Expression of a fluorescent recombinant form of sperm protein phospholipase C zeta in mouse epididymal sperm by in vivo gene transfer into the testis

Fertil Steril. 2006 Apr:85 Suppl 1:1281-9. doi: 10.1016/j.fertnstert.2005.12.012.

Abstract

Objective: To use in vivo gene transfer into the testis by electroporation to express a fluorescent recombinant form of a testis-specific gene in the mature epididymal sperm of mice and thus study the pattern of gene localization.

Design: Controlled animal study.

Setting: Research laboratory at the University of Oxford.

Animal(s): Four- to 6-week-old male mice.

Intervention(s): Phospholipase C zeta (PLCzeta), the putative mammalian egg activation factor, was fused to enhanced yellow fluorescent protein (EYFP), and in vivo gene transfer by electroporation was used to introduce this transgene (PLCzeta-EYFP) into mouse testis. Transgene expression in testis and sperm were analyzed at 20 and 40 days after electroporation.

Main outcome measure(s): Transgene expression in testis and epididymal sperm was analyzed by fluorescence microscopy and an excitation light source suitable for EYFP.

Result(s): Phospholipase C zeta-EYFP was successfully expressed in epididymal sperm when analyzed 40 days after gene transfer and was localized to the head and midpiece regions.

Conclusion(s): Our results provide the first demonstration that in vivo gene transfer can be used to study the localization of proteins in mature sperm and that this represents a powerful new technique for studying male infertility and gene function in sperm.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Cells, Cultured
  • Electroporation / methods*
  • Epididymal Secretory Proteins / biosynthesis*
  • Epididymal Secretory Proteins / genetics*
  • Gene Expression Regulation / physiology
  • Gene Transfer Techniques
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Male
  • Mice
  • Mice, Transgenic
  • Microscopy, Fluorescence
  • Phosphoinositide Phospholipase C
  • Recombinant Fusion Proteins / metabolism
  • Spermatozoa / metabolism*
  • Testis / metabolism*
  • Tissue Distribution
  • Type C Phospholipases / genetics*
  • Type C Phospholipases / metabolism*

Substances

  • Bacterial Proteins
  • Epididymal Secretory Proteins
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • yellow fluorescent protein, Bacteria
  • Type C Phospholipases
  • Phosphoinositide Phospholipase C
  • Plcz1 protein, mouse