Targeted Gene Deletion and Phenotypic Analysis of the Drosophila melanogaster Seminal Fluid Protease Inhibitor Acp62F

Internally fertilizing organisms transfer a complex assortment of seminal fluid proteins, a substantial fraction of which are proteolysis regulators. In mammals, some seminal protease inhibitors have been implicated in male infertility and these same molecular classes of protease inhibitors are also found in Drosophila seminal fluid. Here, we tested the reproductive functions of the Drosophila melanogaster seminal fluid protease inhibitor Acp62F by generating a precise deletion of the Acp62F gene. We did not detect a nonredundant function for Acp62F in modulating the egg laying, fertility, remating frequency, or life span of mated females. However, loss of Acp62F did alter a male's defensive sperm competitive ability, consistent with the localization of Acp62F to sperm storage organs. In addition, the processing of at least one seminal protein, the ovulation hormone ovulin, is slower in the absence of Acp62F.

[1]  Y. Heifetz,et al.  Mating-responsive genes in reproductive tissues of female Drosophila melanogaster , 2006, Proceedings of the National Academy of Sciences.

[2]  T. Aigaki,et al.  Ectopic expression of sex peptide alters reproductive behavior of female D. melanogaster , 1991, Neuron.

[3]  M. Wolfner,et al.  Structure and expression of a Drosophila male accessory gland gene whose product resembles a peptide pheromone precursor. , 1988, Genes & development.

[4]  Laura N. Vandenberg,et al.  Two cleavage products of the Drosophila accessory gland protein ovulin can independently induce ovulation. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[5]  T. Honegger,et al.  Gradual Release of Sperm Bound Sex-Peptide Controls Female Postmating Behavior in Drosophila , 2005, Current Biology.

[6]  A. Clark,et al.  Sperm Competitive Ability in Drosophila melanogaster Associated With Variation in Male Reproductive Proteins , 2005, Genetics.

[7]  Y. Rong,et al.  A targeted gene knockout in Drosophila. , 2001, Genetics.

[8]  H. Lilja,et al.  Protein C inhibitor in human body fluids. Seminal plasma is rich in inhibitor antigen deriving from cells throughout the male reproductive system. , 1992, The Journal of clinical investigation.

[9]  A. Civetta,et al.  Quantitative trait loci and interaction effects responsible for variation in female postmating mortality in Drosophila simulans and D. sechellia introgression lines , 2005, Heredity.

[10]  T. Chapman,et al.  Sex Peptide Causes Mating Costs in Female Drosophila melanogaster , 2005, Current Biology.

[11]  D. Begun,et al.  Molecular population genetics of female-expressed mating-induced serine proteases in Drosophila melanogaster. , 2007, Molecular biology and evolution.

[12]  U. Hengst,et al.  Male fertility defects in mice lacking the serine protease inhibitor protease nexin-1 , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[13]  H. Lilja,et al.  Secretory leucocyte protease inhibitor in the male genital tract: PSA-induced proteolytic processing in human semen and tissue localization. , 1995, Journal of andrology.

[14]  M. Wolfner,et al.  Fates and targets of male accessory gland proteins in mated female Drosophila melanogaster. , 2005, Insect biochemistry and molecular biology.

[15]  M. Wolfner,et al.  The Drosophila seminal fluid protein Acp26Aa stimulates release of oocytes by the ovary , 2000, Current Biology.

[16]  S. Lindquist,et al.  The FLP recombinase of yeast catalyzes site-specific recombination in the drosophila genome , 1989, Cell.

[17]  T. Iwamoto,et al.  Semenogelin, the main protein of semen coagulum, inhibits human sperm capacitation by interfering with the superoxide anion generated during this process. , 2001, Journal of andrology.

[18]  A. Clark,et al.  Variation in sperm displacement and its association with accessory gland protein loci in Drosophila melanogaster. , 1995, Genetics.

[19]  L. Partridge,et al.  Male identity and sperm displacement in Drosophila melanogaster , 1995 .

[20]  D. W. Rogers,et al.  A genome-wide analysis in Anopheles gambiae mosquitoes reveals 46 male accessory gland genes, possible modulators of female behavior , 2007, Proceedings of the National Academy of Sciences.

[21]  L. Partridge,et al.  Cost of mating in Drosophila melanogaster females is mediated by male accessory gland products , 1995, Nature.

[22]  U. Theopold,et al.  The coagulation of insect hemolymph , 2002, Cellular and Molecular Life Sciences CMLS.

[23]  D. Promislow,et al.  SPERM COMPETITIVE ABILITY AND GENETIC RELATEDNESS IN DROSOPHILA MELANOGASTER: SIMILARITY BREEDS CONTEMPT , 2002, Evolution; international journal of organic evolution.

[24]  A. Clark,et al.  Sperm competition and the maintenance of polymorphism , 2002, Heredity.

[25]  M. Wolfner,et al.  Mated Drosophila melanogaster females require a seminal fluid protein, Acp36DE, to store sperm efficiently. , 1999, Genetics.

[26]  A. Wong,et al.  Evolutionary Expressed Sequence Tag Analysis of Drosophila Female Reproductive Tracts Identifies Genes Subjected to Positive Selection , 2004, Genetics.

[27]  M. Wolfner,et al.  Drosophila seminal fluid proteins enter the circulatory system of the mated female fly by crossing the posterior vaginal wall. , 1999, Insect biochemistry and molecular biology.

[28]  A. Clark,et al.  Genes Regulated by Mating, Sperm, or Seminal Proteins in Mated Female Drosophila melanogaster , 2004, Current Biology.

[29]  Y. Rong,et al.  Gene targeting by homologous recombination in Drosophila. , 2000, Science.

[30]  M. Wolfner,et al.  Male and female cooperate in the prohormone-like processing of a Drosophila melanogaster seminal fluid protein. , 1995, Developmental biology.

[31]  M. Wolfner,et al.  Predicted seminal astacin-like protease is required for processing of reproductive proteins in Drosophila melanogaster , 2006, Proceedings of the National Academy of Sciences.

[32]  D. Curtis,et al.  Systematic generation of high-resolution deletion coverage of the Drosophila melanogaster genome , 2004, Nature Genetics.

[33]  M. Wolfner,et al.  An Ectopic Expression Screen Reveals the Protective and Toxic Effects of Drosophila Seminal Fluid Proteins , 2007, Genetics.

[34]  A. Clark,et al.  Female x male interactions in Drosophila sperm competition. , 1999, Science.

[35]  M. Wolfner,et al.  Comparative structural modeling and inference of conserved protein classes in Drosophila seminal fluid. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[36]  A. Civetta Direct visualization of sperm competition and sperm storage in Drosophila , 1999, Current Biology.

[37]  J. Coyne,et al.  Sperm competition between Drosophila males involves both displacement and incapacitation , 1999, Nature.

[38]  G. Rubin,et al.  Genetic transformation of Drosophila with transposable element vectors. , 1982, Science.

[39]  K. Golic,et al.  Gene Deletions by Ends-In Targeting in Drosophila melanogaster , 2004, Genetics.

[40]  A. Clark,et al.  Cross-Species Comparison of Drosophila Male Accessory Gland Protein Genes , 2005, Genetics.

[41]  L. Partridge,et al.  The role of male accessory gland protein Acp36DE in sperm competition in Drosophila melanogaster , 2000, Proceedings of the Royal Society of London. Series B: Biological Sciences.

[42]  A. Clark,et al.  Associations Between Sperm Competition and Natural Variation in Male Reproductive Genes on the Third Chromosome of Drosophila melanogaster , 2007, Genetics.

[43]  D. Cavener,et al.  Glucose dehydrogenase is required for normal sperm storage and utilization in female Drosophila melanogaster , 2004, Journal of Experimental Biology.

[44]  M. Wolfner,et al.  Probing the function of Drosophila melanogaster accessory glands by directed cell ablation. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[45]  M. Wolfner,et al.  Seminal influences: Drosophila Acps and the molecular interplay between males and females during reproduction. , 2007, Integrative and comparative biology.

[46]  M. Wolfner,et al.  Sustained Post-Mating Response in Drosophila melanogaster Requires Multiple Seminal Fluid Proteins , 2007, PLoS genetics.

[47]  M. Wolfner,et al.  A Drosophila seminal fluid protein, Acp26Aa, stimulates egg laying in females for 1 day after mating. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[48]  A. Ciereszko,et al.  Gelatinases and serine proteinase inhibitors of seminal plasma and the reproductive tract of turkey (Meleagris gallopavo). , 2005, Theriogenology.

[49]  M. Wolfner,et al.  Localization of the Drosophila male accessory gland protein Acp36DE in the mated female suggests a role in sperm storage. , 1996, Insect Biochemistry and Molecular Biology.

[50]  A. Clark,et al.  Evolutionary EST analysis identifies rapidly evolving male reproductive proteins in Drosophila , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[51]  M. Wolfner,et al.  The Drosophila melanogaster seminal fluid protein Acp62F is a protease inhibitor that is toxic upon ectopic expression. , 2002, Genetics.

[52]  J. Reichhart,et al.  The immune response of Drosophila melanogaster , 2004, Immunological reviews.

[53]  D. Begun,et al.  A genome-wide analysis of courting and mating responses in Drosophila melanogaster females. , 2004, Genome.