The a3 isoform of subunit a of the vacuolar ATPase localizes to the plasma membrane of invasive breast tumor cells and is overexpressed in human breast cancer.

Kristina Cotter,Rachel Liberman,Gehong Sun-Wada,Stephen Naber,Dennis Sgroi,Yoh Wada,Dennis Brown,Michael Forgac,Sylvie Breton

doi:10.18632/oncotarget.10063

Abstract

The vacuolar (H+)-ATPases (V-ATPases) are a family of ATP-driven proton pumps that acidify intracellular compartments and transport protons across the plasma membrane. Previous work has demonstrated that plasma membrane V-ATPases are important for breast cancer invasion in vitro and that the V-ATPase subunit a isoform a3 is upregulated in and critical for MDA-MB231 and MCF10CA1a breast cancer cell invasion. It has been proposed that subunit a3 is present on the plasma membrane of invasive breast cancer cells and is overexpressed in human breast cancer. To test this, we used an a3-specific antibody to assess localization in breast cancer cells. Subunit a3 localizes to the leading edge of migrating breast cancer cells, but not the plasma membrane of normal breast epithelial cells. Furthermore, invasive breast cancer cells express a3 throughout all intracellular compartments tested, including endosomes, the Golgi, and lysosomes. Moreover, subunit a3 knockdown in MB231 breast cancer cells reduces in vitro migration. This reduction is not enhanced upon addition of a V-ATPase inhibitor, suggesting that a3-containing V-ATPases are critical for breast cancer migration. Finally, we have tested a3 expression in human breast cancer tissue and mRNA prepared from normal and cancerous breast tissue. a3 mRNA was upregulated 2.5-47 fold in all breast tumor cDNA samples tested relative to normal tissue, with expression generally correlated to cancer stage. Furthermore, a3 protein expression was increased in invasive breast cancer tissue relative to noninvasive cancer and normal breast tissue. These studies suggest that subunit a3 plays an important role in invasive human breast cancer.

Highlights

Cancer cells are able to survive and proliferate in tumor microenvironments that are more acidic than normal, maintaining a slightly alkaline intracellular pH despite the lower than normal extracellular pH [1]
Despite the evidence suggesting its importance in breast tumor cell invasion discussed above, the localization of the a3 isoform the V-ATPase subunit a in breast cancer cells is not known
Several studies have examined the role of the a3 isoform of the V-ATPase subunit a in breast cancer cell invasion in vitro [22,23], much remains unknown with regard to the localization, function, and expression of a3 in human breast cancer

Summary

Introduction

Cancer cells are able to survive and proliferate in tumor microenvironments that are more acidic than normal, maintaining a slightly alkaline intracellular pH despite the lower than normal extracellular pH [1]. This unique ability affects the metastatic potential of cancer cells. A more alkaline cytosolic pH can enhance cytoskeletal remodeling necessary for cell migration and an acidic microenvironment enhances cancer cell invasion by promoting the activity of pH-dependent proteases [1]. The primary role of the V-ATPase is to transport protons into intracellular compartments or the extracellular space. These include osteoclasts, where V-ATPases are involved in bone resorption, renal intercalated cells, where they function in pH homeostasis by secreting acid from the blood into the urine, and epididymal clear cells, where they are involved in sperm maturation [3,4,5,6,7]

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