Ubiquitin-protein ligase Ubr5 cooperates with hedgehog signalling to promote skeletal tissue homeostasis.

Anna Thornburn,Ioanna Ch Georgiou,Malgorzata Kubiak,Rob Van’T Hof,Colin Farquharson,Mark Ditzel,Stuart H Ralston,Elaine Kinsella,Silvia Peluso,Robert E Hill,Michela Grillo,Natalia A Riobo-Del Galdo,Natalie Dora,Gregory S Barsh,Katherine A Staines,David Mellis,Donald M Salter

doi:10.1371/journal.pgen.1009275

Abstract

Mammalian Hedgehog (HH) signalling pathway plays an essential role in tissue homeostasis and its deregulation is linked to rheumatological disorders. UBR5 is the mammalian homologue of the E3 ubiquitin-protein ligase Hyd, a negative regulator of the Hh-pathway in Drosophila. To investigate a possible role of UBR5 in regulation of the musculoskeletal system through modulation of mammalian HH signaling, we created a mouse model for specific loss of Ubr5 function in limb bud mesenchyme. Our findings revealed a role for UBR5 in maintaining cartilage homeostasis and suppressing metaplasia. Ubr5 loss of function resulted in progressive and dramatic articular cartilage degradation, enlarged, abnormally shaped sesamoid bones and extensive heterotopic tissue metaplasia linked to calcification of tendons and ossification of synovium. Genetic suppression of smoothened (Smo), a key mediator of HH signalling, dramatically enhanced the Ubr5 mutant phenotype. Analysis of HH signalling in both mouse and cell model systems revealed that loss of Ubr5 stimulated canonical HH-signalling while also increasing PKA activity. In addition, human osteoarthritic samples revealed similar correlations between UBR5 expression, canonical HH signalling and PKA activity markers. Our studies identified a crucial function for the Ubr5 gene in the maintenance of skeletal tissue homeostasis and an unexpected mode of regulation of the HH signalling pathway.

Highlights

Ubiquitin ligases target proteins for ubiquitination which can modulate protein function by regulating protein degradaton, protein–protein interactions, and protein localization [1,2,3,4], and provide important post-translational mechanisms essential for a variety of cellular processes
Ubiquitin Protein Ligase E3 Component N-Recognin 5 (UBR5) is an E3 ubiquitin ligase and in Drosophila is a regulator of Hedgehog signaling
This work shows that UBR5 interacts with the HH pathway to regulate skeletal homeostasis in and around joints of the legs and identifies targets that may be harnessed for biomedical engineering and clinical applications

Summary

Introduction

Ubiquitin ligases target proteins for ubiquitination which can modulate protein function by regulating protein degradaton, protein–protein interactions, and protein localization [1,2,3,4], and provide important post-translational mechanisms essential for a variety of cellular processes. We previously addressed a possible conserved role for UBR5 in HH-mediated processes in mice [9]. No overt effects were seen in patterning of the developing limb bud in mouse embryogenesis; here, we show that the coordinated action of Ubr with HH signalling is crucial to maintain skeletal tissue homeostasis associated with the appendicular skeleton postnatally and in adult mice. HH signalling regulates cell processes that are critical for skeletal tissue development, growth and homeostasis [10]. Binding to PTCH1 results in derepression of the G protein-coupled receptor, smoothened (SMO), and activation of SMOassociated canonical and non-canonical signalling pathways [11,12,13]. Activation of the SMOassociated canonical pathway results in stimulation of GLI-mediated transcription and expression of crucial target genes [7]. Not yet experimentally addressed, non-canonical signalling may contribute to many of the well-described roles for canonical HH signalling in normal skeletal formation, maturation and maintenance [10,18]

Methods

Results

Discussion

Conclusion