus (RSV) promoter-driven expression of the FR-α gene was 7 to 30 times greater in the FR-α-positive than in FR-α-negative cells, both at the protein and mRNA levels, independently of intron sequences. Through the use of chimeric FR-α/FR-β cDNAs, the above pattern of FR-α expression was attributed to a 60-bp sequence in the FR-α open reading frame. This sequence element, when placed in the 5′ untranslated region of RSV promoter-luciferase, decreased the reporter expression approximately 7- to 20-fold in FR-α-negative cells (MG63, Caki1, HT3, BG1, and MCF7) relative to FR-α-positive cells (HeLa, JAR, and JEG3). Substitution of this FR-α element in FR-β increased the in vivo degradation rate of the transcript in the nuclei of MG63 cells but not in the nuclei of HeLa cells or in the cytosol of MG63 or HeLa cells. The results reveal an efficient mechanism by which a novel sequence element causes differential transcript degradation in the nucleus to ensure narrow tissue specificity for a gene (e.g., that for FR-α) whose transcription is weak and relatively nonselective. FR-α exhibited constitutive mRNA and protein synthesis during the cell cycle and a slow protein turnover, presumably ensuring a high steady-state level of the receptor in cells that could override the nuclear mRNA instability determinant.

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