Dataset Information

The C-terminal tail of the NEIL1 DNA glycosylase interacts with the human mitochondrial single-stranded DNA binding protein.

ABSTRACT: The 16.5?kb mitochondrial genome is subjected to damage from reactive oxygen species (ROS) generated in the cell during normal cellular metabolism and external sources such as ionizing radiation and ultraviolet light. ROS cause harmful damage to DNA bases that could result in mutagenesis and various diseases, if not properly repaired. The base excision repair (BER) pathway is the primary pathway involved in maintaining the integrity of mtDNA. Several enzymes that partake in BER within the nucleus have also been identified in the mitochondria. The nei-like (NEIL) DNA glycosylases initiate BER by excising oxidized pyrimidine bases and others such as the ring-opened formamidopyrimidine and the hydantoin lesions. During BER, the NEIL enzymes interact with proteins that are involved with DNA replication and transcription. In the current manuscript, we detected NEIL1 in purified mitochondrial extracts from human cells and showed that NEIL1 interacts with the human mitochondrial single-stranded DNA binding protein (mtSSB) via its C-terminal tail using protein painting, far-western analysis, and gel-filtration chromatography. Finally, we scrutinized the NEIL1-mtSSB interaction in the presence and absence of a partial-duplex DNA substrate using a combination of multi-angle light scattering (MALS) and small-angle X-ray scattering (SAXS). The data indicate that NEIL1 and homotetrameric mtSSB form a larger ternary complex in presence of DNA, however, the tetrameric form of mtSSB gets disrupted by NEIL1 in the absence of DNA as revealed by the formation of a smaller NEIL1-mtSSBmonomer complex.

SUBMITTER: Sharma N

PROVIDER: S-EPMC5911420 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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The C-terminal tail of the NEIL1 DNA glycosylase interacts with the human mitochondrial single-stranded DNA binding protein.

Sharma Nidhi N Chakravarthy Srinivas S Longley Matthew J MJ Copeland William C WC Prakash Aishwarya A

DNA repair 20180306

The 16.5 kb mitochondrial genome is subjected to damage from reactive oxygen species (ROS) generated in the cell during normal cellular metabolism and external sources such as ionizing radiation and ultraviolet light. ROS cause harmful damage to DNA bases that could result in mutagenesis and various diseases, if not properly repaired. The base excision repair (BER) pathway is the primary pathway involved in maintaining the integrity of mtDNA. Several enzymes that partake in BER within the nucleu ...[more]

PMID: 29522991

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Project description:The maintenance of human mitochondrial DNA (mtDNA) is critical for proper cellular function as damage to mtDNA, if left unrepaired, can lead to a diverse array of pathologies. Of the pathways identified to participate in DNA repair within the mitochondria, base excision repair (BER) is the most extensively studied. Protein-protein interactions drive the step-by-step coordination required for the successful completion of this pathway and are important for crosstalk with other mitochondrial factors involved in genome maintenance. Human NEIL1 is one of seven DNA glycosylases that initiates BER in both the nuclear and mitochondrial compartments. In the current work, we scrutinized the interaction between NEIL1 and mitochondrial transcription factor A (TFAM), a protein that is essential for various aspects of mtDNA metabolism. We note, for the first time, that both the N- and C- terminal domains of NEIL1 interact with TFAM revealing a unique NEIL1 protein-binding interface. The interaction between the two proteins, as observed biochemically, appears to be transient and is most apparent at concentrations of low salt. The presence of DNA (or RNA) also positively influences the interaction between the two proteins, and molar mass estimates indicate that duplex DNA is required for complex formation at higher salt concentrations. Hydrogen deuterium exchange mass spectrometry data reveal that both proteins exchange less deuterium upon DNA binding, indicative of an interaction, and the addition of NEIL1 to the TFAM-DNA complex alters the interaction landscape. The transcriptional activity of TFAM appears to be independent of NEIL1 expression under normal cellular conditions, however, in the presence of DNA damage, we observe a significant reduction in the mRNA expression of TFAM-transcribed mitochondrial genes in the absence of NEIL1. Overall, our data indicate that the interaction between NEIL1 and TFAM can be modulated by local environment such as salt concentrations, protein availability, the presence of nucleic acids, as well as the presence of DNA damage.

Dataset Information

The C-terminal tail of the NEIL1 DNA glycosylase interacts with the human mitochondrial single-stranded DNA binding protein.

Publications

The C-terminal tail of the NEIL1 DNA glycosylase interacts with the human mitochondrial single-stranded DNA binding protein.

Similar Datasets

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