Histochemistry and Cell Biology

, Volume 136, Issue 6, pp 677–688 | Cite as

Dictyostelium puromycin-sensitive aminopeptidase A is a nucleoplasmic nucleomorphin-binding protein that relocates to the cytoplasm during mitosis

  • Andrew Catalano
  • Yekaterina Poloz
  • Danton H. O’Day
Original Paper


Nucleomorphin (NumA1) is a nucleolar/nucleoplasmic protein linked to cell cycle in Dictyostelium. It interacts with puromycin-sensitive aminopeptidase A (PsaA) which in other organisms is a Zn2+-metallopeptidase thought to be involved in cell cycle progression and is involved in several human diseases. Here, we have shown that Dictyostelium PsaA contains domains characteristic of the M1 family of Zn2+-metallopeptidases: a GAMEN motif and a Zn2+-binding domain. PsaA colocalized with NumA1 in the nucleoplasm in vegetative cells and was also present to a lesser extent in the cytoplasm. The same localization pattern was observed in cells from slugs, however, in fruiting bodies PsaA was only detected in spore nuclei. During mitosis PsaA redistributed mainly throughout the cytoplasm. It possesses a functional nuclear localization signal (680RKRF683) necessary for nuclear entry. To our knowledge, this is the first nuclear localization signal identified in a Psa from any organism. Treatment with Ca2+ chelators or calmodulin antagonists indicated that neither Ca2+ nor calmodulin is involved in PsaA localization. These results are interpreted in terms of the inter-relationship between NumA1 and PsaA in cell function in Dictyostelium.


Puromycin-sensitive aminopeptidase Nucleoplasm Mitosis Nuclear localization signal Nucleomorphin 



This work was supported by the Natural Sciences and Engineering Research Council of Canada (DHO’D; A6807).


  1. Bhutani N, Venkatraman P, Goldberg AL (2007) Puromycin-sensitive aminopeptidase is the major peptidase responsible for digesting polyglutamine sequences released by proteasomes during protein degradation. EMBO J 26:1385–1396PubMedCrossRefGoogle Scholar
  2. Brooks DR, Hooper NM, Isaac RE (2003) The Caenorhabditis elegans orthologue of mammalian puromycin-sensitive aminopeptidase has roles in embryogenesis and reproduction. J Biol Chem 278:42795–42801PubMedCrossRefGoogle Scholar
  3. Catalano A, O’Day DH (2008) Calmodulin-binding proteins in the model organism Dictyostelium. A complete & critical review. Cell Signal 20:277–291PubMedCrossRefGoogle Scholar
  4. Catalano A, O’Day DH (2011a) Calcium-dependent nucleolar localization of Dictyostelium calcium-binding protein 4a (CBP4a/cbpD1) during interphase and redistribution throughout mitosis. Cell Calcium (Submitted)Google Scholar
  5. Catalano A, O’Day DH (2011b) Nucleolar localization and identification of nuclear/nucleolar localization signals of the calmodulin-binding protein nucleomorphin during growth and mitosis in Dictyostelium. Histochem Cell Biol 135:239–249PubMedCrossRefGoogle Scholar
  6. Chen S-H, Cao M-J, Huang J-Z, Wu G-P (2011) Identification of a puromycin-sensitive aminopeptidase from zebrafish (Danio rerio). Comp Biochem Physiol Part B 159:10–17CrossRefGoogle Scholar
  7. Constam DB, Tobler AR, Rensing-Ehl A, Kemler I, Hersh LB, Fontana A (1995) Puromycin-sensitive aminopeptidase. Sequence analysis, expression, and functional characterization. J Biol Chem 270:26931–26939PubMedCrossRefGoogle Scholar
  8. De Souza CPC, Osmani SA (2007) Mitosis, not just open or closed. Eukaryotic Cell 6:1521–1527PubMedCrossRefGoogle Scholar
  9. Fey P, Kowal AS, Gaudet P, Pilcher KE, Chisholm RL (2007) Protocols for growth and development of Dictyostelium discoideum. Nat Protoc 2:1307–1316PubMedCrossRefGoogle Scholar
  10. Hagedorn M, Neuhaus EN, Soldati T (2006) Optimised fixation and immunofluorescence protocols for Dictyostelium cells. Methods Mol Biol 346:327–338PubMedGoogle Scholar
  11. Huber R, O’Day DH (2011) Nucleocytoplasmic transfer of cyclin dependent kinase 5 and its binding to puromycin-sensitive aminopeptidase in Dictyostelium discoideum. Histochem Cell Biol. doi: 10.1007/s00418-011-0839-6
  12. Hui K-S, Saito M, Hui M, Saito M, Lajtha A, Yamamoto K, Osawa T (1993) Two cytosolic puromycin-sensitive aminopeptidase isozymes in chicken brain: molecular homology to brain-specific 14-3-3 protein. Neurochem Int 22:445–453PubMedCrossRefGoogle Scholar
  13. Karsten SL, Sang T-K, Gehman LT, Chatterjee S, Liu J, Lawless GM, Sengupta S, Berry RW, Pomakian J, Oh HS, Schulz C, Hui K-S, Wiedau-Pazos M, Vinters HV, Binder LI, Geschwind DH, Jackson GR (2006) A genomic screen for modifiers of tauopathy identifies puromycin-sensitive aminopeptidase as an inhibitor of Tau-induced neurodegeneration. Neuron 51:549–560PubMedCrossRefGoogle Scholar
  14. Kudo LC, Parfenova L, Ren G, Vi N, Hui M, Ma Z, Lau K, Gray M, Bardag-Gorce F, Wiedau-Pazos M, Hui K-S, Karsten L (2011) Puromycin-sensitive aminopeptidase (PSA/NPEPPS) impedes development of neuropathology in hPSA/TAUP301L double-transgenic mice. Hum Mol Genet 20:1820–1833PubMedCrossRefGoogle Scholar
  15. Lateef SS, Gupta S, Jayathilaka LP, Krishnanchettiar S, Huang J-S, Lee B-S (2007) An improved protocol for coupling synthetic peptides to carrier proteins for antibody production using DMF to solubilize peptides. J Biomol Tech 18:173–176PubMedGoogle Scholar
  16. Lee SH, Kim HG (2009) Cobalt chloride-induced downregulation of puromycin-sensitive aminopeptidase suppresses the migration and invasion of PC-3 cells. J Microbiol Biotechnol 19:530–536PubMedCrossRefGoogle Scholar
  17. Lyczak R, Zweier L, Group T, Murrow MA, Snyder C, Kulovitz L, Beatty A, Smith K, Bowerman B (2006) The puromycin-sensitive aminopeptidase PAM-1 is required for meiotic exit and anteroposterior polarity in the one-cell Caenorhabditis elegans embryo. Development 133:4281–4292PubMedCrossRefGoogle Scholar
  18. Myre MA (2005) Characterization of nucleomorphin, a novel nuclear breast cancer carboxy-terminal-domain containing calmodulin-binding protein from Dictyostelium discoideum. PhD dissertation, Department of Zoology, University of Toronto, CanadaGoogle Scholar
  19. Myre MA, O’Day DH (2002) Nucleomorphin: a novel, acidic, nuclear calmodulin-binding protein from Dictyostelium that regulates nuclear number. J Biol Chem 277:19735–19744PubMedCrossRefGoogle Scholar
  20. Myre MA, O’Day DH (2004a) Dictyostelium calcium-binding protein 4a interacts with nucleomorphin, a BRCT-domain protein that regulates nuclear number. Biochem Biophys Res Commun 322:665–671PubMedCrossRefGoogle Scholar
  21. Myre MA, O’Day DH (2004b) Dictyostelium nucleomorphin is a member of the BRCT-domain family of cell cycle checkpoint proteins. Biochim Biophys Acta 1675:192–197PubMedGoogle Scholar
  22. O’Day DH, Poloz Y, Myre MA (2009) Differentiation inducing factor-1 (DIF-1) induces gene and protein expression of the Dictyosetlium nuclear calmodulin-binding protein nucleomorphin. Cell Signal 21:317–323PubMedCrossRefGoogle Scholar
  23. Osada T, Watanabe G, Kondo S, Toyoda M, Sakaki Y, Takeuchi T (2001) Male reproductive defects caused by puromycin-sensitive aminopeptidase deficiency in mice. Mol Endocrinol 15:960–971PubMedCrossRefGoogle Scholar
  24. Peer WA (2011) The role of multifunctional M1 metallopeptidases in cell cycle progression. Ann Bot. doi: 10.1093/aob/mcq265
  25. Rawlings ND, Barrett AJ (1995) Evolutionary families of metallopeptidases. Method Enzymol 248:183–228CrossRefGoogle Scholar
  26. Reinders Y, Schulz I, Graf R, Sickmann A (2006) Identification of novel centrosomal proteins in Dictyosetlium discoideum by comparative proteomic approaches. J Proteome Res 5:589–598PubMedCrossRefGoogle Scholar
  27. Sanchez-Moran E, Jones GH, Franklin CH, Santos JL (2004) A puromycin-sensitive aminopeptidase is essential for meiosis in Arabidopsis thaliana. Plant Cell 16:2895–2909PubMedCrossRefGoogle Scholar
  28. Takahashi S, Ohishi Y, Kato H, Noguchi T, Naito H, Aoyagi T, Umezawa H (1989) The effects of bestatin, a microbial aminopeptidase inhibitor, on epidermal growth factor- induced DNA synthesis and cell division in primary cultured hepatocytes of rats. Exp Cell Res 183:399–412PubMedCrossRefGoogle Scholar
  29. Taylor A (1993) Aminopeptidases: structure and function. FASEB J 296:290–298Google Scholar
  30. Veltman DM, Akar G, Bosgraff L, van Haastert PJ (2009) A new set of small, extrachromosomal expression vectors for Dictyostelium discoideum. Plasmid 61:110–118PubMedCrossRefGoogle Scholar
  31. Yamamoto M, Chikuma T, Yajima R, Hirano H, Yamamoto Y, Nishi K, Ohkubo I, Kato T (2002) Axonal transport of puromycin-sensitive aminopeptidase in rat sciatic nerves. Neurosci Res 42:133–140PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Andrew Catalano
    • 1
  • Yekaterina Poloz
    • 1
  • Danton H. O’Day
    • 1
    • 2
  1. 1.Department of Cell and Systems BiologyUniversity of TorontoTorontoCanada
  2. 2.Department of BiologyUniversity of Toronto at MississaugaMississaugaCanada

Personalised recommendations