Changes in Corticotrope Gene Expression Upon Increased Expression of Peptidylglycine α-Amidating Monooxygenase

Abstract

Throughout evolution, secretion has played an essential role in the ability of organisms and single cells to survive in the face of a changing environment. Peptidylglycine α-amidating monooxygenase (PAM) is an integral membrane monooxygenase, first identified for its role in the biosynthesis of neuroendocrine peptides released by the regulated secretory pathway. PAM was subsequently identified in Chlamydomonas reinhardtii, a unicellular green alga, where it plays an essential role in constitutive secretion and in ciliogenesis. Reduced expression of C. reinhardtii PAM resulted in significant changes in secretion and ciliogenesis. Hence, a screen was performed for transcripts and proteins whose expression responded to changes in PAM levels in a mammalian corticotrope tumor cell line. The goal was to identify genes not previously known to play a role in secretion. The screen identified transcription factors, peptidyl prolyl isomerases, endosomal/lysosomal proteins, and proteins involved in tissue-specific responses to glucose and amino acid availability that had not previously been recognized as relevant to the secretory pathway. Perhaps reflecting the dependence of PAM on molecular oxygen, many PAM-responsive genes are known to be hypoxia responsive. The data highlight the extent to which the performance of the secretory pathway may be integrated into a wide diversity of signaling pathways.

Identifying roles for peptidergic signaling in mice

“…The generation of a PAM-derived COOH-terminal fragment called sfCD, which enters the nucleus (25), led to studies on the effects of PAM on gene expression. We know from our studies in a doxycycline-inducible corticotrope tumor cell line that a 2-d 100-fold increase in PAM expression (up close to the level in the adult atrium) altered the expression of a few hundred genes (10). The altered atrial gene expression observed here could reflect the absence of PAM itself or a compensatory response to the consequences of lacking PAM expression since conception.…”

“…The x axis is log 2 of the ratio (WT/knockout [KO]); the y axis is −log10 of the DESeq2 P value (up is more significant). Transcripts with blue dots were decreased when PAM was induced in pituitary cells in our previous study (10), while red dots indicate transcripts increased when PAM was induced. Gray dots indicate transcripts expressed at low levels in pituitary cells (FPKM < 3).…”

“…N, no change in pituitary cells; X, opposite response in pituitary cells; 0, not expressed above FPKM = 2 in pituitary cells. *Significant in pituitary cells (10).…”

“…Expression of 10 atrial transcripts (marked with X in Fig. 8 B and C) decreased with loss of atrial PAM and also decreased in pituitary cells when PAM was increased (10), suggesting that these transcripts are not responding directly to the PAM protein but rather to some other compensatory changes or differences between the 2 tissues. Five of 39 atrial transcript are not expressed at significant levels in pituitary cells (marked with 0 in Fig.…”

“…8 B and C). For example, expression of Per1 increased more than 2-fold in Pam Myh6-cKO/cKO mice compared with WT and decreased to 60% when Pam expression was increased in pituitary cells (10), suggesting that the change may be a direct response to altered PAM levels. Expression of 10 atrial transcripts (marked with X in Fig.…”