# DExH-Box Helicase 9 Participates in De Novo Nrf2 Protein Translation Under Oxidative Stress

## TL;DR

The paper shows how the protein DHX9 helps translate the Nrf2 protein under oxidative stress by interacting with its mRNA.

## Contribution

The study identifies DHX9 as a novel factor involved in de novo Nrf2 translation under oxidative stress.

## Key findings

- DHX9 binds to the Nrf2 mRNA 5′UTR and its binding increases with oxidative stress.
- DHX9 interacts with the 43S preinitiation complex to facilitate Nrf2 translation.
- DHX9 may remove inhibitory RNA elements to allow translation initiation under stress.

## Abstract

Nrf2 transcript factor plays an important role in cellular defense against oxidative stress due to its control for expression of antioxidant and detoxification genes. We have found that Nrf2 gene undergoes de novo protein translation when mammalian cells encounter oxidative stress. Here, we report the discovery of DExH-box helicase-9 (DHX9), also known as RNA helicase A, as a binding protein for Nrf2 mRNA at 5′UTR. DHX9 binding to Nrf2 5′UTR increased with increasing doses (50–300 μM) of H2O2 or treatment time (10–120 min). This incease was in parallel with elevation of Nrf2 protein. Inhibiting DHX9 expression with siRNA or its activity with YK-4-279 inhibitor blocked H2O2 from inducing Nrf2 mRNA recruitment to the ribosomes or Nrf2 protein elevation. As a nuclear protein, DHX9 was found to increase its abundance in the cytosol with oxidative stress. An increase of DHX9 was detected in the ribosomes from cells treated with H2O2, most significantly with 100 μM H2O2, and at 60 min. Ribosomal fractionation revealed an increase of DHX9 protein at 43/48S and 80S fractions in H2O2-treated cells. H2O2 treatment caused an RNA-dependent increase of DHX9 interaction with eIF3η. The binding of DHX9 to Nrf2 5′UTR was enhanced by H2O2-treated cells or by deducting the length of Nrf2 5′UTR. RNase digestion enhanced DHX9 association with the ribosomes. Our data have revealed a novel mechanism of de novo Nrf2 protein translation under oxidative stress involving DHX9 binding to Nrf2 5′UTR, perhaps via removal of a negative RNA element, to recruit 43S preinitiation complex for translation initiation.

•Little is known about how certain proteins can be translated under stress conditions.•DHX9 was discovered as a binding protein for Nrf2 5’UTR under oxidative stress.•DHX9 interacts with 43S pre-initiation complex for Nrf2 protein translation.•RNA structural remodeling participates in translation regulation under stress conditions.

Little is known about how certain proteins can be translated under stress conditions.

DHX9 was discovered as a binding protein for Nrf2 5’UTR under oxidative stress.

DHX9 interacts with 43S pre-initiation complex for Nrf2 protein translation.

RNA structural remodeling participates in translation regulation under stress conditions.

Little is known for how proteins are translated under stress conditions. LC-MS/MS–based proteomics led to the discovery of DHX9 as a binding protein for Nrf2 mRNA under oxidative stress. DHX9 interacts with Nrf2 5′UTR and 43S preinitiation complex for translation initiation of Nrf2 protein. As an RNA helicase, DHX9 may remove inhibitory elements to allow eIFs and translation machinery to carry out translation of Nrf2 protein under oxidative stress conditions.