Retroviral insertional mutagenesis identifies a small protein required for synthesis of diphthamide, the target of bacterial ADP-ribosylating toxins

Shihui Liu; Stephen H Leppla

doi:10.1016/j.molcel.2003.08.003

Retroviral insertional mutagenesis identifies a small protein required for synthesis of diphthamide, the target of bacterial ADP-ribosylating toxins

Mol Cell. 2003 Sep;12(3):603-13. doi: 10.1016/j.molcel.2003.08.003.

Authors

Shihui Liu¹, Stephen H Leppla

Affiliation

¹ Microbial Pathogenesis Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

PMID: 14527407
DOI: 10.1016/j.molcel.2003.08.003

Abstract

Retroviral insertional mutagenesis was used to produce a mutant Chinese hamster ovary cell line that is completely resistant to several different bacterial ADP-ribosylating toxins. The gene responsible for toxin resistance, termed diphtheria toxin (DT) and Pseudomonas exotoxin A (ETA) sensitivity required gene 1 (DESR1), encodes two small protein isoforms of 82 and 57 residues. DESR1 is evolutionally conserved and ubiquitously expressed. Only the longer isoform is functional because the mutant cell line can be complemented by transfection with the long but not the short isoform. We demonstrate that DESR1 is required for the first step in the posttranslational modification of elongation factor-2 at His(715) that yields diphthamide, the target site for ADP ribosylation by DT and ETA. KTI11, the analog of DESR1 in yeast, which was originally identified as a gene regulating the sensitivity of yeast to zymocin, is also required for diphthamide biosynthesis, implicating DESR1/KTI11 in multiple biological processes.

Publication types

Research Support, U.S. Gov't, P.H.S.

MeSH terms

ADP-Ribosylation Factors / metabolism
Animals
Bacterial Infections / immunology*
Bacterial Toxins / metabolism*
CHO Cells
Cricetinae
Eukaryotic Cells / metabolism
Eukaryotic Cells / microbiology
Histidine / analogs & derivatives*
Histidine / biosynthesis
Histidine / metabolism*
Immunity, Innate / genetics
Molecular Sequence Data
Molecular Weight
Mutagenesis, Insertional*
Mutation / genetics
Peptide Elongation Factor 2 / genetics
Peptide Elongation Factor 2 / metabolism
Protein Isoforms / genetics
Protein Isoforms / isolation & purification
Protein Processing, Post-Translational
Repressor Proteins / genetics
Repressor Proteins / isolation & purification*
Retroviridae / genetics
Saccharomyces cerevisiae Proteins / genetics
Saccharomyces cerevisiae Proteins / isolation & purification*
Sequence Homology, Amino Acid
Sequence Homology, Nucleic Acid

Substances

Bacterial Toxins
KTI11 protein, S cerevisiae
Peptide Elongation Factor 2
Protein Isoforms
Repressor Proteins
Saccharomyces cerevisiae Proteins
Histidine
diphthamide
ADP-Ribosylation Factors