Elsevier

Bone

Volume 133, April 2020, 115265
Bone

Full Length Article
A mutation in p62 protein (p. R321C), associated to Paget's disease of bone, causes a blockade of autophagy and an activation of NF-kB pathway

https://doi.org/10.1016/j.bone.2020.115265Get rights and content

Highlights

  • First time that p62 p. R321C mutation is associated to Paget's disease of bone.

  • p62 p.R321C mutation induces blockage of autophagy.

  • p62 p.R321C mutation induces cell proliferation through NF-kB pathway.

Abstract

Paget's disease of bone (PDB) is a bone disorder characterized by an increase in bone turnover in a disorganized way with a large increase in bone resorption followed by bone formation. The most important known genetic factor predisposing to PDB is mutation in Sequestosome1 (SQSTM1) gene. We have studied the prevalence of SQSTM1 mutations and examined genotype-phenotype correlations in a Spanish cohort of PDB patients. Also, we have characterized three PDB patients that carry the c.961C>T SQSTM1 gene mutation that it is localized in exon 6 of SQSTM1 gene and it causes the p. R321C mutation. This mutation has been reported in patients with amyotrophic lateral sclerosis and frontotemporal dementia but in our knowledge this is the first time that p62 p. R321C mutation is associated to PDB. We show that p62 p.R321C mutation could induce blockage of autophagy and cell proliferation through NF-kB pathway. These results reinforce the hypothesis of autophagy involvement in Paget's disease of bone.

Introduction

Paget's disease of bone (PDB) (OMIM: 167250) is the second most common metabolic bone disease after osteoporosis [1]. It is characterized by an increase in bone turnover in a disorganized way with a large increase in bone resorption followed by bone formation. The result is an irregular and anarchic bone with altered mechanical properties. Some patients are asymptomatic, while others have pain, degenerative arthropathy, fractures, bone deformities, deafness or other syndromes of nerve compression [2,3].

PDB is a multifactorial disease in which genetic and environmental factors are involved. The main alteration resides in PDB osteoclasts that increase in number, size and activity [2,3]. The PDB osteoclasts present cytoplasmic structures that have been reported as viral nucleocapsids, however, more recently it has been proposed that these inclusions could be aggregated proteins, resulting from alterations in the autophagy process [[4], [5], [6]].

The strong tendency to familial aggregation (15–40%) supports a genetic susceptibility to PDB. To date, the most important known genetic factor predisposing to PDB is mutation in Sequestosome1 (SQSTM1) gene [[7], [8], [9]]. SQSTM1 gene encodes the p62 protein that has 440 aminoacids (62 kDa) and contains six different domains: PB1, ZZ, TRAF6, PEST, LIR and UBA domain. P62 protein plays an important role as a cellular signalling crossroad in osteoclastogenesis, cellular apoptosis and autophagy among others [[10], [11], [12], [13]].

We have studied the prevalence of SQSTM1 mutations and examined genotype-phenotype correlations in a Spanish cohort of patients with PDB. Moreover, bioinformatics and functional analyses were performed to determine the functional consequences of p62 p.R321C mutation. In our knowledge, this is the first time that p62 p.R321C mutation is associated to PDB. This mutation has been reported also by Fecto et al in patients with Amyotrophic Lateral Sclerosis (ALS) [14] and later other groups reported new cases in ALS as well as in patients with frontotemporal dementia (FTLD) [[15], [16], [17], [18]].

Section snippets

Subjects

We have studied 310 patients suffering PDB: 257 patients without family history (termed sporadic PDB patients) and 53 patients from 32 different PDB families. Patients were recruited in the Metabolic Bone Unit at the University Hospital of Salamanca between January 1990 and February 2015. Clinical and analytical variables at diagnosis such as gender, age, family history, number of affected bones, Renier's index, presence of complications (fracture, coxopathy and cranial nerve involvement) and

Mutational screening of the SQSTM1 gene

The clinical variables of the PDB patients included in our cohort are summarized in Table 2. Mutations in the SQSTM1 gene were identified in 10 sporadic PDB patients (3,89%) and in 15 familial PDB patients (28,3%). 8 sporadic PDB patients carried of the p.P392L mutation and 2 of them carried the p.R321C mutation. 12 patients with familial PDB carried the p.P392L mutation; 1 patient had the p.M404T mutation; 1 patient was a carrier of the p.A426V mutation, and 1 patient had the p.R321C mutation.

Discussion

PDB is a bone disorder characterized by an irregular and anarchic structure of the bone that alter their mechanical properties [2,3]. Currently, the most frequent gene mutated in the disease is SQSTM1, that codes for p62 protein [[7], [8], [9]]. P62 protein plays an important role in cellular signal crossroads related with osteoclastogenesis [[10], [11], [12], [13]]. It has been reported that 20–40% of patients with a family history of PDB and 5–10% of sporadic patients are carriers of SQSTM1

CRediT authorship contribution statement

Ricardo Usategui-Martín: Conceptualization, Investigation, Formal analysis, Writing - original draft, Writing - review & editing. Nerea Gestoso-Uzal: Investigation, Formal analysis, Writing - review & editing. Ismael Calero-Paniagua: Investigation, Formal analysis, Writing - review & editing. José María De Pereda: Investigation, Formal analysis, Writing - review & editing. Javier del Pino-Montes: Conceptualization, Investigation, Formal analysis, Writing - review & editing. Rogelio

Acknowledgements

We are grateful to patients with Paget's disease and their families for their participation in this study. Furthermore, we thank Ms. Nieves Mateos for technical help. This work was supported by grants from Instituto de Salud Carlos III (Ministry of Economy and Competitiveness) (ISC IIII-FEDER: PI10/00219 and PI13/01741).

Declaration of competing interest

The authors have no potential conflict of interest.

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