Protein sequence editing defines distinct and overlapping functions of SKN-1A/Nrf1 and SKN-1C/Nrf2.

Nrf/NFE2L family transcription factors regulate redox balance, metabolism, proteostasis, and aging. Nrf1/NFE2L1 is responsible for stress-responsive upregulation of proteasome subunit genes and is essential for adaptation to proteotoxic stress. The closely related Nrf2/NFE2L2 is responsible for activation of oxidative stress responses and xenobiotic detoxification. Although regulated by different mechanisms, Nrf1 and Nrf2 contain very similar DNA binding domains and can drive similar transcriptional responses. However, the extent to which functions of Nrf1/2 are distinct or overlapping has been unclear. In C. elegans, a single gene, skn-1, encodes distinct protein isoforms, SKN-1A and SKN-1C, that function analogously to mammalian Nrf1 and Nrf2, respectively. We previously showed that regulation of the proteasome by SKN-1A/Nrf1 requires post-translational conversion of N-glycosylated asparagine residues to aspartate by the PNG-1/NGLY1 peptide:N-glycanase, a process we term 'sequence editing'. Here, we reveal the consequences of sequence editing for the transcriptomic output of activated SKN-1A. We show that whilst activation of proteasome subunit genes is strictly dependent on sequence editing, sequence edited SKN-1A also activate genes linked to redox homeostasis and xenobiotic detoxification that are also activated by non-sequence edited forms of SKN-1. Using mutant alleles that selectively inactivate either SKN-1A or SKN-1C, we show that SKN-1A/Nrf1 and SKN-1C/Nrf2 function coordinately to promote optimal oxidative stress resistance and confirm that they are regulated by discrete genetic pathways. This work demonstrates that sequence editing plays a critical role in tailoring SKN-1/Nrf functions by tuning the SKN-1A/Nrf1 regulated transcriptome. Overall design: We used RNAseq to compare gene expression in animals expressing trunacted forms of the SKN-1 transcription factor that are altered with different patterns of Asn to Asp substitutions. For each transgenic strain, we compared gene expression to stage-matched wild type (non-transgenic) control samples. Samples were prepared from synchronized L4 stage animals. We analyzed four replicate samples for each of three transgenic strains (GR3105, NJL2494, NJL2492) and four replicate samples of the wild type control strain (N2).