Poly(A)-specific ribonuclease deficiency impacts telomere biology and causes dyskeratosis congenita
Hemanth Tummala, … , Thomas Vulliamy, Inderjeet Dokal
Hemanth Tummala, … , Thomas Vulliamy, Inderjeet Dokal
Published April 20, 2015
Citation Information: J Clin Invest. 2015;125(5):2151-2160. https://doi.org/10.1172/JCI78963.
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Research Article Aging Genetics Hematology Article has an altmetric score of 23

Poly(A)-specific ribonuclease deficiency impacts telomere biology and causes dyskeratosis congenita

Abstract

Dyskeratosis congenita (DC) and related syndromes are inherited, life-threatening bone marrow (BM) failure disorders, and approximately 40% of cases are currently uncharacterized at the genetic level. Here, using whole exome sequencing (WES), we have identified biallelic mutations in the gene encoding poly(A)-specific ribonuclease (PARN) in 3 families with individuals exhibiting severe DC. PARN is an extensively characterized exonuclease with deadenylation activity that controls mRNA stability in part and therefore regulates expression of a large number of genes. The DC-associated mutations identified affect key domains within the protein, and evaluation of patient cells revealed reduced deadenylation activity. This deadenylation deficiency caused an early DNA damage response in terms of nuclear p53 regulation, cell-cycle arrest, and reduced cell viability upon UV treatment. Individuals with biallelic PARN mutations and PARN-depleted cells exhibited reduced RNA levels for several key genes that are associated with telomere biology, specifically TERC, DKC1, RTEL1, and TERF1. Moreover, PARN-deficient cells also possessed critically short telomeres. Collectively, these results identify a role for PARN in telomere maintenance and demonstrate that it is a disease-causing gene in a subset of patients with severe DC.

Authors

Hemanth Tummala, Amanda Walne, Laura Collopy, Shirleny Cardoso, Josu de la Fuente, Sarah Lawson, James Powell, Nicola Cooper, Alison Foster, Shehla Mohammed, Vincent Plagnol, Thomas Vulliamy, Inderjeet Dokal

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Figure 2

Mutations in PARN cause deadenylation deficiency.

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Mutations in PARN cause deadenylation deficiency. (A and B) Immunocytoch...
(A and B) Immunocytochemistry and Western blotting on EBV-transformed LCLs revealed no apparent differences in localization or expression of PARN between control, father, and case 1. Antibody against TATA binding protein (TBP) was used as a loading control. Immunostaining images show DAPI (blue) or PARN (red) from case 1, father, or control. Scale bar: 8 μm. (C) A gel-based deadenylase assay shows reduced deadenylation activity of an RNA substrate in whole-cell extracts from case 1 LCLs and PARN siRNA–treated (PARNsi-treated) HEK293 cells. The upper arrow (A9) indicates intact RNA substrates containing nine 3′ adenosine residues, the lower arrow (A1) indicates the reaction product containing a single 3′ adenosine residue, and * denotes related deadenylated products. (D and E) Fluorescence-based measurement shows reduced deadenylation kinetics in case 1 LCLs and PARNsi-treated cells. (F–H) PARN topology shows the position of p.Ala383Val missense change in the ND2 domain. (F) In silico analysis of PARN catalytic site (PDB 2A1R) denotes the amino acids from each promoter (ball and stick model; green) and bound RNA poly(A) (black). The mutated alanine 383 (Ala 383) residue is shown in blue. The adjacent aspartic acid 382 (Asp 382) residue has been shown to be involved in RNA poly(A) processivity (19). (G) Ribbon diagram (yellow) showing part of PARN nuclease domain and RNA poly(A) complex (black). (H) The missense change Ala 383 (blue, in G) to valine 383 (Val 383) (red) introduces a side chain (arrow head) in the α-helices (yellow) of PARN nuclease domain.