pH dependence of melanoma cell migration: protons extruded by NHE1 dominate protons of the bulk solution

Abstract

Migration and morphology of human melanoma cells (MV3) depend on extracellular pH (pHe) and the activity of the Na+/H+ exchanger NHE1. To distinguish effects of NHE1 activity per se from effects of pHe we compared an NHE1-deficient mutant with rescued and wild-type cells. Time lapse video microscopy was used to investigate migratory and morphological effects caused by pHe and NHE1 activity, and a membrane-bound fluorescein conjugate was employed for ratiometric pH measurements at the outer leaflet of the cell membrane. As long as NHE1 remained inactive due to deficiency or inhibition by cariporide (HOE642) neither migration nor morphology was affected by changes in pHe. Under these conditions pH at the outer leaflet of the plasma membrane was uniform all over the cell surface. The typical pH dependence of MV3 cell migration and morphology could be reconstituted by restoring NHE1 activity. At the same time the proton gradient at the outer leaflet of the plasma membrane with the higher proton concentration at the leading edge and the lower one at the cell rear was re-established as well. Hence, NHE1 activity generates a proton gradient at the cell surface accompanied by the cells' ability to respond to changes in pHe (bulk pH). We conclude that NHE1 activity contributes to the generation of a well-defined cell surface pH by creating a proton gradient at the outer leaflet of the plasma membrane that is needed for (i) the development of a variety of morphologies including a distinct polarity and (ii) migration. A missing proton gradient at the cell surface cannot be compensated for by varying pHe.

Cancer metabolism in space and time: Beyond the Warburg effect

“…This re sults from the ab nor mal meta bolic ac tiv ity of can cer cells, in clud ing in creased gly colytic flux and lac tic acid re lease [150]; car bon diox ide pro duc tion by the TCA cy cle and the pen tose phos phate path way (PPP) [151][152][153][154]; over ex pres sion or up reg u lated ac tiv ity of car bonic an hy drases [155,156] and pro ton trans porters [156]; and de creased pro ton clear ance in the tu mor en vi ron ment [157]. Pro ton ex tru sion and ex tra cel lu lar acid i fi ca tion make ad her ent junc tions lose [158], stim u late mi gra tion and cy toskele ton re or ga ni za tion [159][160][161][162][163], and pro mote the ex pres sion [164,165], re lease [166,167] and ac tiv ity [167,168] of pro teases that de grade the ex tra cel lu lar ma trix. Hence, acidic prim ing was shown to be suf fi cient to in crease lung col o niza tion fol low ing sar coma and melanoma tu mor cell in jec tion in the tail vein of mice [163,167,169,170].…”

miR-145-dependent targeting of Junctional Adhesion Molecule A and modulation of fascin expression are associated with reduced breast cancer cell motility and invasiveness

“…Time-lapse and immunofluorescence microscopy Time-lapse microscopy experiments were essentially performed as described previously (Stu¨we et al, 2007). A detailed description of time-lapse microscopy experiments is available in the Supplementary Information.…”

Proton production, regulation and pathophysiological roles in the mammalian brain

“…It is reported that the asymmetrical expression and activation of NHE1 in cells generates a local intracellular pH gradient and thus induces or at least supports the rearrangement of the cytoskeleton by affecting cofilin, an actin-binding protein that increases the recycling of actin monomers [1] . While a great deal of effort has been focused on the involvement of NHE1 in cell migration and polarity [33][34][35][36] , two studies have demonstrated that protons, released via C. elegans intestinal NHE PBO-4 (also known as NHX-7, the ortholog of mammalian NHE1), act on a proton-gated cation channel (comprised of PBO-5 and PBO-6 subunits) located in the adjacent muscle cells to induce contraction during the defecation cycle [5,6] . These studies provided characterized [37] .…”