In spite of the advances in diagnosis and treatment, HCC patients are still diagnosed at a late stage and possess a poor overall survival rate [20–21]. Therefore, more strategies are needed for HCC early diagnosis and treatment. A growing body of articles has explored numerous molecules year on year, hoping to clear the underlying molecular mechanisms of HCC and contribute to its diagnosis and treatment. Some of them have manifested vital roles in HCC carcinogenesis, such as metadherin [22–24]. However, so far, there has been no valid target protein—especially a protein located in the Golgi complex which could be the target for HCC diagnosis and treatment, making it necessary to be explored. Recent studies have concentrated on proteins of the Golgi apparatus in HCC. Yang et al. (2014) focused on the Golgi apparatus proteomes in HCC tissues and adjacent liver tissues; proteins related to the different BPs were found [25]. It is reported that GP73 acts as a key oncogene in HCC by regulating metastasis. For HCC diagnosis, the accuracy of Golgi protein 73 was higher than alpha-fetoprotein, which is used for clinical diagnosis [26–27]. Liu et al. (2018) suggested that GOLGPH3 acts an oncogene in HCC development and progression by activating the mTOR pathway, which is also a potential target for HCC diagnosis and therapy [28]. Lee et al. (2018) indicated that the Golgi transmembrane protein TEME165 contributes to the progression of HCC [29]. GOLGA8B—a gene that encodes protein golgin-67, which is located in the Golgi complex to maintain its structure—may also serve as a key character in HCC carcinogenesis.
We were the first group to focus on GOLGA8B in HCC. Some previous studies discovered that GOLGA8B manifested different expressions in variant cancers through different methods [11–15]. However, there has been no research so far that has directly studied the relationship between GOLGA8B and cancers. To investigate the accurate function and underlying molecular mechanisms of GOLGA8B, we identified the expression and clinical significance of GOLGA8B, at mRNA level, in HCC by different data combination. IHC was also performed to access the expression of GOLGA8B at the protein level. Consequently, potential functions and pathways were explored.
First, we investigated the role GOLGA8B plays in HCC by an RT-qPCR analysis. Big data which contained RNA-sequencing data and microarray chip data were also utilised for GOLGA8B expression elucidation. There was an higher expression of GOLGA8B in HCC, according to different resources. Based on RNA-sequencing data, GOLGA8B expression was higher in the late pathologic tumour stages (T3–T4) than the early pathologic tumour stages (T1–T2), indicating GOLGA8B might serve as an oncogene in HCC carcinogenesis. However, no correlations were found between GOLGA8B expression and the survival of HCC patients. The expression of GOLGA8B in IHC revealed a similar trend with mRNA expression. GOLGA8B protein exhibited significantly higher expressions in HCC tissues. GOLGA8B protein exhibited higher expressions in late stages (III–IV) than in early stages (I–II), which further elucidated its tumour promotor role in HCC development and progression.
We integrated the expression data obtained from different resources at mRNA and protein level. The results showed SMD = 0.893 (95% CI: 0.279–1.506, P = 0.004). A significantly higher expression in HCC at mRNA and protein level further confirmed the function of GOLGA8B in tumour promotion. Moreover, GOLGA8B might be of clinical value due to the result of sROC with an AUC of 0.79. However, a study with larger samples is still necessary for GOLGA8B expression and clinical value elucidation.
Enrichment analyses also reveal the vital functions and pathways of GOLGA8B in HCC. Within all the biological pathways, the spliceosome pathway was the most significantly enriched term. The spliceosome includes more than 150 different proteins; it yields mature mRNA by removing introns and joining extrons of pre-mRNA, which is responsible for gene expression [30–31]. The spliceosome pathway could participate in the carcinogenesis, cancer development and chemoresistance [32]. By the regulation of mRNA and RNA splicing, phosphorylation of spliceosome proteins may also take effect in the metastasis of HCC [33]. Besides, the spliceosome pathway also takes part in the tumour progression from cirrhosis to HCC [34]. Many genes associated with spliceosome, such as the MYC genes, manifested different expressions in HCC samples compared to the corresponding adjacent healthy liver samples. A number of studies have found that spliceosome could be a target for many anticancer drugs [35].
In addition, classic tumour suppressor phosphatase and tensin homologue could affect proteins on the Golgi apparatus to play a role in tumour suppression by interacting with spliceosome [36]. However, no relevant study related to GOLGA8B and spliceosome was found. In this study, we hypothesised GOLGA8B could be a vital element in the spliceosome pathway, but the potential molecular mechanism of GOLGA8B in HCC needs further confirmation. In addition, the MAPK pathway, which consists of extracellular signal-regulated kinase, functions as regulating fundamental cellular process, which includes survival, proliferation, progression and migration [37]. As reported, in 50–100% of HCC cases, the MAPK pathway is activated and is related to poor prognosis [38]. Carbohydrate-responsive element-binding protein, which acts an important role in lipid and glucose metabolism in liver, takes effect in the development of HCC by regulating the MAPK pathway [39].
Factors such as anticancer drugs act on HCC regulation through triggering the MAPK pathway. This indicates the MAPK pathway might be a potential therapeutic target [40–41]. The MAPK pathway also participates in liver-disease progression—initiated from inflammation, followed by fibrosis, cirrhosis and HCC [42]. A number of studies have elucidated that MAPK pathway plays a significant role in HCC. In this study, we assumed that GOLGA8B is a vital factor in the MAPK pathway. However, no studies related to GOLGA8B and the MAPK pathway could be found; more experiments are necessary to verify this hypothesis.
Hub genes SF3B1, HNRNPA2B1, HNRNPA1 and SRRM2 were selected for further analyses according to degrees and interactions. SF3B1 was overexpressed in HCC tissues and was identified to be a target antigen of HCC-associated antibodies. Also, SF3B1 was significantly mutated through DNA sequencing and mutation analyses [43–44]. Both HNRNPA2B2 and HNRNPA1 belong to the hnRNP A/B family, which is a subset of hnRNP proteins [45]. HNRNPA2B1 was overexpressed in HCC tissues. The expression of HNRNPA2B1 was significantly correlated with tumour differentiation, microvascular invasion and survival rate. As a splicing factor, HNRNPA2B1 participates in cancer development by different pathways—including the MAPK pathway and NF-κB pathway, which provides an interplay between HNRNPA2B1 and GOLGA8B. Furthermore, HNRNPA2B1 interacts with factors like long noncoding RNA and human telomerase reverse transcriptase to serve as an important marker in HCC [45–46]. Similarly, HNRNPA1 expression was upregulated in HCC cell lines and tissues. Through the regulation of CD44v6, overexpression of HNRNPA1 promotes HCC invasion; this indicates shorter overall survival and a higher tumour recurrence rate [47].
SRRM2 is a factor of spliceosome. The mutation in SRRM2 predisposes people to thyroid carcinoma by affecting the alternative splicing of downstream target genes [48]. However, studies related to SRRM2 and HCC have not been found. These four hub genes were all overexpressed in HCC based on TCGA data and showed an inverse trend with GOLGA8B expression. The expression of GOLGA8B and hub genes in HCC informed us that GOLGA8B may target these hub genes to hinder their expression through different mechanisms or signalling pathways. Further explorations are necessary to elucidate the relationship between GOLGA8B and hub genes in HCC.
miRNAs are endogenous, non-coding small RNAs which participate in posttranscriptional regulation by targeting the 3’-untranslated region (UTR) of mRNAs, causing degradation or translation suppressing of mRNAs, and act as suppressors or oncogenes in tumorigenesis [49–50]. In this study, miRNAs which may influence GOLGA8B expression were selected only when they were predicted by six algorithms. The expression of those selected miRNAs were obtained for further analysis. miR-203a, miR-139-5p, miR-144-3p, miR-369-3p and miR-374b-5p were significantly decreased in HCC tissues. It is reported that miR-203a is downregulated in HCC and is associated with the prognosis of HCC patients[51]. Wang et al. (2018) found that miR-203a could suppress the metastasis, migration, invasion and angiogenesis of HCC cells[52]. miR-139-5p exhibited lower expressions in HCC and HCC cell lines. miR-139-5p also plays a role as a tumour suppressor via inhibiting migration, invasion and growth of HCC cells[53-54]. Wu et al. (2017) found that miR-144-3p could suppress the migration and growth of HCC cells[55]. Moreover, the downregulated miR-144-3p was correlated to poor disease-free survival. As for miR-369-3p and miR-374b-5p, they were dysregulated in various types of cancers, but no study has been conducted to explore their functions and mechanisms in HCC[56-60]. These five miRNAs may target GOLGA8B to function as tumour suppressors. However, in vivo and in vitro experiments are necessary for further elucidation.