TMEM165 a new player in proteoglycan synthesis: loss of TMEM165 impairs elongation of chondroitin- and heparan-sulfate glycosaminoglycan chains of proteoglycans and triggers early chondrocyte differentiation and hypertrophy

Sajida Khan,Mohamed Ouzzine,Lydia Barré,François Foulquier,Malak Sbeity

doi:10.1038/s41419-021-04458-1

Abstract

TMEM165 deficiency leads to skeletal disorder characterized by major skeletal dysplasia and pronounced dwarfism. However, the molecular mechanisms involved have not been fully understood. Here, we uncover that TMEM165 deficiency impairs the synthesis of proteoglycans by producing a blockage in the elongation of chondroitin-and heparan-sulfate glycosaminoglycan chains leading to the synthesis of proteoglycans with shorter glycosaminoglycan chains. We demonstrated that the blockage in elongation of glycosaminoglycan chains is not due to defect in the Golgi elongating enzymes but rather to availability of the co-factor Mn2+. Supplementation of cell with Mn2+ rescue the elongation process, confirming a role of TMEM165 in Mn2+ Golgi homeostasis. Additionally, we showed that TMEM165 deficiency functionally impairs TGFβ and BMP signaling pathways in chondrocytes and in fibroblast cells of TMEM165 deficient patients. Finally, we found that loss of TMEM165 impairs chondrogenic differentiation by accelerating the timing of Ihh expression and promoting early chondrocyte maturation and hypertrophy. Collectively, our results indicate that TMEM165 plays an important role in proteoglycan synthesis and underline the critical role of glycosaminoglycan chains structure in the regulation of chondrogenesis. Our data also suggest that Mn2+ supplementation may be a promising therapeutic strategy in the treatment of TMEM165 deficient patients.

Highlights

Glycosylation is one of the most common and important posttranslational modifications of proteins [1, 2]
Here, we generated tmem165-knockout pre-chondrocyte mouse ATDC5 and HEK293 cells and showed that loss of TMEM165 led to strong defects in the synthesis of PGs
We showed a dramatical reduction in the length of HS- and CS-GAG chains, revealing for the first time that elongation of PG-GAG chains is impaired in TMEM165 deficient cells

Summary

Introduction

Glycosylation is one of the most common and important posttranslational modifications of proteins [1, 2]. Genetic defects in protein glycosylation can lead to Congenital Disorders of Glycosylation (CDGs) which is a group of inherited diseases associated with a broad variety of pathological symptoms [3]. The recently identified CDG subtype linked to mutations in TMEM165 (transmembrane protein 165) [4]. Mutation of the yeast ortholog of TMEM165, named Gdt induced sensitivity to high Ca2+external concentrations, suggesting its participation to Ca2+ transport and reduction of the concentration of Ca2+ in the cytosol [3]. TMEM165 gene deficiency was associated with a slight defect in sialylation and galactosylation of N-glycans in TMEM165-deficient patients. The transport activity of TMEM165 in human cells have not been demonstrated yet, several indirect observations suggest that TMEM165 may be involved in maintaining Golgi Ca2+, H+, Mn2+ homeostasis [5]

Methods

Results

Conclusion