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C‐terminal processing and C‐terminus of the mature pokeweed antiviral protein are required for its retrotranslocation from the ER to the cytosol
Author(s) -
Tumer Nilgun Ereken,
Baykal Ulku
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a1353-d
Subject(s) - signal peptide , endoplasmic reticulum , ricin , cytosol , ribosome , biology , biochemistry , n terminus , signal recognition particle , c terminus , peptide sequence , amino acid , microbiology and biotechnology , rna , toxin , enzyme , gene
Pokeweed antiviral protein (PAP) is a single chain (type I) ribosome‐inactivating protein (RIP), which depurinates ribosomes at the a‐sarcin/ricin loop of the large rRNA, resulting in inhibition of translation. Unlike the type II RIPs, which have an active and a binding moiety, PAP has only the active moiety. The mechanism by which toxins without a binding moiety enter cells is not known. We recently showed that PAP enters the cytosol from the endoplasmic reticulum (ER) in yeast and exhibits unusual stability in the cytosol. Using a series of PAP mutants, we identified the signal for this retrotranslocation event. Our results demonstrate that processing of the C‐terminal signal is critical for transport of the mature form into the cytosol. Sequential deletion of amino acids from the C‐terminus led to a gradual reduction in retrotranslocation, which was lost upon deletion of the last 13 amino acids. Single mutations in this sequence delayed retrotranslocation, indicating that the C‐terminal hydrophobic peptide is critical for ER export. This peptide showed sequence similarity to ribosome inactivating AB‐toxins that retrotranslocate. This is the first demonstration that a conserved C‐terminal sequence mediates retrotranslocation of a type I RIP and suggests that type I and type II RIPs may use a common signal to enter the cytosol. This work is supported by NSF MCB 0348299.