Mechanism of Sorting Proopiomelanocortin and Proenkephalin to the Regulated Secretory Pathway of Neuroendocrine Cells

Abstract

Proopiomelanocortin (POMC) and proenkephalin (PE) are synthesized at the endoplasmic reticulum and transported to the trans-Golgi network (TGN) where they are sorted and packaged into dense-core granules of the regulated secretory pathway (RSP). The mechanism of sorting POMC and PE to the RSP in neuroendocrine cells was investigated. Consensus sorting signals comprising two acidic residues and two hydrophobic residues exposed on the surface of N-POMC(1-26) and N-PE(1-32) were identified and shown to be sufficient and necessary for targeting POMC and PE to the RSP in PC12, Neuro2a, and AtT-20 cells. The acidic residues of these sorting signals bind specifically to basic residues on the sorting receptor membrane, carboxypeptidase E (CPE), to effect sorting to the RSP. Analysis of POMC and PE sorting in Neuro2a cells depleted of CPE by CPE antisense RNA, and Cpe(fat/fat) mouse pituitary cells lacking CPE showed missorting of both these molecules to the constitutive pathway in vivo. Thus, POMC and PE are sorted to the RSP at the TGN by a mechanism involving the interaction of a specific sorting signal on these molecules with the sorting receptor, CPE.

Salivary glands as a potential gene transfer target for gene therapeutics of some monogenetic endocrine disorders

“…amino acid sequence dependent; e.g. Kelly 1985, Gerdes & Glombik 1999, Loh et al 2002. The example of pro-insulin mis-sorting both in humans and in transgenic mice due to point mutations in the insulin gene is well described (Chan et al 1987, Carroll et al 1988.…”

60 YEARS OF NEUROENDOCRINOLOGY: TRH, the first hypophysiotropic releasing hormone isolated: control of the pituitary–thyroid axis

“…The protein codified by the rat (r) Trh gene is a precursor (pre-proTRH; 255 amino acids) containing five Gln-His-Pro-Gly sequences flanked by a pair of basic residues and cryptic peptides in between (Lechan et al 1986). As for other neuropeptides (Loh et al 2002), proTRH is processed in the secretory pathway through sequential enzyme activities: convertases, carboxypeptidase, pyroglutamyl cyclase, and peptidylglycine a-hydroxylating monooxygenase (Wu & Jackson 1988, Nillni 2010, Fekete & Lechan 2014.…”

“…The protein codified by the rat (r) Trh gene is a precursor (pre-proTRH; 255 amino acids) containing five Gln-His-Pro-Gly sequences flanked by a pair of basic residues and cryptic peptides in between (Lechan et al 1986). As for other neuropeptides (Loh et al 2002), proTRH is processed in the secretory pathway through sequential enzyme activities: convertases, carboxypeptidase, pyroglutamyl cyclase, and peptidylglycine a-hydroxylating monooxygenase (Wu & Jackson 1988, Nillni 2010, Fekete & Lechan 2014.Antibodies specific for proTRH, together with Trh cDNA, were used in immunocytochemical and in situ hybridization analyses that enabled the final identification of the paraventricular nucleus (PVN) as the hypothalamic nucleus with the highest expression of proTRH precursor (Lechan & Segerson 1989 et al 1985et al , Heuer et al 1998. In the hypothalamus, PPII is expressed in neurons and in tanycytes whose cytoplasmic extensions reach the external layer of the ME, in proximity to TRH terminals (Joseph-Bravo et al 1998, Sánchez et al 2009).…”

Sorting of growth hormone–erythropoietin fusion proteins in rat salivary glands

“…In the first, proteins are secreted as fast as they are synthesized, in a constitutive, non-directional manner and independent from changes in second messenger levels. In the second pathway, the regulated secretory pathway (RSP), proteins destined for secretion are sorted and stored in high concentrations in secretory granules where they await an external secretory stimulus [3][4][5]. RSP proteins require an amino acid-based sorting signal and evidence supports both the "sortingfor-entry" and "sorting-by-retention" hypotheses for these proteins [6][7][8][9][10].…”