Prohormone convertase 1 (PC1) is a serine proteinase responsible for the proteolytic processing of many precursor proteins within the regulated secretory pathway. The activity of PC1 is potentially regulated by two endogenous inhibitors, the PC1 propeptide and proSAAS. Here we have investigated the effect of proSAAS and propeptide-containing constructs on PC1 carboxy-terminal processing and activity. In AtT-20 cells, proSAAS expression inhibited both C-terminal PC1 processing and proopiomelanocortin (POMC) processing under pulse/chase conditions. SAAS CT peptide-propeptide chimeric constructs had no effect on the cleavage of PC1 and POMC under pulse/chase conditions. However, a construct containing the propeptide alone reduced C-terminal PC1 processing under pulse/chase conditions and also inhibited POMC processing. In contrast, experiments using HEK293 cells transiently expressing PC1 plus the respective constructs demonstrated significant inhibition of zymogen processing and decreased C-terminal processing of PC1 by the SAAS CT peptide portion of the chimera. Our results suggest that the PC1 propeptide expressed in trans is able to act as an endogenous inhibitor of PC1, but that SAAS CT peptide-containing/propeptide constructs cannot function as effective inhibitors of precursor maturation in the regulated pathway.
SN Lee, E Prodhomme, and I Lindberg
E. L. Sheldrick and A. P. F. Flint
Peptidyl glycine α-amidating mono-oxygenase (PGA), the terminal enzyme in the pathway of oxytocin synthesis, was measured in extracts of ovine corpora lutea throughout the oestrous cycle. Activity of PGA was low early in the cycle but increased between days 2 and 10 (from 2·3 to 9·0 pmol/mg protein per h) and remained high until day 15. Thereafter, activity declined rapidly at structural luteolysis and was low in corpora albicantia collected 18 and 20 days after ovulation (1·28 and 1·07 pmol/mg protein per h respectively). Luteal concentrations of ascorbic acid, a cofactor for PGA, were high (4·7 μmol/g wet wt tissue) by day 4 after oestrus; concentrations fell rapidly after day 15 (to 2·1 μmol/g on day 16). Concentrations of ascorbic acid were also high in the pituitary gland and in the adrenal medulla and cortex. Concentrations of oxytocin in luteal tissue, which were low (0·3 nmol/g wet wt) on day 2 after oestrus, were highest (2·73 nmol/g) on day 6 and declined thereafter (0·56 nmol/g on day 10, 0·08 nmol/g on day 15 and not detectable on days 18 and 20).
Concentrations of oxytocin, progesterone, PGA and protein were measured in subcellular fractions obtained after density gradient centrifugation of extracts of corpora lutea collected on days 6, 7 and 12 of the oestrous cycle, and on day 7 from an anaesthetized ewe before and after treatment with the prostaglandin F2α analogue, cloprostenol. PGA co-localized with particle-associated oxytocin in fractions of density 1·049–1·054 g/ml. Exogenous [3H]oxytocin and [3H]progesterone and endogenous progesterone localized in fractions of density 1·035 g/ml. Oxytocin and PGA were depleted from fractions of density 1·049–1·054 g/ml following cloprostenol treatment in vivo.
Fractionation of extracts of ovine corpora lutea by high-performance liquid chromatography (HPLC) followed by radioimmunoassay and radioreceptor assay for oxytocin demonstrated the presence of at least two cross-reacting substances with elution characteristics distinct from oxytocin. Concentrations of these peptides increased as the cycle progressed. These compounds differed from the putative C-terminally extended post-translational processing intermediates, oxytocinyl-glycine, oxytocinyl-glycine-lysine and oxytocinyl-glycine-lysine-arginine, as indicated by their elution positions on HPLC and the specificities of the assays used to detect them, and no conclusions could be drawn on which post-translational processing step was rate-limiting in oxytocin synthesis.
These data are consistent with the suggestion that post-translational processing of oxytocin-neurophysin prohormone takes place in secretory granules in luteal cells. The low activity of PGA early in the cycle may account for the lag previously observed between concentrations of the oxytocin-neurophysin prohormone mRNA and the mature peptide, but post-translational processing intermediates could not be identified. The rate of α-amidation is unlikely to be controlled by availability of ascorbic acid.
Journal of Endocrinology (1989) 122, 313–322
Niamh X Cawley, Guida Portela-Gomes, Hong Lou, and Y Peng Loh
performed by Kex2, a subtilisin-like serine protease, it was speculated that yapsins may be backup enzymes for the serine proteases involved in prohormone processing. Mammalian aspartic proteases with similar properties to the yapsins have been characterized
Rebecca McGirr, Leonardo Guizzetti, and Savita Dhanvantari
, processing and storage of peptide hormones. Peptide hormones are first synthesised as larger precursors, or prohormones, and are selectively targeted to the regulated secretory pathway via the trans -Golgi network (TGN). Proteins destined for the regulated
N M Whalley, L E Pritchard, D M Smith, and A White
Introduction Like many prohormones, proglucagon is processed in a cell type-specific manner. In the α-cells of the pancreas, proglucagon is processed to glucagon by prohormone convertase 2 (PC2), but it undergoes alternative processing in the L
A Alidibbiat, C E Marriott, K T Scougall, S C Campbell, G C Huang, W M Macfarlane, and J A M Shaw
pathway necessary for post-translational processing and storage of proinsulin was shown by negative prohormone convertase RT-PCR. This accounted for absent immunocytochemical staining with an antibody raised against mature insulin and undetectable insulin
Uxía Gurriarán-Rodríguez, Omar Al-Massadi, Ana Belén Crujeiras, Carlos S Mosteiro, María Amil-Diz, Daniel Beiroa, Rubén Nogueiras, Luisa María Seoane, Rosalía Gallego, Yolanda Pazos, Felipe F Casanueva, and Jesús P Camiña
talk coordinates a variety of biological processes including energy metabolism, neuroendocrine function, and immune function ( Scherer et al . 2006 , Virtue & Vidal-Puig 2010 ). Added to this capacity, adipose tissue expresses a broad spectrum of
Abdullah Cim, Greta J Sawyer, Xiaohong Zhang, Haibin Su, Louise Collins, Peter Jones, Michael Antoniou, Jean-Paul Reynes, Hans-Joachim Lipps, and John W Fabre
probably represents the result of a more complex transdifferentiation process. To investigate this further, the presence of other pancreatic β cell proteins was evaluated. The results in Fig. 5 demonstrate that the prohormone convertase 1/3, essential for
Deiodinases: the balance of thyroid hormone
Ana Luiza Maia, Iuri Martin Goemann, Erika L Souza Meyer, and Simone Magagnin Wajner
D2), via outer (5′)-ring deiodination of the pro-hormone T 4 . Type 3 iodothyronine deiodinase (D3) catalyzes the inner (5)-ring deiodination of T 4 and T 3 , thus inactivating the thyroid hormone action. In the last decades, several studies have
B. G. Jenks, A. G. H. Ederveen, J. H. M. Feyen, and A. P. van Overbeeke
Pro-opiomelanocortin (POMC) is a glycoprotein precursor for a number of neuropeptides and peptide hormones. The functional significance of the glycosylation of POMC has never been established. Using the antibiotic tunicamycin to block glycosylation of the prohormone in the mouse pars intermedia, we have compared processing of non-glycosylated prohormone with that of glycosylated prohormone in pulse-chase experiments. The peptides produced from non-glycosylated prohormone were shown to be correct cleavage products. Therefore it was concluded that, with the possible exception of peptides from the N-terminal region of the prohormone, the carbohydrate on POMC plays no role in directing cleavage or in protecting the prohormone from random proteolysis. Tunicamycin treatment retarded N-terminal acetylation of melanotrophin but had no apparent effect on acetylation of β-endorphin. The mouse pars intermedia synthesizes two forms of POMC which differ in their degree of glycosylation. Our results indicated that, during secretion, the melanotrophs make no distinction between peptides derived from the two prohormones.
J. Endocr. (1985) 107, 365–374