Document Type


Date of Degree

Fall 2009

Degree Name

PhD (Doctor of Philosophy)

Degree In


First Advisor

Sheff, David R

First Committee Member

Gardinier, Minnetta

Second Committee Member

Stamnes, Mark

Third Committee Member

Strack, Stefan

Fourth Committee Member

Yeaman, Charles


Shiga toxin and cholera toxin are members of the AB5 family of protein exotoxins. The A subunit is the enzymatic subunit, whereas the pentameric B subunit binds cell surface receptors and carries the A subunit to the endoplasmic reticulum (ER) where it can be released into the cytosol. The B-subunits (STxB or CTxB) mediate toxin traffic along the retrograde pathway from the plasma membrane to the ER via early/recycling endosomes and the Golgi apparatus. It is unknown if STxB requires transport through the Golgi, or if it is just kinetically favorable. It is also unknown if the recycling endosome (RE) plays a role in the retrograde transport of STxB and CTxB. The first goal of this dissertation research was to demonstrate that transport through the Golgi is required for STxB to reach the ER. Using aluminum fluoride treatment, a simple temperature block, and cytoplast studies, I show that Golgi transport is necessary for STxB to reach the ER. The second goal of this dissertation research was to tease apart how STxB and CTxB move through early and recycling endosomes as well as elucidate a mechanism of how STxB exits endosomes en route to the Golgi. The role of the RE in STxB and CTxB transport is unclear. I used transferrin colocalization and temperature block studies to show that STxB and CTxB traffic through the RE. I then used HRP ablation of the RE to show that STxB requires the RE to reach the Golgi. I also examined the role of an RE-specific protein, EHD1, in exit of STxB from the RE. EHD1 has been previously shown to regulate recycling Tfn exit from the RE but its role in STxB transport is unknown. Expression of a dominant negative form of EHD1 arrested STxB at the RE and prevented it from reaching the Golgi. Together, these results suggest that STxB and CTxB transit the RE, STxB requires a functional RE for normal retrograde trafficking, and that STxB exit from the RE is regulated by EHD1.


cholera toxin, EHD1, endosomes, membrane transport, recycling endosome, Shiga toxin


xii, 93 pages


Includes bibliographical references (pages 84-93).


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Copyright © 2009 Jenna Elyse McKenzie

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