Characterization of Structural and Trans-Acting Elements of PLAC1 5’UTRs That Affect Translation of PLAC1

Abstract

The highly conserved developmental protein, Placenta Specific Protein 1 (PLAC1) is widely expressed in placental and fetal tissue during development. Its expression is restricted in normal tissue after birth, though it is reactivated and broadly expressed in many types of cancer. Expression of PLAC1 is controlled by two promoters, producing alternatively spliced PLAC1 transcripts, each with a unique 5’untranslated region (UTR) and identical protein coding sequences. Patterns of differential transcript expression resulting from alternative promoter usage are observed in both fetal development and cancer. To date, little is known about the post-transcriptional regulation of PLAC1 expression or the significance that 5’UTRs have on the translation of PLAC1. Using a GFP reporter system, the translational efficiency of each 5’UTR was measured. Overall, the 5’UTR sequences of promoter 2 transcripts had slightly lower efficiency than those from promoter 1 and mutation of the uORFs were minimally effective in restoring %GFP activity. Binding site motifs in PLAC1 were also analyzed using the RPBMapper and ATtRACT database. Binding sites for RBFOX proteins and ELAVL2, important alternative splicing proteins during fetal development, were identified 14 nucleotides apart in exon 4. Many of intronic binding sites for both of these proteins were also identified. The presence of binding sites for RBFOX and ELAVL2 proteins suggests that PLAC1 undergoes extensive alternative splicing during fetal development, potentially encoding additional isoforms that have not yet been identified.