Continuous stress-induced dopamine dysregulation augments PAP-I and PAP-II expression in melanotrophs of the pituitary gland.

Under continuous stress (CS) in rats, melanotrophs, the predominant cell-type in the intermediate lobe (IL) of the pituitary, are hyperactivated to secrete α-melanocyte-stimulating hormone and thereafter degenerate. Although these phenomena are drastic, the molecular mechanisms underlying the cellular changes are mostly unknown. In this study, we focused on the pancreatitis-associated protein (PAP) family members of the secretory lectins and characterized their expression in the IL of CS model rats because we had identified two members of this family as up-regulated genes in our previous microarray analysis. RT-PCR and histological studies demonstrated that prominent PAP-I and PAP-II expression was induced in melanotrophs in the early stages of CS, while another family member, PAP-III, was not expressed. We further examined the regulatory mechanisms of PAP-I and PAP-II expression and revealed that both were induced by the decreased dopamine levels in the IL under CS. Because the PAP family members are implicated in cell survival and proliferation, PAP-I and PAP-II secreted from melanotrophs may function to sustain homeostasis of the IL under CS conditions in an autocrine or a paracrine manner.

V-ATPase-mediated granular acidification is regulated by the V-ATPase accessory subunit Ac45 in POMC-producing cells.

The vacuolar (H(+))-ATPase (V-ATPase) is an important proton pump, and multiple critical cell-biological processes depend on the proton gradient provided by the pump. Yet, the mechanism underlying the control of the V-ATPase is still elusive but has been hypothesized to involve an accessory subunit of the pump. Here we studied as a candidate V-ATPase regulator the neuroendocrine V-ATPase accessory subunit Ac45. We transgenically manipulated the expression levels of the Ac45 protein specifically in Xenopus intermediate pituitary melanotrope cells and analyzed in detail the functioning of the transgenic cells. We found in the transgenic melanotrope cells the following: i) significantly increased granular acidification; ii) reduced sensitivity for a V-ATPase-specific inhibitor; iii) enhanced early processing of proopiomelanocortin (POMC) by prohormone convertase PC1; iv) reduced, neutral pH-dependent cleavage of the PC2 chaperone 7B2; v) reduced 7B2-proPC2 dissociation and consequently reduced proPC2 maturation; vi) decreased levels of mature PC2 and consequently reduced late POMC processing. Together, our results show that the V-ATPase accessory subunit Ac45 represents the first regulator of the proton pump and controls V-ATPase-mediated granular acidification that is necessary for efficient prohormone processing.