Nuclear shuttling of the peptidase nardilysin

Zhangliang Ma, K. Martin Chow, Jia Yao, Louis B. Hersh

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

The metalloendopeptidase nardilysin contains a putative N-terminal nuclear localization signal. The functionality of this sequence was tested with nardilysin-GFP fusion constructs. Expression in NIH3T3 cells showed ∼90-95% of nardilysin-GFP as cytoplasmic. However, 3-6% of transfected cells showed both cytosolic and nuclear staining, while 2-4% showed predominantly nuclear staining. A nuclear localization signal mutant and an N-terminally truncated nardilysin-GFP with the nuclear localization signal deleted were completely cytoplasmic. Although endogenous nardilysin was barely detectable in the nucleus, after treatment with leptomycin B, nuclear nardilysin rose to ∼15% and to over 25% after addition of spermine. The ability of a methionine 49 to act as the sole initiator methionine, as previously proposed, was tested by inserting a c-myc epitope between leucine28 and glycine 29. Expression in HEK293 cells showed the presence of the c-myc tag, demonstrating that the enzyme can be translated from the first methionine and contains the nuclear localization signal.

Original languageEnglish
Pages (from-to)153-160
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume422
Issue number2
DOIs
StatePublished - Feb 15 2004

Bibliographical note

Funding Information:
We thank Dr. Sidney W. Whiteheart (Department of Molecular and Cellular Biochemistry, University of Kentucky) for kindly providing thiolase antisera. We also thank Dr. Daniel Noonan (Department of Molecular and Cellular Biochemistry, University of Kentucky) for helpful discussions. This research was supported in part by NIH/NIDA Grant DA02243 to L.B.H.

Keywords

  • GFP
  • Nardilysin
  • Neuropeptidase
  • Nuclear export
  • Nuclear localization
  • Spermine

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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