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Histochemistry and Cell Biology

, Volume 123, Issue 1, pp 51–60 | Cite as

Fluorescently labeled inhibitors detect localized serine protease activities in Drosophila melanogaster pole cells, embryos, and ovarian egg chambers

  • Rasmus Kragh Jakobsen
  • Shin Ono
  • James C. Powers
  • Robert DeLotto
Original Paper

Abstract

Serine proteases are typically synthesized as proteolytically inactive zymogens that often become activated in a limited and highly localized manner. Consequently, determination of the spatial and temporal activation pattern of these molecules is of great importance to understanding the biological processes that they mediate. Until only recently, the tools to conveniently address the question of where and when serine proteases are active within complex tissues have been lacking. In order to detect spatially restricted serine protease activities in Drosophila embryos and ovaries we introduce a technique using fluorescent synthetic and protein-based inhibitors. With this approach we have detected a novel serine protease activity with a relative mobility of 37 kDa, localized to the surface of pole cells, the germ-line precursors, in embryos between nuclear cycles 11 and 14 in development. A second novel cell-specific protease activity was localized to the tissues of early gastrulating embryos. Microinjection of inhibitors into the perivitelline space of stage 2 embryos perturbed normal embryonic development. Fluorescein-conjugated chymotrypsin inhibitor and Bowman-Birk inhibitor labeled protease activity localized to the oocyte–somatic follicle cell interface of the developing egg chamber. Our results suggest that this technique holds promise to identify new spatially restricted activities in adult Drosophila tissues and developing embryos.

Keywords

Oogenesis Embryonic patterning Chloromethyl ketone Phosphonate Zymogen activation 

Notes

Acknowledgements

We would like to thank Yvonne DeLotto for excellent technical assistance with the microinjections and Jakob Winther for constructive experimental criticism. This work was supported by the Danish Natural Science Research Council, the Danish Cancer Fund, the Vera and Carl Johan Michaelsens Legacy, and the US National Science Foundation to R.D., and by grants from the National Institute of General Medical Sciences (grants GM54401 and GM61964) to J.C.P.

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Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  • Rasmus Kragh Jakobsen
    • 1
  • Shin Ono
    • 2
  • James C. Powers
    • 3
  • Robert DeLotto
    • 1
  1. 1.Department of Genetics, Institute of Molecular BiologyUniversity of CopenhagenCopenhagen KDenmark
  2. 2.Department of System Engineering of Materials and Life Science, Faculty of EngineeringToyama UniversityToyamaJapan
  3. 3.Department of Chemistry and BiochemistryGeorgia Institute of TechnologyAtlantaUSA

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