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Vol. 19, Issue 3, 877-884, March 2008

This Article Full Text Full Text (PDF) All Versions of this Article: E07-08-0755v1 19/3/877    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Bernardi, K. M. Articles by Tsai, B. PubMed PubMed Citation Articles by Bernardi, K. M. Articles by Tsai, B.

Derlin-1 Facilitates the Retro-Translocation of Cholera Toxin

Kaleena M. Bernardi^*, Michele L. Forster^*, Wayne I. Lencer, and Billy Tsai^*

*Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109; and GI Cell Biology, Children's Hospital, Harvard Medical School, Boston, MA 02115

Submitted August 6, 2007; Revised December 5, 2007; Accepted December 12, 2007
Monitoring Editor: Jeffrey Brodsky

Cholera toxin (CT) intoxicates cells by using its receptor-binding B subunit (CTB) to traffic from the plasma membrane to the endoplasmic reticulum (ER). In this compartment, the catalytic A1 subunit (CTA1) is unfolded by protein disulfide isomerase (PDI) and retro-translocated to the cytosol where it triggers a signaling cascade, leading to secretory diarrhea. How CT is targeted to the site of retro-translocation in the ER membrane to initiate translocation is unclear. Using a semipermeabilized-cell retro-translocation assay, we demonstrate that a dominant-negative Derlin-1-YFP fusion protein attenuates the ER-to-cytosol transport of CTA1. Derlin-1 interacts with CTB and the ER chaperone PDI as assessed by coimmunoprecipitation experiments. An in vitro membrane-binding assay showed that CTB stimulated the unfolded CTA1 chain to bind to the ER membrane. Moreover, intoxication of intact cells with CTB stabilized the degradation of a Derlin-1–dependent substrate, suggesting that CT uses the Derlin-1 pathway. These findings indicate that Derlin-1 facilitates the retro-translocation of CT. CTB may play a role in this process by targeting the holotoxin to Derlin-1, enabling the Derlin-1–bound PDI to unfold the A1 subunit and prepare it for transport.

Address correspondence to: Billy Tsai (btsai{at}umich.edu)

Abbreviations used: CT, cholera toxin; PDI, protein disulfide isomerase; YFP, yellow fluorescent protein.