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Flow cytometric sex sorting affects CD4 membrane distribution and binding of exogenous DNA on bovine sperm cells

Published online by Cambridge University Press:  13 July 2017

William Borges Domingues ,
Tony Leandro Rezende da Silveira ,
Eliza Rossi Komninou ,
Leonardo Garcia Monte ,
Mariana Härter Remião ,
Odir Antônio Dellagostin ,
Carine Dahl Corcini ,
Antônio Sergio Varela Junior ,
Fabiana Kömmling Seixas  and
Tiago Collares
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William Borges Domingues
Affiliation:
Laboratório de Genômica Estrutural, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Tony Leandro Rezende da Silveira
Affiliation:
Laboratório de Genômica Estrutural, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Eliza Rossi Komninou
Affiliation:
Laboratório de Biotecnologia do Cancer, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Leonardo Garcia Monte
Affiliation:
Laboratório de Imunodiagnóstico, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Mariana Härter Remião
Affiliation:
Laboratório de Biotecnologia do Cancer, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Odir Antônio Dellagostin
Affiliation:
Laboratório de Vacinologia, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Carine Dahl Corcini
Affiliation:
Reprodução Animal Comprada, Instituto de Ciências Biológicas, Universidade Federal de Rio Grande, RS, Brasil.
Antônio Sergio Varela Junior
Affiliation:
Reprodução Animal Comprada, Instituto de Ciências Biológicas, Universidade Federal de Rio Grande, RS, Brasil.
Fabiana Kömmling Seixas
Affiliation:
Laboratório de Biotecnologia do Cancer, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Tiago Collares
Affiliation:
Laboratório de Biotecnologia do Cancer, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brasil.
Vinicius Farias Campos*
Affiliation:
Laboratório de Genômica Estrutural, Centro de Desenvolvimento Tecnológico, Campus Universitário Capão do Leão s/n°, CEP 96160–000, Capão do Leão, RS, Brasil.
*
All correspondence to: Vinicius Farias Campos, Laboratório de Genômica Estrutural, Centro de Desenvolvimento Tecnológico, Campus Universitário Capão do Leão s/n°, CEP 96160–000, Capão do Leão, RS, Brasil. Tel: +55 53 3275 7350. E-mail: fariascampos@gmail.com
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Summary

Bovine sex-sorted sperm have been commercialized and successfully used for the production of transgenic embryos of the desired sex through the sperm-mediated gene transfer (SMGT) technique. However, sex-sorted sperm show a reduced ability to internalize exogenous DNA. The interaction between sperm cells and the exogenous DNA has been reported in other species to be a CD4-like molecule-dependent process. The flow cytometry-based sex-sorting process subjects the spermatozoa to different stresses causing changes in the cell membrane. The aim of this study was to elucidate the relationship between the redistribution of CD4-like molecules and binding of exogenous DNA to sex-sorted bovine sperm. In the first set of experiments, the membrane phospholipid disorder and the redistribution of the CD4 were evaluated. The second set of experiments was conducted to investigate the effect of CD4 redistribution on the mechanism of binding of exogenous DNA to sperm cells and the efficiency of lipofection in sex-sorted bovine sperm. Sex-sorting procedure increased the membrane phospholipid disorder and induced the redistribution of CD4-like molecules. Both X-sorted and Y-sorted sperm had decreased DNA bound to membrane in comparison with the unsorted sperm; however, the binding of the exogenous DNA was significantly increased with the addition of liposomes. Moreover, we demonstrated that the number of sperm-bound exogenous DNA was decreased when these cells were preincubated with anti-bovine CD4 monoclonal antibody, supporting our hypothesis that CD4-like molecules indeed play a crucial role in the process of exogenous DNA/bovine sperm cells interaction.

Keywords

BindingProtein redistributionSMGTSpermatozoaTransfection
Type
Research Article
Information
Zygote , Volume 25 , Issue 4 , August 2017 , pp. 519 - 528
Copyright
Copyright © Cambridge University Press 2017 

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