Molecular mechanisms associated with acidification and alkalization along the larval midgut of Musca domestica

To disclose the molecular mechanisms involved in luminal midgut buffering of M. domestica, we used RNA-seq analyses from triplicate samples of seven sections along the midgut to evaluate the expression levels of genes coding for selected manually curated protein sequences. Channels, pumps and transporters were confirmed as being apical by proteomics of purified microvillar membranes. Midgut pH determinations with a microsensor and a pH indicator were carried out in larvae in different diets with or without added compounds to evaluate the role of proteins in buffering. The data suggested that acidification occurs at middle midgut by the action of H⁺ V-ATPase with protons produced by carbonic anhydrase, followed by chloride ions transported by a K⁺Cl^- symporter. K⁺ ions are recovered by an apical K⁺ channel and K⁺ homeostasis maintained by a basolateral Na⁺/K⁺-ATPase. Acidification is also affected by a Na⁺/H⁺ exchanger and a multidrug resistance protein. Posterior midgut alkalization results from the action of a NH₃ transporter and H⁺-coupled peptide transporter, mainly in a diet rich in free peptides. A working model was proposed for the midgut luminal acidification and alkalization, as well as for mucosal protection against acid by a neutralized mucus layer.