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The Caenorhabditis elegans orthologue of mammalian puromycinsensitive aminepeptidas has roles in embryogenesis and reproduction

Brooks, DR, Hooper, NM and Isaac, RE 2003, 'The Caenorhabditis elegans orthologue of mammalian puromycinsensitive aminepeptidas has roles in embryogenesis and reproduction' , The Journal of Biological Chemistry, 278 (44) , pp. 42795-42801.

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Abstract

Mammals possess membrane-associated and cytosolic forms of the puromycin-sensitive aminopeptidase (PSA; EC 3.4.11.14). Increasing evidence suggests the membrane PSA is involved in neuromodulation within the central nervous system and in reproductive biology. The functional roles of the cytosolic PSA are less clear. The genome of the nematode Caenorhabditis elegans encodes an aminopeptidase, F49E8.3 (PAM-1), that is orthologous to PSA, and sequence analysis predicts it to be cytosolic. We have determined the spatio/temporal gene expression pattern of pam-1 by using the promoter region of F49E8.3 to control expression in the nematode of a second exon translational fusion of the aminopeptidase to green fluorescent protein. Cytosolic fluorescence was observed throughout development in the intestine and nerve cells of the head. Neuronal expression was also observed in the tail of adult males. Recombinant PAM-1, expressed and purified from Escherichia coli, hydrolyzed the N-terminal amino acid from peptide substrates. Favored substrates had positively charged or small neutral amino acids in the N-terminal position. Peptide hydrolysis was inhibited by the metal-chelating agent 1,10-phenanthroline and by the aminopeptidase inhibitors actinonin, amastatin, and leuhistin. However, the enzyme was ~100-fold less sensitive toward puromycin (IC50, 135 µM) than other PSA homologues. Following inactivation of the enzyme, aminopeptidase activity was recovered with Zn2+, Co2+, and Ni2+. Silencing expression of pam-1 by RNA interference resulted in 30% embryonic lethality. Surviving adult hermaphrodites deposited large numbers of oocytes throughout the self-fertile period. The overall brood size was, however, unaffected. We conclude that pam-1 encodes an aminopeptidase that clusters phylogenetically with the PSAs, despite attenuated sensitivity toward puromycin, and that it functions in embryo development and reproduction of the nematode.

Item Type: Article
Themes: Subjects / Themes > Q Science > Q Science (General)
Subjects outside of the University Themes
Schools: Colleges and Schools > College of Science & Technology
Colleges and Schools > College of Science & Technology > School of Environment and Life Sciences
Colleges and Schools > College of Science & Technology > School of Environment and Life Sciences > Biomedical Research Centre
Journal or Publication Title: The Journal of Biological Chemistry
Publisher: American Society for Biochemistry and Molecular Biology
Refereed: Yes
ISSN: 1083351X
Depositing User: H Kenna
Date Deposited: 07 Aug 2007 11:08
Last Modified: 20 Aug 2013 16:45
URI: http://usir.salford.ac.uk/id/eprint/128

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