SLC4A2 anion exchanger promotes tumour cell malignancy via enhancing net acid efflux across golgi membranes

Elham Khosrowabadi,Tuula Salo,Maija Risteli,Sakari Kellokumpu,Anne Tuomisto,Markus J Mäkinen,Antti Rivinoja

doi:10.1007/s00018-021-03890-y

Abstract

Proper functioning of each secretory and endocytic compartment relies on its unique pH micro-environment that is known to be dictated by the rates of V-ATPase-mediated H+ pumping and its leakage back to the cytoplasm via an elusive “H+ leak” pathway. Here, we show that this proton leak across Golgi membranes is mediated by the AE2a (SLC4A2a)-mediated bicarbonate-chloride exchange, as it is strictly dependent on bicarbonate import (in exchange for chloride export) and the expression level of the Golgi-localized AE2a anion exchanger. In the acidic Golgi lumen, imported bicarbonate anions and protons then facilitate a common buffering reaction that yields carbon dioxide and water before their egress back to the cytoplasm via diffusion or water channels. The flattened morphology of the Golgi cisternae helps this process, as their high surface-volume ratio is optimal for water and gas exchange. Interestingly, this net acid efflux pathway is often upregulated in cancers and established cancer cell lines, and responsible for their markedly elevated Golgi resting pH and attenuated glycosylation potential. Accordingly, AE2 knockdown in SW-48 colorectal cancer cells was able to restore these two phenomena, and at the same time, reverse their invasive and anchorage-independent growth phenotype. These findings suggest a possibility to return malignant cells to a benign state by restoring Golgi resting pH.

Highlights

Altered cellular metabolism, tumour acidosis, and aberrant glycosylation are all hallmarks of cancers and contribute to tumorigenesis and its progression by various means [1,2,3,4] Cancer-associated glycosylation changes most often include increased branching and/or fucosylation of N-linked glycans, synthesis of truncated mucin-type O-glycans, increased sialylation and decreased sulphation of glycosaminoglycans such as heparan sulphate [5]
Because cell-surface glycans regulate a vast number of different cell–cell and cell–matrix interactions [6, 7], their alterations can modulate a variety of basic cellular functions, including inflammatory responses, immune evasion, apoptosis, cell attachment as well as cancer cell dissemination, motility, invasion, and metastasis [8,9,10,11,12]
By using ratiometric and Golgi-targeted pHluorin as the probe [38], we show that the AE2a variant regulates Golgi resting pH by facilitating net acid efflux across Golgi membranes via the well-known chemical buffering reaction that yields carbon dioxide and water from luminal bicarbonate anions and protons

Summary

Introduction

Tumour acidosis, and aberrant glycosylation are all hallmarks of cancers and contribute to tumorigenesis and its progression by various means [1,2,3,4] Cancer-associated glycosylation changes most often include increased branching and/or fucosylation of N-linked glycans, synthesis of truncated mucin-type O-glycans, increased sialylation and decreased sulphation of glycosaminoglycans such as heparan sulphate [5]. ◂Fig. 1 AE2a overexpression in COS-7 cells alters Golgi resting pH and its glycosylation potential. C Quantification of the AE2a-mCherry protein level in wild-type COS-7 cells (control), in mock-transfected cells, and AE2a-mCherry expressing cells. D Ratiometric Golgi pH measurements in wild type, mock-transfected, or AE2a-mCherry (red) expressing COS-7 cells. E Regression analysis between Golgi resting pH and AE2a-mCherry expression level. Single-cell data points (dots) were classified into eight equal classes based on AE2a-mCherry intensity (200–5000 AU units, 600 AU units/class), and plotted against the Golgi resting pH. F Lectin binding differences between AE2a-mCherry overexpressing COS-7 cells and mock-transfected control cells. H A cartoon that illustrates main N- and O-glycan differences between AE2a-mCherry overexpressing cells and mock-transfected controls and inability to form functionally relevant complexes in the Golgi [13, 14]. Previous work from our laboratory suggests that Golgi resting pH in cancer cells is abnormally high [15] and that pH gradient dissipating compounds (chloroquine, ammonium chloride) can induce cancer-associated glycosylation changes in normal cells and lead to mistargeting of apical glycoproteins in epithelial cells [16,17,18]

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Results

Conclusion