Glycogene Expression Profiling of Hepatic Cells by RNA-Seq Analysis for Glyco-Biomarker Identification

Abstract

Glycans are primarily generated by "glycogenes," which consist of more than 200 genes for glycosynthesis, including sugar-nucleotide synthases, sugar-nucleotide transporters, and glycosyltransferases. Measuring the expression level of glycogenes is one of the approaches to analyze the glycomes of particular biological and clinical samples. To develop an effective strategy for identifying the glycosylated biomarkers, we performed transcriptome analyses using quantitative real-time polymerase chain reaction (qRT-PCR) arrays and RNA sequencing (RNA-Seq). First, we measured and analyzed the transcriptome from the primary culture of human liver cells and hepatocarcinoma cells using RNA-Seq. This analysis revealed similar but distinctive expression profiles of glycogenes among hepatic cells as indicated by the qRT-PCR arrays, which determined a copy number of 186 glycogenes. Both data sets indicated that altered expression of glycosyltransferases affect the glycosylation of particular glycoproteins, which is consistent with the mass analysis data. Moreover, RNA-Seq analysis can uncover mutations in glycogenes and search differently expressed genes out of more than 50,000 distinct human gene transcripts including candidate biomarkers that were previously reported for hepatocarcinoma cells. Identification of candidate glyco-biomarkers from the expression profile of the glycogenes and proteins from liver cancer tissues available from public database emphasized the possibility that even though the expression level of biomarkers might not be altered, the expression of the glycogenes modifying biomarkers, generating glyco-biomarkers, might be different. Pathway analysis revealed that ∼20% of the glycogenes exhibited different expression levels in normal and cancer cells. Thus, transcriptome analyses using both qRT-PCR array and RNA-Seq in combination with glycome and glycoproteome analyses can be advantageous to identify "glyco-biomarkers" by reinforcing information at the expression levels of both glycogenes and proteins.

Association between the expression of core 3 synthase and survival outcomes of patients with cholangiocarcinoma

“…Frozen tissues were obtained from resected surgical specimens and were stored at -80˚C until use. For comparison between pathological specimens and cancer cell lines, frozen CCA tissue samples from the G8-144 positive cases and HAL8 cells, a lung adenocarcinoma cell line (22), were lysed using the lysis buffer. After centrifugation, the supernatant was collected, and the protein concentration was measured using a micro-BCA kit (Thermo Fisher Scientific, Inc.).…”

LM-GlycomeAtlas Ver. 2.0: An Integrated Visualization for Lectin Microarray-based Mouse Tissue Glycome Mapping Data with Lectin Histochemistry

“…26 LMA-based tissue data can be compared with MSbased tissue glycomic and glycoproteomics data provided by GlycomeAtlas 22 and GlycoProtDB, 27 respectively. In addition, information on the characteristics and expression profiles of glycogenes obtained from the GlycoGene Database (GGDB) 28 and GlycoMaple may facilitate the subsequent analysis of the glycosylation machinery underlying tissue-and region-specific protein modifications observed in tissue glycome maps in LM-GlycomeAtlas. The integrated use of these multiomics databases is expected to facilitate a further understanding of spatial protein glycosylation.…”

Bridging Glycomics and Genomics: New Uses of Functional Genetics in the Study of Cellular Glycosylation

“…This body of work uncovered alterations to GAG biosynthesis in a number of breast cancer subtypes ( Huang et al, 2021c ). Similar approaches have been used to investigate regulation of glycogene expression in CRC ( Wu et al, 2021 ), cervical cancer ( Martinez-Morales et al, 2021 ), hepatic cancer ( Angata et al, 2020 ) and pluripotent stem cells ( Pecori et al, 2021 ). To facilitate such studies, Thambu et al also recently developed anexVis, a visualization tool that similarly facilitates transcriptomic analysis of glycan biosynthesis genes ( Thambu and Balagurunathan 2022 ).…”