Sodium proton exchanger NHE9 pHine-tunes exosome production by impairing Rab7 activity

Cell-to-cell communication is mediated by vesicles ranging from 30 to 150 nm, known as exosomes. These exosomes shuttle bioactive molecules such as proteins, lipids, and nucleic acids, thus playing crucial roles in both health and disease mechanisms. Exosomes form within the endocytic pathway through the process of inward budding of the endosomal membrane, facilitated by the progressive acidification of the endosomal lumen. Although endosomal pH is known to be critical for exosome production, the precise molecular mechanisms involved remain poorly defined. Maintaining optimal endosomal pH involves meticulous coordination between proton pumping and leakage mechanisms. The sodium-proton exchanger NHE9, located on the endosomal membrane, modulates endosomal pH by transporting protons out of the endosomes in exchange for sodium or potassium ions. Here, we use genetic engineering, biochemistry, and advanced microscopy to demonstrate that the sodium-proton exchanger NHE9 significantly affects exosome production by regulating endosomal pH. NHE9-mediated endosomal alkalization impairs Rab7 activation, thereby disrupting the delivery of multivesicular endosomes to lysosomes. Moreover, luminal alkalization promotes the recruitment of Rab27b. This enhances the targeting of multivesicular endosomes to the cell periphery, their fusion with the plasma membrane, and subsequent exosome secretion. Our findings reveal the detailed molecular mechanisms through which endosomal pH regulates exosome production. Additionally, we identify NHE9 as a potential target for therapeutic strategies aimed at controlling exosome dynamics.