Premium
The Genome‐wide Landscape of Oxidative Base Damage in Hypoxia; Potential Role in Metabolic Reprogramming in Chronic Lung Disease
Author(s) -
Daly Grant,
Houserova Dominika,
Francis Christopher Michael,
Andrews Joel,
Langley Raymond J.,
Pastukh Viktor M.,
Bass Hank W.,
Borchert Glen M.,
Gillespie Mark N.
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.04056
Subject(s) - biology , reprogramming , transcriptome , chromatin , microbiology and biotechnology , transcription factor , dna damage , gene , genetics , gene expression , dna
Hypoxia, a common attribute of many lung diseases, causes an adaptive response characterized by glycolytic reprogramming, which is initiated in part by the master transcriptional regulator, Hif‐1, accumulating in response to reactive oxygen species (ROS) generated as second messengers in hypoxic signaling. ROS produced in hypoxia also appear to engage a DNA damage and repair pathway in promoter regulatory elements that impacts core events in transcription (Pastukh, et al. , 2015). Because the genome‐wide disposition of base damage in hypoxia could provide clues about mechanisms of persistent metabolic reprogramming in the lung, we executed a proof‐of‐concept study to define the landscape of the common base oxidation product 8‐oxoguanine (8‐oxoG) in the genomes of normoxic and acutely hypoxic human umbilical vein endothelial cells (HUVECs), focusing on localization of base damage within genes comprising the hypoxic transcriptome. Using ChIP‐seq analysis to delineate sequences harboring 8‐oxoG, we found that base damage was enriched in multiple regulatory regions, including those defined by transcription factor binding motifs and epigenetic marks. Of particular interest, base damage in these regions occurred in distinctly different genes in normoxic and hypoxic cells. SIM super‐resolution immunofluorescence microscopy revealed prominent base damage in facultative heterochromatin residing in close proximity to chromatin‐free channels that appeared to be in direct communication with the nuclear membrane. A comparison of sequences demarcated by 8‐oxoG ChIP‐seq signals to the RNA‐seq‐defined hypoxic transcriptome revealed that base damage was prominent in genes occupying regulatory nodes in pathways controlling the metabolic response to hypoxia as well as other key adaptive pathways. Finally, after noting that sequences harboring 8‐oxoG displayed elevated variant densities, we discovered that a high proportion of these variants were likely mutational artifacts since they were corrected by enzymatic excision of damaged bases in template DNA prior to PCR‐based sequencing library preparation. Collectively, these findings suggest that hypoxia fundamentally alters the genomic landscape of 8‐oxoG in HUVECs, causing accumulation or dissipation of the base oxidation product in non‐coding regulatory regions in differentially‐transcribed genes. The presence of base damage‐associated mutational artifacts supports the concept that the DNA damage and repair pathway engaged during hypoxic transcriptional signaling could impose a mutagenic threat leading to formation of permanent and currently uncharacterized somatic variants with the potential to drive persistent metabolic dysregulation in hypoxia‐related lung diseases. Support or Funding Information NIH