title: Ultrastructural localization and distribution of Nardilysin in mammalian male germ cells.
abstract: BACKGROUND
  NRD convertase, also termed Nardilysin, is a Zn(++) metalloendopeptidase that specifically cleaves the N-terminus of arginine and lysine residues into dibasic moieties. Although this enzyme was found located within the testis, its function in male reproduction is largely unknown. In addition, the precise distribution of this enzyme within germ cells remains to be determined.
  
  
  METHODS
  To answer these questions, we developed an immuno-gold electron microscopy analysis to detect Nardilysin at ultrastructural level in mice. In addition, we performed a quantitative analysis of these gold particles to statistically estimate the distribution of Nardilysin in the different subcellular compartments of differentiating late spermatids/spermatozoa.
  
  
  RESULTS
  Expression of Nardilysin in wild-type mice was restricted to germ cells and markedly increased during the last steps of spermiogenesis. In elongated spermatids, we found the enzyme mainly localized in the cytoplasm, more precisely associated with two microtubular structures, the manchette and the axoneme. No labelling was detected over the membranous organelles of the spermatids. To test whether this localization is dependent of the functional microtubules organization of the flagella, we analysed the localization into a specific mouse mutant ebo/ebo (ébouriffé) known to be sterile due to an impairment of the final organization of the flagellum. In the ebo/ebo, the enzyme was still localized over the microtubules of the axoneme and over the isolated cytoplasmic microtubules doublets. Quantification of gold particles in wild-type and mutant flagella revealed the specific association of the enzyme within the microtubular area of the axoneme.
  
  
  CONCLUSIONS
  The strong and specific accumulation of Nardilysin in the manchette and axoneme suggests that the enzyme probably contributes either to the establishment of these specific microtubular structures and/or to their functional properties.
authors.affiliation: UMR S 1147 Université Paris Descartes, 45 rue des Saint-Pères, 75006 Paris, France ; Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), Versailles, 78000 France.
authors.forename: D
authors.initials: D
authors.lastname: Segretain
authors.affiliation: INSERM U 1065, Université Nice Sophia-Antipolis, 151 route Saint-Antoine de Ginestière BP 2 3194, 06204, Nice, cedex 3 France.
authors.forename: J
authors.initials: J
authors.lastname: Gilleron
authors.affiliation: Institut de Mathématiques et de Modélisation de Montpellier (I3M), UMR CNRS 5149 Université Montpellier, CC 51; 4 place Eugène Bataillon 34095, Montpellier, cedex 5 France.
authors.forename: J N
authors.initials: JN
authors.lastname: Bacro
authors.affiliation: UMR S 1147 Université Paris Descartes, 45 rue des Saint-Pères, 75006 Paris, France ; Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), Versailles, 78000 France.
authors.forename: M
authors.initials: M
authors.lastname: Di Marco
authors.affiliation: UMR S 1147 Université Paris Descartes, 45 rue des Saint-Pères, 75006 Paris, France ; Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), Versailles, 78000 France.
authors.forename: D
authors.initials: D
authors.lastname: Carette
authors.affiliation: INSERM U 1065, Université Nice Sophia-Antipolis, 151 route Saint-Antoine de Ginestière BP 2 3194, 06204, Nice, cedex 3 France.
authors.forename: G
authors.initials: G
authors.lastname: Pointis
keywords: Flagellum; Gold immunohistochemistry; Nardilysin; Spermiogenesis