Identification of CED-3 Substrates by a Yeast-Based Screening Method

Author: Kim Sung Yun   Valencia Marcela   Lee Eui Seung   Park Daeho   Oh Myungsok   Xue Ding   Park Woo Jin  

Publisher: Humana Press, Inc

ISSN: 1073-6085

Source: Molecular Biotechnology, Vol.27, Iss.1, 2004-05, pp. : 1-6

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Abstract

Identifying cellular substrates repertoire of individual proteases will facilitate our understanding of their physiological and pathological roles. In this article, we employed a yeast-based screening method to isolate CED-3 substrates. This method uses a transcription factor anchored to the plasma membrane by fusion to a library of cellular protein sequences. When a fusion protein is cleaved by CED-3, the transcription factor is released from the plasma membrane and enters the nucleus where it turns on the expression of reporter genes. We identified seven candidate clones by screening a genomic library using this method. Of these seven clones, two were cleaved by purified CED-3 in vitro. Therefore, the method described here may be generally used for genomewide screening to isolate potential substrates of specific proteases.