Cloning and Functional Analysis of<i>C. elegans</i>7B2

Lindberg, Iris; Tu, Bin; Muller, Laurent; Dickerson, Ian M.

doi:10.1089/dna.1998.17.727

Cited by 32 publications

(34 citation statements)

References 25 publications

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“…Both PC2 and 7B2 are highly conserved in evolution; homologues of 7B2 have recently been described in the molluscan Lymnaea stagnalis (52) and in Caenorhabditis elegans (53). Recently, mice with a disruption of the 7B2 gene have been produced and, as expected, they lack active PC2 but have other defects (54).…”

mentioning

confidence: 57%

Proteolytic Processing in the Secretory Pathway

Zhou¹,

Webb²,

Zhu³

et al. 1999

Journal of Biological Chemistry

453

416

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mentioning

confidence: 57%

Proteolytic Processing in the Secretory Pathway

Zhou¹,

Webb²,

Zhu³

et al. 1999

Journal of Biological Chemistry

453

416

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“…The lack of importance of this chaperone-like domain for proPC2 activation is in agreement with its weak conservation through evolution. Whereas the sequences of X. laevis (34) and mammalian 7B2s (35)(36)(37) share more than 90% homology, the recent cloning of invertebrate 7B2s, from Lymnea stagnalis (31), C. elegans (27), and D. melanogaster 2 shows no significant conservation in the amino-terminal half of these 7B2s. Whereas 7B2 86 -150 , which excludes the 7B2 chaperone-like domain, could not facilitate proPC2 maturation, the 7B2 68 -150 construct was active.…”

Section: Discussionmentioning

confidence: 99%

“…The recent cloning of 7B2 from Caenorhabditis elegans (27) and Drosophila melanogaster 2 showed that this putative ␣-helix is conserved, even though the overall conservation of 7B2 through evolution is quite poor. In order to test the requirement for the ␣-helix, we introduced two mutations: the replacement of an arginine by a proline (R116P), which should disrupt the helical structure, and the insertion of an alanine at position 115 (ϩA115), which should preserve the helical structure but modify the distribution of the residues on its surface (Fig.…”

Section: Fig 1 the Amino-terminal Half Of 7b2 Is Not Required For Bmentioning

confidence: 99%

“…The Disulfide Bond Is Required for PC2 Activation-Another feature of the 86 -121 peptide is the presence of the sole two cysteine residues in mammalian 7B2s, which are also conserved in all of the invertebrate 7B2s cloned thus far (27,31). 2 These cysteines form a disulfide bond that is apparently not required for correct processing and for secretion of 7B2 (17,32).…”

Section: Fig 1 the Amino-terminal Half Of 7b2 Is Not Required For Bmentioning

confidence: 99%

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A 36-Residue Peptide Contains All of the Information Required for 7B2-mediated Activation of Prohormone Convertase 2

Muller¹,

Zhu²,

Juliano³

et al. 1999

Journal of Biological Chemistry

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The prohormone convertases (PCs) are serine proteinases responsible for the processing of secretory protein precursors. PC2 is the only member of this family whose activation requires intracellular interaction with a helper protein, the neuroendocrine protein 7B2. In order to gain a better understanding of the mechanism of proPC2 activation, we have characterized the structural determinants of 7B2 required for proPC2 activation. We had already identified a proline-rich binding determinant in the 21-kDa domain, the portion of 7B2 responsible for proPC2 activation. We have now investigated the function of the weakly conserved amino-terminal portion of 21-kDa 7B2 by sequential deletions. Mutant proteins were analyzed in four assays: binding to proPC2, facilitation of proPC2 maturation, and activation of proPC2 in vivo and in vitro. We found that the aminoterminal half of 7B2 is not involved in proPC2 activation, and we identified an active 36-residue peptide that contains the previously characterized proline-rich sequence as well as an ␣-helix and the only disulfide bond of 7B2. Mutation of the ␣-helix and of the cysteines demonstrated that these determinants are absolutely required for PC2 activation. Thus, the 186-residue fulllength 7B2 rat protein can be functionally reduced to an internal segment of only 36 residues.Endoproteolytic processing is one of the major post-translational modifications that hormones and neuropeptides precursors must undergo during their biosynthesis. A family of mammalian subtilisin-like endoproteases responsible for these processing events has been recently identified, the prohormone convertases (PCs) 1 (reviewed in Refs. 1 and 2). PC1 and PC2 are the prohormone convertases specific for neuroendocrine cells; both enzymes are active late in the secretory pathway, i.e. the trans-Golgi network (TGN) and the secretory granules.Precursors such as proinsulin or proopiomelanocortin are processed sequentially, first by PC1, then by PC2 (reviewed in Ref.3). In agreement with the ordered activity of these enzymes, PC1 and PC2 have different activation pathways (reviewed in Ref. 4). Whereas the propeptide of proPC1 is first processed in the endoplasmic reticulum (ER) (5-7), as are those of furin (8, 9), PC5 (10), and LPC/PC7/PC8 (11), the proPC2 propeptide is processed only in the acidic compartments of the TGN/secretory granules. In addition, proPC2 is the only PC that specifically interacts with a helper protein, the neuroendocrine-specific protein 7B2 (12)(13)(14). We have demonstrated that this interaction is absolutely required for proPC2 activation, both in vivo in transfected and in 7B2 null mice (16), as well as in vitro (17).The neuroendocrine protein 7B2 was originally purified from pituitary extracts (18). It is an 185-residue precursor that is cleaved at a pentabasic site at residue 155 (Fig. 1), most probably by furin (19 -21). The two domains generated by this cleavage have distinct functions; the amino-terminal domain of 21 kDa is sufficient for PC2 activation (14), and the COOHt...

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“…5B) from 7B2 (P12961). 7B2 was first discovered in 1982 (39,40); it is a bifunctional polypeptide that is highly conserved in multiple species (41)(42)(43)(44)(45)(46). The N-terminal peptide of the polypeptide binds to PC2 and is essential for its activation; in contrast the C-terminal peptide functions as its inhibitor (47,48).…”

Section: Figmentioning

confidence: 99%

Neuropeptidomics Strategies for Specific and Sensitive Identification of Endogenous Peptides

Fälth

Sköld

Svensson

et al. 2007

Molecular & Cellular Proteomics

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A new approach using targeted sequence collections has been developed for identifying endogenous peptides. This approach enables a fast, specific, and sensitive identification of endogenous peptides. Three different sequence collections were constituted in this study to mimic the peptidomic samples: SwePep precursors, SwePep peptides, and SwePep predicted. The searches for neuropeptides performed against these three sequence collections were compared with searches performed against the entire mouse proteome, which is commonly used to identify neuropeptides. These four sequence collections were searched with both Mascot and X! Tandem. Evaluation of the sequence collections was achieved using a set of manually identified and previously verified peptides. By using the three new sequence collections, which more accurately mimic the sample, 3 times as many peptides were significantly identified, with a false-positive rate below 1%, in comparison with the mouse proteome. The new sequence collections were also used to identify previously uncharacterized peptides from brain tissue; 27 previously uncharacterized peptides and potentially bioactive neuropeptides were identified. These novel peptides are cleaved from the peptide precursors at sites that are characteristic for prohormone convertases, and some of them have post-translational modifications that are characteristic for neuropeptides. The targeted protein sequence collections for different species are publicly

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Cloning and Functional Analysis ofC. elegans7B2

Cited by 32 publications

References 25 publications

Proteolytic Processing in the Secretory Pathway

Proteolytic Processing in the Secretory Pathway

A 36-Residue Peptide Contains All of the Information Required for 7B2-mediated Activation of Prohormone Convertase 2

Neuropeptidomics Strategies for Specific and Sensitive Identification of Endogenous Peptides

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