Identification of the human PMR1 mRNA endonuclease as an alternatively processed product of the gene for peroxidasin-like protein

Abstract

The PMR1 endonuclease was discovered in Xenopus liver and identified as a member of the large and diverse peroxidase gene family. The peroxidase genes arose from multiple duplication and rearrangement events, and their high degree of sequence similarity confounded attempts to identify human PMR1. The functioning of PMR1 in mRNA decay depends on the phosphorylation of a tyrosine in the C-terminal polysome targeting domain by c-Src. The sequences of regions that are required for c-Src binding and phosphorylation of Xenopus PMR1 were used to inform a bioinformatics search that identified two related genes as potential candidates for human PMR1: peroxidasin homolog (PXDN) and peroxidasin homolog-like (PXDNL) protein. Although each of these genes is predicted to encode a large, multidomain membrane-bound peroxidase, alternative splicing of PXDNL pre-mRNA yields a transcript whose predicted product is a 57-kDa protein with 42% sequence identity to Xenopus PMR1. Results presented here confirm the existence of the predicted 57-kDa protein, show this is the only form of PXDNL detected in any of the human cell lines examined, and confirm its identity as human PMR1. Like the Xenopus protein, human PMR1 binds to c-Src, is tyrosine phosphorylated, sediments on polysomes, and catalyzes the selective decay of a PMR1 substrate mRNA. Importantly, the expression of human PMR1 stimulates cell motility in a manner similar to that of the Xenopus PMR1 expressed in human cells, thus providing definitive evidence linking endonuclease decay to the regulation of cell motility.

The human PMR1 endonuclease stimulates cell motility by down regulating miR-200 family microRNAs

“…The 57 kDa hPMR1 protein is cytoplasmic, and it is the only form of PXDNL detectable in a number of cancer cell lines, including U2OS, K562, MCF-7 and MDA-MB-231. We previously showed that the motility of U2OS cells was increased following expression of Xenopus PMR1 from a tetracycline-inducible promoter ( 6 ), and similar results were seen for hPMR1 in MCF-7 breast cancer cells ( 5 ). MCF-7 cells are not particularly motile or invasive, but become both motile and invasive following suppression of miR-200 family microRNAs ( 7 ).…”

“…The creation of tetracycline-inducible lines of MCF-7 cells and cells knocked down for hPMR1 were described in ( 5 ). These were maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS), 2 mM l -glutamine, 1.0 mM sodium pyruvate, and 10 mM Hepes and 4.5 g/l glucose until 3 days before the start of each experiment.…”

“…hPMR1 induction was achieved by adding 100 or 400 ng/ml doxycycline to the medium at the indicated times. siRNA knockdowns were performed as described previously ( 5 ).…”

“…Human PMR1 (hPMR1) is a 57 kDa protein that is expressed from an alternatively spliced form of peroxidasin homolog (Drosophila)-like protein (PXDNL) mRNA ( 5 ). PXDNL, also known as cardiac peroxidase, is a 164 kDa membrane-bound protein that is found predominately in heart and aorta.…”

A six-mRNA signature-based model for the prognosis prediction of breast cancer

“…Finally, we constructed a prognostic gene signature model by using multivariate Cox analysis, and screened 6 BC immunity-related feature genes (PXDNL, FABP7, STX11, PIGR, SHISAL2A, WDR17) which could be used as independent prognostic factors. PXDNL (Peroxidasin like), a member of the peroxidase gene family, has a variety of biological functions, including hormone biosynthesis, host defense and cell movement26 . A recent study by Li et al has reported that PXDNL is closely related to the development of BC, and the high expression of PXDNL is a potential independent prognostic indicator of BC27 .…”