A Dominant Heterozygous Mutation in COG4 Causes Saul–Wilson Syndrome, a Primordial Dwarfism, and Disrupts Zebrafish Development via Wnt Signaling

Abstract

Saul–Wilson syndrome (SWS) is a rare, skeletal dysplasia with progeroid appearance and primordial dwarfism. It is caused by a heterozygous, dominant variant (p.G516R) in COG4, a subunit of the conserved oligomeric Golgi (COG) complex involved in intracellular vesicular transport. Our previous work has shown the intracellular disturbances caused by this mutation; however, the pathological mechanism of SWS needs further investigation. We sought to understand the molecular mechanism of specific aspects of the SWS phenotype by analyzing SWS-derived fibroblasts and zebrafish embryos expressing this dominant variant. SWS fibroblasts accumulate glypicans, a group of heparan sulfate proteoglycans (HSPGs) critical for growth and bone development through multiple signaling pathways. Consistently, we find that glypicans are increased in zebrafish embryos expressing the COG4p.G516R variant. These animals show phenotypes consistent with convergent extension (CE) defects during gastrulation, shortened body length, and malformed jaw cartilage chondrocyte intercalation at larval stages. Since non-canonical Wnt signaling was shown in zebrafish to be related to the regulation of these processes by glypican 4, we assessed wnt levels and found a selective increase of wnt4 transcripts in the presence of COG4p.G516R. Moreover, overexpression of wnt4 mRNA phenocopies these developmental defects. LGK974, an inhibitor of Wnt signaling, corrects the shortened body length at low concentrations but amplifies it at slightly higher concentrations. WNT4 and the non-canonical Wnt signaling component phospho-JNK are also elevated in cultured SWS-derived fibroblasts. Similar results from SWS cell lines and zebrafish point to altered non-canonical Wnt signaling as one possible mechanism underlying SWS pathology.

COG4 mutation in Saul-Wilson syndrome selectively affects secretion of proteins involved in chondrogenesis in chondrocyte-like cells

“…Interestingly, we also found exclusively elevated components in the medium of COG4 p.G516R cells, including WNT signaling receptor FZD2, glypican 1 (GPC1), and LTBP4. These results reinforce the critical roles of WNT signaling pathway in chondrogenic hypertrophy and SWS pathogenesis ( Studer et al, 2012 ; Xia et al, 2021 ). We are aware that FZD2 and GPC1 are membrane proteins instead of secreted or ECM proteins.…”

“…We are aware that FZD2 and GPC1 are membrane proteins instead of secreted or ECM proteins. The possible reason of the detection of GPC1 could be proteoglycan shedding which is considered a powerful post-translational modification ( Niehrs, 2012 ; Xia et al, 2021 ). Also, the detection of some cytoplasmic proteins in the chondrocyte secretome using proteomic techniques has been reported previously and one of the potential explanations is considered as secretion of ECM-derived vesicles by chondrocytes.…”

“…Zebrafish models for both COG4-CDG and COG4-SWS have been reported and they all display small body length, abnormal pectoral fins, and abnormal chondrocyte stacking ( Ferreira et al, 2018 ; Clement et al, 2019 ; Xia et al, 2021 ) . But COG4-null zebrafish exhibit more severely impaired proteoglycan synthesis based on reduced Alcian blue staining of the jaw compared to COG4 p.G516R zebrafish.…”

“…Our previous work also revealed that the COG4 p.G516R variant disturbed WNT4 signaling at the embryonic stage in zebrafish development. These differences likely indicate distinctive mechanisms for the dominant verses recessive COG4 disorders ( Xia et al, 2021 ).…”

“…It is not surprising that the COG4 p.G516R variant could disturb the secretion of multiple proteins. Our previous findings of impaired decorin glycosylation and glypican turnover prompted a more thorough study of protein secretion in a suitable cell line carrying the COG4 p.G516R variant ( Ferreira et al, 2018 ; Xia et al, 2021 ). Sumya et al have shown increased secretion of intracellular protein SIL1 and ERGIC53 in COG4-G516R RPE1 cells compared to WT.…”