EFFECT OF DIPHTHERIA TOXIN T-DOMAIN ON ENDOSOMAL pH.

Palladin Institute Of Biochemistry, Nas Of Ukraine, Kyiv ,A J Labyntsev,D V Kolybo,N V Korotkevych,S V Komisarenko

doi:10.15407/ubj87.04.013

Palladin Institute Of Biochemistry, Nas Of Ukraine, Kyiv , A J Labyntsev + Show 3 more

Open Access

https://doi.org/10.15407/ubj87.04.013

Copy DOI

Abstract

A key step in the mode of cytotoxic action of diphtheria toxin (DT) is the transfer of its catalytic domain (Cd) from endosomes into the cytosol. The main activity in this process is performed by the transport domain (Td), but the molecular mechanism of its action remains unknown. We have previously shown that Td can have some influence on the endosomal transport of DT The aim of this work was to study the effect of diphtheria toxin on the toxin compartmentalization in the intracellular transporting pathway and endosomal pH. We used recombinant fragments of DT which differed only by the presence of Td in their structure, fused with fluorescent proteins. It was shown that the toxin fragment with Td moved slower by the pathway early-late endosomes-lysosomes, and had a slightly different pattern of colocalization with endosomal markers than DT fragment without Td. In addition, endosomes containing DT fragments with Td had a constant pH of about 6.5 from the 10th to 50th minute of observation, for the same time endosomes containing DT fragments without Td demonstrated a decrease in pH from 6.3 to 5.5. These results indicate that Td inhibits acidification of endosomal medium. One of possible explanations for this may be the effect of the ion channel formed by the T-domain on the process of the endosomal acidification. This property of Td may not only inhibit maturation of endosomes but also inhibit activation of endosomal pH-dependent proteases, and this promotes successful transport of Cd into the cell cytosol.

Highlights

D iphtheria is a bacterial infection of the upper respiratory tract with possible complications in tissues and organs of the affected organism
Toxin is assigned to AB group of bacterial toxins, it is divided into a subunit A responsible for the enzymatic activity, which corresponds to Cdomain, and a subunit B (SubB), includes Td and Rd and responsible for the transport of the subunit A
The main marker of maturation and late endosomes (LE) formation is the formation of multivesicular bodies (MVB) which are the endosomes included as the bodies in EE

Summary

Introduction

D iphtheria is a bacterial infection of the upper respiratory tract with possible complications in tissues and organs of the affected organism (the kidneys, heart, nervous system, etc.). The disease is caused by toxigenic strains of Corynebacterium diphtheriae, which can synthesize the main pathogenicity factor – diphtheria toxin (DT). This protein is a simple polypeptide of 535 amino acids long. The endosomal cell apparatus consists of seve ral main compartments, where vesicles can arrive after the process of its formation. Such compartments may include early endosomes (EE) which can be headed to recycling endosomes (RE) or maturate to late endosomes (LE) and to lysosomes [2]. The aim of this work was to investigate the peculiarities of such effect studying the pathways of intracellular transportation of DT fragments and Td effect on the endosomal pH

Objectives

Methods

Results