Abstract
The interaction of an 18-base cis-element in the 5'-untranslated region of human folate receptor (FR)-alpha mRNA with a cytosolic trans-factor protein is critical for the translation of FR (Sun, X.-L., and Antony, A. C. (1996) J. Biol. Chem. 271, 25539-25547). This trans-factor was isolated to apparent homogeneity as a 43- and 38-kDa doublet from human placenta using poly(U)-Sepharose, followed by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electro-elution as major purification steps. Amino acid microsequencing of two cyanogen bromide-generated peptide fragments of the 43-kDa trans-factor revealed complete identity with 43-kDa heterogeneous nuclear ribonucleoprotein E1 (hnRNP E1). Purified specific rabbit anti-hnRNP E1 peptide antibodies (generated against a synthetic oligopeptide that was not represented in microsequenced peptides of the trans-factor) also recognized the purified trans-factor on Western blots. Conversely, the 18-base FR RNA cis-element also bound hnRNP E1 protein on Northwestern blots. Moreover, a 19-base RNA cis-element in the 3'-untranslated region of 15-lipoxygenase mRNA that is known to bind hnRNP E1 also interacted with placental 43-kDa trans-factor. In addition, several murine tissues containing a hnRNP E1-related protein (also known as alphaCP-1) readily interacted with the 18-base FR RNA cis-element. Finally, anti-hnRNP E1 antibodies specifically inhibited translation of FR in vitro in a dose-dependent manner, and the antibody effect could be reversed in a dose-dependent manner by either purified trans-factor or hnRNP E1. Collectively, the data favor identity of the FR mRNA-binding trans-factor and hnRNP E1, confirm its critical role in the translation of FR, and highlight yet another role of multifunctional hnRNP E1 in eukaryotic mRNA regulation.
Highlights
The interaction of an 18-base cis-element in the 5untranslated region of human folate receptor (FR)-␣ mRNA with a cytosolic trans-factor protein is critical for the translation of FR
In studies designed to identify the basis for the inverse relationship between the extracellular folate concentration and FR expression [8], we identified that the up-regulation of FR in cervical carcinoma cells is primarily at the translational level [9]
Because this FR mRNA-binding trans-factor was widely distributed in cells expressing little to no FR-␣, we predicted that this RNA-binding protein probably has additional functions that are unrelated to FR translation [10]
Summary
Further studies identified a 46-kDa cytosolic protein in human cervical carcinoma cells, which interacted with a CU-rich 18-base cis-element in the 5Ј-untranslated region (5Ј-UTR) of human FR-␣ mRNA, and that this interaction was critical for the translation of FR in vitro [10]. Because this FR mRNA-binding trans-factor was widely distributed in cells expressing little to no FR-␣, we predicted that this RNA-binding protein probably has additional functions that are unrelated to FR translation [10].
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