Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis

Alistair Taverner, Julia Mackay, Floriane Laurent, Tom Hunter, Keyi Liu, Khushdeep Mangat, Lisa Song, Elbert Seto, Sally Postlethwaite, Aatif Alam, Apurva Chandalia, Minji Seung, Mazi Saberi, Weijun Feng, Randy Mrsny

Research output: Contribution to journalArticle

Abstract

Cholix (Chx) is expressed by the intestinal pathogen Vibrio cholerae as a single chain of 634 amino acids (~70.7 kDa protein) that folds into three distinct domains, with elements of the second and third domains being involved in accessing the cytoplasm of nonpolarized cells and inciting cell death via ADP-ribosylation of elongation factor 2, respectively. In order to reach nonpolarized cells within the intestinal lamina propria, however, Chx must cross the polarized epithelial barrier in an intact form. Here, we provide invitro and invivo demonstrations that a nontoxic Chx transports across intestinal epithelium via a vesicular trafficking pathway that rapidly achieves vesicular apical to basal (A→B) transcytosis and avoids routing to lysosomes. Specifically, Chx traffics in apical endocytic Rab7+ vesicles and in basal exocytic Rab11+ vesicles with a transition between these domains occurring in the ER-Golgi intermediate compartment (ERGIC) through interactions with the lectin mannose-binding protein 1 (LMAN1) protein that undergoes an intracellular re-distribution that coincides with the re-organization of COPI+ and COPII+ vesicular structures. Truncation studies demonstrated that domain I of Chx alone was sufficient to efficiently complete A→B transcytosis and capable of ferrying genetically conjoined human growth hormone (hGH). These studies provide evidence for a pathophysiological strategy where native Chx exotoxin secreted in the intestinal lumen by nonpandemic V. cholerae can reach nonpolarized cells within the lamina propria in an intact form by using a nondestructive pathway to cross in the intestinal epithelial that appears useful for oral delivery of biopharmaceuticals. One-Sentence Summary: Elements within the first domain of the Cholix exotoxin protein are essential and sufficient for the apical to basal transcytosis of this Vibrio cholerae-derived virulence factor across polarized intestinal epithelial cells.
Original languageEnglish
Article number1710429
JournalTissue Barriers
Early online date13 Jan 2020
DOIs
Publication statusE-pub ahead of print - 13 Jan 2020

Cite this

Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis. / Taverner, Alistair; Mackay, Julia; Laurent, Floriane; Hunter, Tom; Liu, Keyi; Mangat, Khushdeep; Song, Lisa; Seto, Elbert; Postlethwaite, Sally; Alam, Aatif; Chandalia, Apurva; Seung, Minji; Saberi, Mazi; Feng, Weijun; Mrsny, Randy.

In: Tissue Barriers, 13.01.2020.

Research output: Contribution to journalArticle

Taverner, A, Mackay, J, Laurent, F, Hunter, T, Liu, K, Mangat, K, Song, L, Seto, E, Postlethwaite, S, Alam, A, Chandalia, A, Seung, M, Saberi, M, Feng, W & Mrsny, R 2020, 'Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis', Tissue Barriers. https://doi.org/10.1080/21688370.2019.1710429
Taverner, Alistair ; Mackay, Julia ; Laurent, Floriane ; Hunter, Tom ; Liu, Keyi ; Mangat, Khushdeep ; Song, Lisa ; Seto, Elbert ; Postlethwaite, Sally ; Alam, Aatif ; Chandalia, Apurva ; Seung, Minji ; Saberi, Mazi ; Feng, Weijun ; Mrsny, Randy. / Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis. In: Tissue Barriers. 2020.
@article{6cca67f27f2a4395b7e4052974c4b597,
title = "Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis",
abstract = "Cholix (Chx) is expressed by the intestinal pathogen Vibrio cholerae as a single chain of 634 amino acids (~70.7 kDa protein) that folds into three distinct domains, with elements of the second and third domains being involved in accessing the cytoplasm of nonpolarized cells and inciting cell death via ADP-ribosylation of elongation factor 2, respectively. In order to reach nonpolarized cells within the intestinal lamina propria, however, Chx must cross the polarized epithelial barrier in an intact form. Here, we provide invitro and invivo demonstrations that a nontoxic Chx transports across intestinal epithelium via a vesicular trafficking pathway that rapidly achieves vesicular apical to basal (A→B) transcytosis and avoids routing to lysosomes. Specifically, Chx traffics in apical endocytic Rab7+ vesicles and in basal exocytic Rab11+ vesicles with a transition between these domains occurring in the ER-Golgi intermediate compartment (ERGIC) through interactions with the lectin mannose-binding protein 1 (LMAN1) protein that undergoes an intracellular re-distribution that coincides with the re-organization of COPI+ and COPII+ vesicular structures. Truncation studies demonstrated that domain I of Chx alone was sufficient to efficiently complete A→B transcytosis and capable of ferrying genetically conjoined human growth hormone (hGH). These studies provide evidence for a pathophysiological strategy where native Chx exotoxin secreted in the intestinal lumen by nonpandemic V. cholerae can reach nonpolarized cells within the lamina propria in an intact form by using a nondestructive pathway to cross in the intestinal epithelial that appears useful for oral delivery of biopharmaceuticals. One-Sentence Summary: Elements within the first domain of the Cholix exotoxin protein are essential and sufficient for the apical to basal transcytosis of this Vibrio cholerae-derived virulence factor across polarized intestinal epithelial cells.",
author = "Alistair Taverner and Julia Mackay and Floriane Laurent and Tom Hunter and Keyi Liu and Khushdeep Mangat and Lisa Song and Elbert Seto and Sally Postlethwaite and Aatif Alam and Apurva Chandalia and Minji Seung and Mazi Saberi and Weijun Feng and Randy Mrsny",
year = "2020",
month = "1",
day = "13",
doi = "10.1080/21688370.2019.1710429",
language = "English",
journal = "Tissue Barriers",
issn = "2168-8362",
publisher = "Taylor and Francis",

}

TY - JOUR

T1 - Cholix Protein Domain I Functions as a Carrier Element for Efficient Apical to Basal Epithelial Transcytosis

AU - Taverner, Alistair

AU - Mackay, Julia

AU - Laurent, Floriane

AU - Hunter, Tom

AU - Liu, Keyi

AU - Mangat, Khushdeep

AU - Song, Lisa

AU - Seto, Elbert

AU - Postlethwaite, Sally

AU - Alam, Aatif

AU - Chandalia, Apurva

AU - Seung, Minji

AU - Saberi, Mazi

AU - Feng, Weijun

AU - Mrsny, Randy

PY - 2020/1/13

Y1 - 2020/1/13

N2 - Cholix (Chx) is expressed by the intestinal pathogen Vibrio cholerae as a single chain of 634 amino acids (~70.7 kDa protein) that folds into three distinct domains, with elements of the second and third domains being involved in accessing the cytoplasm of nonpolarized cells and inciting cell death via ADP-ribosylation of elongation factor 2, respectively. In order to reach nonpolarized cells within the intestinal lamina propria, however, Chx must cross the polarized epithelial barrier in an intact form. Here, we provide invitro and invivo demonstrations that a nontoxic Chx transports across intestinal epithelium via a vesicular trafficking pathway that rapidly achieves vesicular apical to basal (A→B) transcytosis and avoids routing to lysosomes. Specifically, Chx traffics in apical endocytic Rab7+ vesicles and in basal exocytic Rab11+ vesicles with a transition between these domains occurring in the ER-Golgi intermediate compartment (ERGIC) through interactions with the lectin mannose-binding protein 1 (LMAN1) protein that undergoes an intracellular re-distribution that coincides with the re-organization of COPI+ and COPII+ vesicular structures. Truncation studies demonstrated that domain I of Chx alone was sufficient to efficiently complete A→B transcytosis and capable of ferrying genetically conjoined human growth hormone (hGH). These studies provide evidence for a pathophysiological strategy where native Chx exotoxin secreted in the intestinal lumen by nonpandemic V. cholerae can reach nonpolarized cells within the lamina propria in an intact form by using a nondestructive pathway to cross in the intestinal epithelial that appears useful for oral delivery of biopharmaceuticals. One-Sentence Summary: Elements within the first domain of the Cholix exotoxin protein are essential and sufficient for the apical to basal transcytosis of this Vibrio cholerae-derived virulence factor across polarized intestinal epithelial cells.

AB - Cholix (Chx) is expressed by the intestinal pathogen Vibrio cholerae as a single chain of 634 amino acids (~70.7 kDa protein) that folds into three distinct domains, with elements of the second and third domains being involved in accessing the cytoplasm of nonpolarized cells and inciting cell death via ADP-ribosylation of elongation factor 2, respectively. In order to reach nonpolarized cells within the intestinal lamina propria, however, Chx must cross the polarized epithelial barrier in an intact form. Here, we provide invitro and invivo demonstrations that a nontoxic Chx transports across intestinal epithelium via a vesicular trafficking pathway that rapidly achieves vesicular apical to basal (A→B) transcytosis and avoids routing to lysosomes. Specifically, Chx traffics in apical endocytic Rab7+ vesicles and in basal exocytic Rab11+ vesicles with a transition between these domains occurring in the ER-Golgi intermediate compartment (ERGIC) through interactions with the lectin mannose-binding protein 1 (LMAN1) protein that undergoes an intracellular re-distribution that coincides with the re-organization of COPI+ and COPII+ vesicular structures. Truncation studies demonstrated that domain I of Chx alone was sufficient to efficiently complete A→B transcytosis and capable of ferrying genetically conjoined human growth hormone (hGH). These studies provide evidence for a pathophysiological strategy where native Chx exotoxin secreted in the intestinal lumen by nonpandemic V. cholerae can reach nonpolarized cells within the lamina propria in an intact form by using a nondestructive pathway to cross in the intestinal epithelial that appears useful for oral delivery of biopharmaceuticals. One-Sentence Summary: Elements within the first domain of the Cholix exotoxin protein are essential and sufficient for the apical to basal transcytosis of this Vibrio cholerae-derived virulence factor across polarized intestinal epithelial cells.

U2 - 10.1080/21688370.2019.1710429

DO - 10.1080/21688370.2019.1710429

M3 - Article

C2 - 31928299

JO - Tissue Barriers

JF - Tissue Barriers

SN - 2168-8362

M1 - 1710429

ER -