Glyceraldehyde-3-phosphate dehydrogenase is required for efficient repair of cytotoxic DNA lesions in Escherichia coli

Abstract

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional protein with diverse biological functions in human cells. In bacteria, moonlighting GAPDH functions have only been described for the secreted protein in pathogens or probiotics. At the intracellular level, we previously reported the interaction of Escherichia coli GAPDH with phosphoglycolate phosphatase, a protein involved in the metabolism of the DNA repair product 2-phosphoglycolate, thus suggesting a putative role of GAPDH in DNA repair processes. Here, we provide evidence that GAPDH is required for the efficient repair of DNA lesions in E. coli. We show that GAPDH-deficient cells are more sensitive to bleomycin or methyl methanesulfonate. In cells challenged with these genotoxic agents, GAPDH deficiency results in reduced cell viability and filamentous growth. In addition, the gapA knockout mutant accumulates a higher number of spontaneous abasic sites and displays higher spontaneous mutation frequencies than the parental strain. Pull-down experiments in different genetic backgrounds show interaction between GAPDH and enzymes of the base excision repair pathway, namely the AP-endonuclease Endo IV and uracil DNA glycosylase. This finding suggests that GAPDH is a component of a protein complex dedicated to the maintenance of genomic DNA integrity. Our results also show interaction of GAPDH with the single-stranded DNA binding protein. This interaction may recruit GAPDH to the repair sites and implicates GAPDH in DNA repair pathways activated by profuse DNA damage, such as homologous recombination or the SOS response.

Identification of Bacterial Protein Interaction Partners Points to New Intracellular Functions of Francisella tularensis Glyceraldehyde-3-Phosphate Dehydrogenase

“…These so-called moonlighting roles are associated with GAPDH's unusual, non-predictable localization. Such multifunctionality also has been reported for bacterial GAPDH in a number of independent studies (Pancholi and Fischetti, 1992;Modun and Williams, 1999;Boël et al, 2005;Butland et al, 2005;Arifuzzaman et al, 2006;Jin et al, 2011;Ferreira et al, 2015). In some human pathogens, the surface-localized and/or -secreted GAPDH is able to bind to several host proteins (Kopeckova et al, 2020), thus contributing to its virulence.…”

“…The results of a transcriptomic analysis on Streptococcus pyogenes mutant strain displaying higher intracellular levels of GAPDH suggest it has a role in transcriptional modulation of virulence and metabolic genes ( Boël et al, 2005 ). Several large-scale protein–protein interaction studies performed on Escherichia coli have revealed that bacterial GAPDH, similar to its eukaryotic orthologs or analogs, is able to interact with other intracellular proteins, including metabolic enzymes as well as proteins involved in transcription, protein synthesis, and DNA repair ( Butland et al, 2005 ; Arifuzzaman et al, 2006 ; Ferreira et al, 2013 , 2015 ).…”

Dimethyl fumarate, a two‐edged drug: Current status and future directions

“…It plays a fundamental role in energy metabolism and the generation of ATP along with pyruvate during glycolysis. It is also involved in DNA repair, DNA replication, transcription activation, and cell death . GAPDH has a number of cysteine residues that can be easily modified by fumarate leading to change in function of the enzyme .…”

Diverse Localization and Protein Binding Abilities of Glyceraldehyde-3-Phosphate Dehydrogenase in Pathogenic Bacteria: The Key to its Multifunctionality?

“…For further verification and functional analyses, Ferreira et al (2013) selected phosphoglycolate phosphatase (Gph), an enzyme involved in DNA repair of 3 ′ -phosphoglycolate ends caused by oxidative stress mediated by the radiomimetic agent bleomycin (Povirk, 1996;Pellicer et al, 2003). Further observations have pointed to a role of bacterial DNA repair machinery also in E. coli (Ferreira et al, 2015). GAPDH was shown to interact with other repair enzymes AP-endonuclease Endo IV and uracil DNA glycosylase.…”