| Summary: | 博士 === 國防醫學院 === 生命科學研究所 === 93 === Apoptosis is a morphologically distinct form of cell death involved in many physiological and pathological processes. And the regulation of Fas/Apo-1 involved in membrane-mediated apoptosis has also been known to play crucial roles in many systems. Here, we identified one novel alternative splicing variants of human Fas gene (containing additional 48-bp) and detected in various cell lines examined. However, we could not see any apparently difference from original functional Fas by cloning and transfection. Interestingly, we found several IMAGE clones containing this 48-bp but in a reverse orientation to Fas transcript. If this antisense transcript is true, it might regulate the Fas gene. Therefore, we shifted our focus to this antisene transcript.
More and more naturally occurring antisense RNAs are now known to regulate, at least in part, a growing number of eukaryotic genes. In this report we describe the findings of a novel RNA transcribed from the opposite strand of the intron 1 of the human Fas gene. Using orientation-specific RT-PCR and Northern blot analysis we show that this transcript is 1.5 kb in length and was expressed in several human tissues and cell lines. This transcript was cloned by 5’ and 3’RACE (rapid amplification of cDNA ends) and the transcription start site was determined by primer extension. This novel gene was named Saf.
To assess the functions of Saf, Jurkat cells transfected with human Saf or control vector was prepared. The stable Saf-transfectant was highly resistant to Fas-mediated but not to TNF-a-mediated apoptosis. While the overall mRNA expression level of Fas was not affected, expression of some novel forms of Fas transcripts was increased in Saf-transfectant, especially the inhibitory soluble forms. These findings collectively suggest that Saf might protect T lymphocytes from Fas-mediated apoptosis by blocking the binding of FasL or its agonistic Fas antibody. Saf might regulate the expression of Fas alternative splice forms through pre-mRNA processing.
To the best of our knowledge, this was the first report that Fas might be regulated by an antisense mechanism. In this study, we proposed that Saf directly, at least in part, involved in regulation of Fas pre-mRNA alternative splicing in providing a possible explanation for antiapototic activity of Saf.
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