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The Zα domain of ADARp150 is critical for proper Z-RNA substrate binding and is a key factor in the type-I interferon response pathway. Two point-mutations in this domain (N173S and P193A), which cause neurodegenerative disorders, are linked to decreased A-to-I editing in disease models. To understand this phenomenon at the molecular level, we biophysically and structurally characterized these two mutated domains, revealing that they bind Z-RNA with a decreased affinity. Less efficient binding to Z-RNA can be explained by structural changes in beta-wing, part of the Z-RNA-protein interface, and alteration of conformational dynamics of the proteins.

Running head: Structures of two disease causing Zα Domains of ADAR1

Section snippetsMain

A well-functioning immune system can distinguish non-self from self RNA. This task is in part carried out by the adenosine deaminase acting on RNA (ADAR1), which catalyzes the conversion of some adenosines in self RNA to inosines1, 2, 3. In humans, this ‘A-to-I’ editing is augmented upon infection, primarily through the interferon-induced longer isoform of ADAR1 (p150)1, 2. p150 contains a ∼65-amino acid long Z-RNA binding domain (Zα) at its N-terminus (Fig. 1A). Zα enhances substrate

Methods

Expression of unlabeled human ADAR1p150’s Zα domain: The Zα domain of hADAR1p150 (residues 140-202) (UniProt ID: P55265) cloned in the pET28a(+) plasmid (N-terminal 6x His-tag and thrombin cleavage site between His-tag and the Zα sequence) was a gift from Drs. Peter Dröge and Alekos Athanasiadis. The ZαN173S and ZαP193A mutants were synthesized and cloned into pET28a(+) vectors by Genscript and confirmed through sequencing. Different Zα constructs were expressed and purified similarly as

CRediT authorship contribution statement

Conner J. Langeberg: Conceptualization, Investigation. Parker J. Nichols: Conceptualization, Investigation. Morkos A. Henen: Conceptualization, Investigation, Supervision. Quentin Vicens: Conceptualization, Supervision, Funding acquisition. Beat Vögeli: Conceptualization, Supervision, Funding acquisition.

Acknowledgements

The authors thank Jeff Kieft for support; current and former Kieft Lab and Vögeli Lab members for thoughtful discussions and technical assistance; and the Biophysics core and NMR facilities at the University of Colorado, Anschutz Medical Campus. This research is funded by NSF grant 1917254 for Infrastructure Innovation for Biological Research and a start-up package from the University of Colorado to B.V., University of Colorado Cancer Center Support Grant P30 CA046934, NIH grant S10OD025020 for

References (37)M. Koeris et al.Modulation of ADAR1 editing activity by Z-RNA in vitro

Nucleic Acids Res.

(2005)

G.I. RiceMutations in ADAR1 cause Aicardi-Goutières syndrome associated with a type i interferon signature

Nat. Genet.

(2012)

J.H. LivingstonA type i interferon signature identifies bilateral striatal necrosis due to mutations in ADAR1

J. Med. Genet.

(2014)

A. HerbertMendelian disease caused by variants affecting recognition of Z-DNA and Z-RNA by the Zα domain of the double-stranded RNA editing enzyme ADAR

Eur. J. Hum. Genet.

(2020)

J.E. Heraud-Farlow et al.What do editors do? Understanding the physiological functions of A-to-I RNA editing by adenosine deaminase acting on RNAs: In vivo functions of A-to-I RNA editing

Open Biol.

(2020)

M.S. AnielloBilateral striatal necrosis, dystonia and multiple mitochondrial DNA deletions: Case study and effect of deep brain stimulation

Mov. Disord.

(2008)

S. DaouA phenolic small molecule inhibitor of RNase L prevents cell death from ADAR1 deficiency

Proc. Natl. Acad. Sci. U. S. A.

(2020)

T. Schwartz et al.Crystal structure of the Zα domain of the human editing enzyme ADAR1 bound to left-handed Z-DNA

Science (80-.)

(1999)

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