Molecular and Functional Characterization of Choline Transporter-Like Proteins in Esophageal Cancer Cells and Potential Therapeutic Targets

Abstract

In this study, we examined the molecular and functional characterization of choline uptake in the human esophageal cancer cells. In addition, we examined the influence of various drugs on the transport of [3H]choline, and explored the possible correlation between the inhibition of choline uptake and apoptotic cell death. We found that both choline transporter-like protein 1 (CTL1) and CTL2 mRNAs and proteins were highly expressed in esophageal cancer cell lines (KYSE series). CTL1 and CTL2 were located in the plasma membrane and mitochondria, respectively. Choline uptake was saturable and mediated by a single transport system, which is both Na+-independent and pH-dependent. Choline uptake and cell viability were inhibited by various cationic drugs. Furthermore, a correlation analysis of the potencies of 47 drugs for the inhibition of choline uptake and cell viability showed a strong correlation. Choline uptake inhibitors and choline deficiency each inhibited cell viability and increased caspase-3/7 activity. We conclude that extracellular choline is mainly transported via a CTL1. The functional inhibition of CTL1 by cationic drugs could promote apoptotic cell death. Furthermore, CTL2 may be involved in choline uptake in mitochondria, which is the rate-limiting step in S-adenosylmethionine (SAM) synthesis and DNA methylation. Identification of this CTL1- and CTL2-mediated choline transport system provides a potential new target for esophageal cancer therapy.

Choline transport for phospholipid synthesis: An emerging role of choline transporter-like protein 1

“…Similar to the high-affinity choline transporter CHT1, CTL1 is selectively inhibited by the choline analogue hemicholinium-3 (CH-3), yet with a much lower sensitivity. In recent years, further choline uptake studies confirmed the expression and choline transporter function of CTL1/SLC44A1 in human lung alveolar cells, 28 mouse bone marrow-derived macrophages, 42 rat renal tubule epithelial cells, 30 human keratinocytes, 31 human brain microvascular endothelial cells, 14 rheumatoid arthritis synovial fibroblasts, 40 mouse muscle and liver cell lines, 32,33 and numerous cancer cell lines 26,28,[34][35][36][37][38][39]41 as summarized in Table 1.…”

“…In choline-deficient skin fibroblast from the postural orthostatic tachycardia syndrome (POTS) patient mentioned above, CTL1/SLC44A1 protein is reduced, and at the same time, oxygen consumption, mitochondrial potential, and glycolytic activity are diminished and mitochondrial function therefore impaired. 43 Recent studies could also localize CTL2/SLC44A2 protein to the mitochondria of human esophageal cells, 41 renal tubule epithelial cells, 30 human tongue carcinoma cells, 39 and human brain microvascular epithelial cells. 14 The purpose of a widely expressed choline transporter in the mitochondrial membrane is not clear yet, but it is likely that there is a link between CTL1/SLC44A1-mediated choline transport and mitochondrial phospholipid metabolism.…”

“…Brain, breast, prostate, and colon cancer cells have been characterized by elevated choline and phosphocholine levels and abnormal choline metabolism, which correlates with malignancy of the cancer. In recent years, CTL/SLC44A protein overexpression was detected in various cancer cell lines 25,39,41,50 , and the proteins are involved in choline transport in these cells. 15 For example, expression of CTL1/SLC44A1, CTL2/SLC44A2, and CTL4/SLC44A4 messenger RNA is higher in cancerous MCF7 breast epithelial cells than in the MCF10A non-cancerous comparative cell line.…”

“…51 In human esophageal cancer cells, both CTL1/SLC44A1 and CTL2/SLC44A2 are expressed and apparently mediate an Na þ -independent, HC3-inhibitable choline transport in these cells. 41 The increased expression of the choline transporter in cancer cells has been shown to be accompanied by a high expression of the Na þ /H þ exchanger protein to elevate Hþ levels inside the cell and Naþ outside. 26,37 The acidification of the extracellular milieu by inhibition of the exchanger was shown to block choline transport in human colon carcinoma cells 34 and small cell lung carcinoma cells.…”

Protein kinase�C promotes choline transporter‑like protein�1 function via improved cell surface expression in immortalized human hepatic cells

“…These results do not support the theory that ocTs play a predominant role in choline transportation in Fa2n-4 cells. To date, it has been suggested that a cTl1-mediated choline uptake system functions in primary human tumor cell cultures, such as colon carcinoma cells (19), keratinocytes (20), esophageal cancer cells (21), small cell lung carcinoma cells (22), tongue cancer cells (13), and trophoblastic cells (23). likewise, cTl1 is ubiquitously expressed in mammalian tissues, suggesting an important role in choline transportation.…”