GP73 N-glycosylation at Asn144 reduces hepatocellular carcinoma cell motility and invasiveness.

Kai Jiang,Qinle Zhang,Wei Li,Shu Zhang,Guoquan Yan,Kun Guo,Yinkun Liu

doi:10.18632/oncotarget.8120

Abstract

Golgi Protein 73 (GP73) is a potential liver disease glycobiomarker warranting comprehensive analyses of its glycan structure and glycosylation function. In this study, we used mass spectrometry to identify glycosylation sites and the glycan structure, high-throughput lectin microarray to provide rapid and sensitive profiling of glycoconjugates, and site-directed mutagenesis to clarify the impact of glycans on target glycoproteins in vivo. We identified three GP73 N-glycosylation sites: Asn109, Asn144 and Asn398. We found five glycoforms on Asn144, including biantennary, triantennary and fucosylated glycans. Removal of N-glycans at Asn144 enhanced the motility and invasiveness of hepatocellular carcinoma cells, possibly due to inhibition of cell adhesion related to the changes of cell membrane glycosylation. This study increases our understanding of the functional relevance of GP73 glycosylation and suggests that Asn144-deleted GP73 can influence the progression and metastasis of hepatocellular carcinoma.

Highlights

Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer mortality globally [1, 2] with a poor 5-year survival rate due to tumor recurrence and metastasis [3]
Norton et al used conventional lectin affinity chromatography and mass spectrometry to identify glycosylation sites and glycan structure of Golgi Protein 73 (GP73) secreted from cultured hepatoma cells [10]
Two N-glycosylation sites Asn109 and Asn144 were identified by liquid chromatography (LC)-mass spectrometry (MS)/MS

Summary

Introduction

Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer mortality globally [1, 2] with a poor 5-year survival rate due to tumor recurrence and metastasis [3]. Elucidation of the detailed mechanism of HCC cell growth and metastasis is crucial to improve HCC therapeutic intervention. Combining differential lectinbased glycoprotein capture with mass spectrometry (MS) analysis, Drake et al discovered that the serum level of fucosylated GP73 varied with HCC disease state [8]. GP73 was linked to HCC through precise large scale identification of core-fucosylated glycoproteins with low- and high-normalized collision energy [9]. Norton et al used conventional lectin affinity chromatography and mass spectrometry to identify glycosylation sites and glycan structure of GP73 secreted from cultured hepatoma cells [10]. Comprehensive analysis of the GP73 glycan structure and glycosylation function could provide additional insight into its role in HCC progression and provide a new avenue for therapeutic intervention

Methods

Results

Conclusion