title: Retrotransposon expression and incorporation of cloned human and mouse retroelements in human spermatozoa.
abstract: To investigate the expression of long interspersed element (LINE) 1, human endogenous retrovirus (HERV) K10, and short interspersed element-VNTR-Alu element (SVA) retrotransposons in ejaculated human spermatozoa by means of reverse-transcription (RT) polymerase chain reaction (PCR) analysis as well as the potential incorporation of cloned human and mouse active retroelements in human sperm cell genome.
  
  
  
  Laboratory study.
  
  
  
  University research laboratories and academic hospital.
  
  
  
  Normozoospermic and oligozoospermic white men.
  
  
  
  RT-PCR analysis was performed to confirm the retrotransposon expression in human spermatozoa. Exogenous retroelements were tagged with a plasmid containing a green fluorescence (EGFP) retrotransposition cassette, and the de novo retrotransposition events were tested with the use of PCR, fluorescence-activated cell sorting analysis, and confocal microscopy.
  
  
  
  Retroelement expression in human spermatozoa, incorporation of cloned human and mouse active retroelements in human sperm genome, and de novo retrotransposition events in human spermatozoa.
  
  
  
  RT-PCR products of expressed human LINE-1, HERV-K10, and SVA retrotransposons were observed in ejaculated human sperm samples. The incubation of human spermatozoa with either retrotransposition-active human LINE-1 and HERV-K10 or mouse reverse transcriptase-deficient VL30 retrotransposons tagged with an EGFP-based retrotransposition cassette led to EGFP-positive spermatozo; 16.67% of the samples were positive for retrotransposition. The respective retrotransposition frequencies for the LINE-1, HERV-K10, and VL30 retrotransposons in the positive samples were 0.34 ± 0.13%, 0.37 ± 0.17%, and 0.30 ± 0.14% per sample of 10,000 spermatozoa.
  
  
  
  Our results show that: 1) LINE-1, HERV-K10, and SVA retrotransposons are transcriptionally expressed in human spermatozoa; 2) cloned active retroelements of human and mammalian origin can be incorporated in human sperm genome; 3) active reverse transcriptases exist in human spermatozoa; and 4) de novo retrotransposition events occur in human spermatozoa.
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece; Medical Genetics and Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Ioannina University Hospital, Ioannina, Greece; Genesis-Genoma Lab, Chalandri-Athens, Greece.
authors.forename: Leandros
authors.initials: L
authors.lastname: Lazaros
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Chrysoula
authors.initials: C
authors.lastname: Kitsou
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Charilaos
authors.initials: C
authors.lastname: Kostoulas
authors.affiliation: Foundation for Research & Technology-Hellas Institute of Molecular Biology and Biotechnology, Department of Biomedical Research, Ioannina, Greece.
authors.forename: Sofia
authors.initials: S
authors.lastname: Bellou
authors.affiliation: Medical Genetics and Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Ioannina University Hospital, Ioannina, Greece.
authors.forename: Elissavet
authors.initials: E
authors.lastname: Hatzi
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Paris
authors.initials: P
authors.lastname: Ladias
authors.affiliation: Medical Genetics and Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Ioannina University Hospital, Ioannina, Greece.
authors.forename: Theodoros
authors.initials: T
authors.lastname: Stefos
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Sofia
authors.initials: S
authors.lastname: Markoula
authors.affiliation: Department of Anatomy-Histology-Embryology, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Vasiliki
authors.initials: V
authors.lastname: Galani
authors.affiliation: Hematology Laboratory, Molecular Biology Unit, Ioannina University Hospital, Ioannina, Greece.
authors.forename: Georgios
authors.initials: G
authors.lastname: Vartholomatos
authors.affiliation: Department of General Biology, Medical School, Ioannina University, Ioannina, Greece.
authors.forename: Theodore
authors.initials: T
authors.lastname: Tzavaras
authors.affiliation: Laboratory of Medical Genetics of Human Reproduction, Medical School, Ioannina University, Ioannina, Greece; Medical Genetics and Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Ioannina University Hospital, Ioannina, Greece. Electronic address: igeorgio@uoi.gr.
authors.forename: Ioannis
authors.initials: I
authors.lastname: Georgiou
mesh_terms: D000818:Animals; D002469:Cell Separation / Q000379:methods; D003001:Cloning, Molecular; D010450:Endopeptidases / Q000096:biosynthesis / Q000235:genetics*; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D020084:Long Interspersed Nucleotide Elements; D008297:Male; D051379:Mice; D018613:Microscopy, Confocal; D018598:Minisatellite Repeats; D009845:Oligospermia / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013094:Spermatozoa / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D000085242:Viral Proteases
chemical_list: C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D010450:Endopeptidases; D000085242:Viral Proteases; C429524:human endogenous retrovirus K10 protease
keywords: HERV-K10; LINE-1; human spermatozoa; retroelements; retrotransposition