Carboxypeptidase E Is a Regulated Secretory Pathway Sorting Receptor: Genetic Obliteration Leads to Endocrine Disorders in Cpefat Mice

Cool, David R.; Normant, Emmanuel; Shen, Fusheng; Chen, Hao-Chia; Pannell, Lewis K.; Zhang, Ying; Loh, Y. Peng

doi:10.1016/s0092-8674(00)81860-7

Cited by 433 publications

(388 citation statements)

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“…POMC is a pro-hormone that is post-translationally processed at paired basic residues to yield several biologically active peptide hormones, including adrenocortico-tropin (ACTH), aMSH, b-endorphin and gMSH. [1][2][3] The process that results in the generation of aMSH begins when POMC is directed to the lumen of the endoplasmic reticulum (ER) by the POMC signal peptide (amino acid residues . The sorting peptide (amino acid residues 27-52) subsequently directs POMC to secretion granules where it is further processed by pro-hormone convertase 1 (PC1) into the 39 amino acid peptide, ACTH (Figure 1).…”

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“…After the initial cleavage of POMC that creates ACTH, the first 17 amino acids of the ACTH peptide are then liberated by pro-hormone convertase 2 (PC2) to form the backbone of what will become native aMSH ( Figure 1). 2,[4][5][6] The final steps in the production of native aMSH involve cleavage of ACTH1-17 to ACTH1-13, followed by amidation of the carboxyl terminus and acetylation of the amino terminus. The amidation modification requires a signal, located at amino acids 14-16 of ACTH (Gly, Lys, Lys; Figure 1).…”

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“…Like most other hormones and neuropeptides, the subsequent release of aMSH from the cell occurs in response to certain stimuli, although there is some level of constitutive secretion. 2,7 Originally, it was thought that aMSH was produced primarily in the pituitary gland. More recently, the peptide has been found in several other regions of the brain and various peripheral organs including the skin.…”

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Generation of expression constructs that secrete bioactive αMSH and their use in the treatment of experimental autoimmune encephalomyelitis

et al. 2003

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a Melanocyte-stimulating hormone (aMSH) is a 13 amino acid peptide with potent anti-inflammatory effects. We created two DNA expression constructs (miniPOMC and pACTH1-17) that encode bioactive versions of the aMSH peptide, and tested these constructs for therapeutic effects in experimental autoimmune encephalomyelitis (EAE). Each construct contained the sequences for aMSH, as well as the sequences that are involved in the secretion and processing of the POMC gene with the assumption that these sequences would promote processing and release of the encoded aMSH peptide. The differences between the two constructs lie at the C-terminal end where amino acids necessary for amidation of aMSH were included in only the pACTH1-17 construct. These two constructs were tested in vitro in bioassays, and in vivo in a mouse model of EAE. The results show that although bioactive peptides are secreted from cells transfected with either construct, there appears to be a significant therapeutic effect only with the pACTH1-17 construct which contains the extra C-terminal amino acids. The data suggest that it is possible to engineer DNA expression vectors encoding small secreted peptides such as aMSH, and that similar type constructs may be useful as therapeutics for the treatment of inflammatory diseases.

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Generation of expression constructs that secrete bioactive αMSH and their use in the treatment of experimental autoimmune encephalomyelitis

et al. 2003

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“…Very recently, CPE has been proposed as the targeting receptor for POMC (18) with evidence that it interacts with the N-terminal sorting domain on this prohormone. Further, proinsulin and other proproteins, which are normally secreted through the regulated pathway but not proteins which are released through the constitutive pathway, were shown to displace POMC (or a peptide encompassing its sorting domain) from this putative sorting receptor (18).…”

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Proinsulin Targeting to the Regulated Pathway Is Not Impaired in Carboxypeptidase E-deficientCpe /Cpe Mice

Irminger

Verchere

Meyer

et al. 1997

Journal of Biological Chemistry

102

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Sorting of proinsulin from the trans-Golgi network to secretory granules is critical for its conversion to insulin as well as for regulated insulin secretion. The proinsulin sorting mechanism is unknown. Recently, carboxypeptidase E (CPE) was proposed as a sorting receptor for prohormones. To know whether CPE is implicated in proinsulin sorting, pancreatic islets were isolated from CPE-deficient Cpe fat /Cpe fat mice and Cpe fat /؉ controls, pulse-labeled ([ 3 H]leucine), and then chased in basal medium (90 min) to examine constitutive secretion followed by medium with secretagogues (60 min) to stimulate regulated secretion. Secretion of labeled proinsulin via the constitutive pathway was <2% even in Cpe fat /Cpe fat islets. After a 150-min chase, only 13% of radioactivity remained as proinsulin in Cpe fat /؉ islets compared with 46% in Cpe fat /Cpe fat islets, reflecting slower conversion. Regulated secretion was stimulated to an equal extent from Cpe fat /؉ and Cpe fat /Cpe fat mice with 20% of the total content of labeled (pro)insulin released during the 60-min stimulatory period. It is concluded that in CPE-deficient Cpe fat / Cpe fat mice, proinsulin is efficiently routed to the regulated pathway and its release can be effectively stimulated by secretagogues. CPE is thus not essential for sorting proinsulin to granules.Proinsulin is directed to the regulated secretory pathway of the pancreatic ␤-cell and is converted to bioactive insulin in immature secretory granules (1). At least three enzymes are needed for generating native insulin, namely the conversion endoproteases PC1 (also known as PC3) and PC2 (2, 3) and carboxypeptidase E (also called carboxypeptidase H or CPE 1 ) for trimming residual basic C-terminal residues (4 -6). Correct targeting to secretory granules is essential for the cell to produce and store bioactive insulin, which can then be released upon stimulation of the ␤-cell by glucose or other secretagogues. One of the crucial steps in targeting takes place in the TGN (trans-Golgi network) (7). The proprotein must somehow be recognized selectively in the TGN and then be transferred to the immature secretory granules, instead of being released through the constitutive or default pathway (3).One hypothesis (reviewed in Ref.3) accounting for targeting within the TGN implicates a sorting receptor with a broad specificity in the TGN membrane, which recognizes a sorting domain and thereby binds proteins to be routed to the regulated pathway. Sorting domains have been reported for several prohormones including prosomatostatin (8, 9), chromogranin (10), and the basic proline-rich protein (11). An N-terminal sorting domain has been reported for proopiomelanocortin (POMC) (12, 13), although this is controversial (14). A putative C-terminal sorting domain has also recently been proposed for the prohormone convertase PC2 (15). There is not as yet any direct experimental evidence for a sorting domain for proinsulin, although the comparison of structural features of many prohormones suggests that a regio...

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“…Although no universal regulated pathway-sorting motifs or signals have been identified [7,8], studies of prohormone/proneuropeptide trafficking have provided evidence that sorting of these molecules to the RSP involves a receptor-mediated mechanism [29,30]. For example, in endocrine cells, sorting motifs of pro-opiomelanocortin (POMC), brain derived neurotrophic factor and proinsulin were shown to interact with membrane carboxypeptidase E, which acts as a sorting or retention receptor to target these prohormones to the RSP [31].…”

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

Samuni

Zheng

Cawley

et al. 2008

Biochemical and Biophysical Research Communications

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Neuroendocrine and exocrine cells secrete proteins in either a constitutive manner or via the regulated secretory pathway (RSP), but the specific sorting mechanisms involved are not fully understood. After gene transfer to rat salivary glands, the transgenic model proteins human growth hormone (hGH) and erythropoietin (hEpo) are secreted primarily into saliva (RSP; exocrine) and serum (constitutive; endocrine), respectively. We hypothesized that fusion of hGH at either the C-terminus or the N-terminus of hEpo would re-direct hEpo from the bloodstream into saliva. We constructed and expressed two fusion proteins, hEpo-hGH and hGH-hEpo, using serotype 5-adenoviral vectors, and delivered them to rat submandibular glands in vivo via retroductal cannulation. Both the hEpohGH and hGH-hEpo fusion proteins, but not hEpo alone, were secreted primarily into saliva (p<0.0001 and P=0.0083, respectively). These in vivo studies demonstrate for the first time that hGH, in an N-as well as C-terminal position, influences the secretion of a constitutive pathway protein.

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Carboxypeptidase E Is a Regulated Secretory Pathway Sorting Receptor: Genetic Obliteration Leads to Endocrine Disorders in Cpefat Mice

Cited by 433 publications

References 40 publications

Generation of expression constructs that secrete bioactive αMSH and their use in the treatment of experimental autoimmune encephalomyelitis

Generation of expression constructs that secrete bioactive αMSH and their use in the treatment of experimental autoimmune encephalomyelitis

Proinsulin Targeting to the Regulated Pathway Is Not Impaired in Carboxypeptidase E-deficientCpe /Cpe Mice

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

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