Hypoxia alters splicing of the cancer associated Fas gene

Abstract

The removal of introns from mRNA precursors (pre-mRNAs) is an essential step in eukaryotic gene expression. The splicing machinery heavily contributes to biological complexity and especially to the ability of cells to adapt to altered cellular conditions. Hypoxia also plays a key role in the pathophysiology of many disease states. Recent studies have revealed that tumorigenesis and hypoxia involve large-scale alterations in alternative pre-mRNA splicing. Cancer associated Fas protein plays a central role in the physiological regulation of programmed cell death and has been implicated in the pathogenesis of various malignancies and diseases of the immune system. Fas pre-mRNA is alternatively spliced by excluding exon 6 to produce soluble Fas (sFas) protein that lacks a transmembrane domain and acts by inhibiting Fas mediated apoptosis. Another cancer related protein Rac1 plays an important regulatory role specifically in cells' motility, growth and survival. Rac pre-mRNA is alternatively spliced to produce Rac1b protein, which is upregulated in metastatic diseases. In the present study we, for the first time, show that anti-apoptotic Fas mRNA isoform formation is regulated by cellular microenvironment - hypoxia. Hypoxic microenvironment, however, does not influence Rac1 pre-mRNAs alternative splicing. Also our presented results indicate that splicing factors hnRNP A1 and SPF45, previously shown to regulate Fas alternative splicing in normoxic cells, are not involved in hypoxia dependent alternative Fas pre-mRNA splicing regulation in an amount dependent manner. Our observations on hypoxia dependent alternative Fas pre-mRNA splicing regulation indicate a probable involvement of other, yet unidentified splicing factors. Presented data also shows the contribution of pre-mRNA splicing to cell survival under unfavorable conditions.

Keywords: Cancer; Fas; Hypoxia; Pre-mRNA splicing; Rac1; SPF45; hnRNPA1.