In previous studies, the functional studies of GAP43 were focused on the neuronal development, and the protein was considered to play crucial roles in effective regenerative responses, and learning and memory processes of the nervous system[20–22]. GAP43 is highly expressed in nervous system, and also biasedly expressed in indigestive tissues, including duodenum, small intestine, colon and appendix. Recent study has shown that GAP43 serves as a specific biomarker of relapsed or refractory neuroblastoma[23, 24]. However, the relationship between GAP43 and CRC development has been uncovered. In this study, we confirmed that GAP43 is expressed in human colon tissues, but is downregulated in CRC cells and tissues due to the methylation of its promoter region.
Further transcriptome analysis indicated multiple potential biological functions of GAP43 in CRC. The most significant different expression gene is upregulated FILIP1L, downregulated KRT6A and S100A14. While FILIP1L is a WNT pathway inhibitor[12], and the reduction of FILIP1L in ovarian cancer is associated with poor survival, progression and chemoresistance[25]. Previous studies showed that KRT6A and S100A14 were involved in cell migration and cell proliferation[13, 14, 17]. These results remind us that the poor prognosis of reduced GAP43 in CRC may due to its transcriptional alterations and the regulation of some potential signaling pathways, such as WNT signaling.
In KEGG_PATHWAY category, we found that the upregulated genes were clustered into 6 subcategories including ATP-binding cassette (ABC) transporters (p = 0.001). ABC transporters constitute a superfamily of membrane proteins that couple the hydrolysis of adenosine triphosphate (ATP) to vectorial transport of substrates across biological membranes[26]. Previous studies have showed that ABC superfamily utilizes the free-energy from ATP hydrolysis to shuttle many different substrates across various biological membranes, and consequently, are involved in both normal and abnormal physiology[27].In this study, we found that with the overexpression of GAP43 in CRC cells, the expression of ABC transporters is also upregulated. However, the expression of bothGAP43 and ABC transporters is down-regulated in CRC tissues and cell lines, which indicates that ABC transporters might be related with the development of CRC.
In addition, we analyzed the transcriptome of GAP43 overexpressed CRC cells through DAVID and IPA. The IPA analysis showed that EIF2 signaling pathway is the most significant psychological phenomena (-log(p-value) = 25.5 and z-score=-4.802) (Fig. 6a), and three important members(eIF1, ATF4, ATF5) of EIF2 signaling pathway were significantly downregulated (Fig. 6b). Incidentally, we also found that GAP43 is differentially expressed, elevated or decreased in gastric cancer. Survival analysis showed that there was no significant change in survival of gastric cancer patients with low or high expression of GAP43 (Fig. S3). However, we found that patients with gastric cancer who were treated with the 5-FU anticancer drug had higher survival rates in patients with high GAP43 expression than those with low expression (Fig. S3). This suggests that GAP43 is likely to be associated with drug resistance to cancer chemotherapy drugs. Considering that the tolerance of chemotherapy drugs is related with ABC transporters, and GAP43 level is associated with the expression of ABC transporters. Therefore, GAP43 might affect the chemotherapy drug resistance in cancer patients through ABC transporter-mediated signaling pathway.