Impaired oxidative stress response characterizes HUWE1-promoted X-linked intellectual disability

Matthias Bosshard,Vuk Palibrk,Matthias Altmeyer,Guy Froyen,Paul Hoff Backe,Cristina Gattiker,Enni Markkanen,Magnar Bjørås,F Lucy Raymond,Stefania Pellegrino,Barbara Van Loon,Grigory L Dianov,Rossana Aprigliano

doi:10.1038/s41598-017-15380-y

Abstract

Mutations in the HECT, UBA and WWE domain-containing 1 (HUWE1) E3 ubiquitin ligase cause neurodevelopmental disorder X-linked intellectual disability (XLID). HUWE1 regulates essential processes such as genome integrity maintenance. Alterations in the genome integrity and accumulation of mutations have been tightly associated with the onset of neurodevelopmental disorders. Though HUWE1 mutations are clearly implicated in XLID and HUWE1 regulatory functions well explored, currently much is unknown about the molecular basis of HUWE1-promoted XLID. Here we showed that the HUWE1 expression is altered and mutation frequency increased in three different XLID individual (HUWE1 p.R2981H, p.R4187C and HUWE1 duplication) cell lines. The effect was most prominent in HUWE1 p.R4187C XLID cells and was accompanied with decreased DNA repair capacity and hypersensitivity to oxidative stress. Analysis of HUWE1 substrates revealed XLID-specific down-regulation of oxidative stress response DNA polymerase (Pol) λ caused by hyperactive HUWE1 p.R4187C. The subsequent restoration of Polλ levels counteracted the oxidative hypersensitivity. The observed alterations in the genome integrity maintenance may be particularly relevant in the cortical progenitor zones of human brain, as suggested by HUWE1 immunofluorescence analysis of cerebral organoids. These results provide evidence that impairments of the fundamental cellular processes, like genome integrity maintenance, characterize HUWE1-promoted XLID.

Highlights

Mutations in the HECT, UBA and WWE domain-containing 1 (HUWE1) E3 ubiquitin ligase cause neurodevelopmental disorder X-linked intellectual disability (XLID)
While HUWE1 levels were significantly lower in XLID cells with HUWE1 p.R4187C, they were higher in the cells with HUWE1 p.R2981H or duplicated HUWE1, when compared to the healthy individual cells (Fig. 1a,b)
Since alterations in the HUWE1 levels result in increased genome instability[14], and accumulation of genomic mutations has been associated with the onset of neurodevelopmental disorders, we tested if the observed alterations in the HUWE1 levels affect the mutation frequency in XLID cells by using the PigA-mutation frequency assay[26]

Summary

Introduction

Mutations in the HECT, UBA and WWE domain-containing 1 (HUWE1) E3 ubiquitin ligase cause neurodevelopmental disorder X-linked intellectual disability (XLID). The observed alterations in the genome integrity maintenance may be relevant in the cortical progenitor zones of human brain, as suggested by HUWE1 immunofluorescence analysis of cerebral organoids. These results provide evidence that impairments of the fundamental cellular processes, like genome integrity maintenance, characterize HUWE1-promoted XLID. We showed that the levels of mutated HUWE1 p.G4310R are significantly lower in XLID individual cells when compared to cells from healthy individual[3] It remains unknown, if other XLID-associated mutations result in changed HUWE1 levels, as well as what is the impact of potentially altered expression and function on the HUWE1-regulated processes, such as genome integrity maintenance. Tough HUWE1 mutations are clearly implicated in XLID and HUWE1 regulatory functions well explored, much is unknown about the cellular processes that are altered by XLID-causing HUWE1 and that contribute to the onset of the disease

Methods

Results

Conclusion