Retrotransposon RNA expression and evidence for retrotransposition events in human oocytes.
Although human diseases of retrotransposition-derived etiology have been documented, retrotransposon RNA expression and the occurrence of retrotransposition events in the human oocyte are not studied. We investigated the RNA expression of L1 and HERV-K10 retrotransposons in human oocytes by RT-PCR analysis with designed primers. Using denucleated germinal vesicles (GVs), we detected RT-PCR products of expressed L1, HERV-K10 and, unexpectedly, SINE-R, VNTR and Alu (SVA) retrotransposons. Their transcript specificities were identified as such following RNA-FISH and their origin by cloning and sequence alignment analyses. Assessing the expression level in comparison with somatic cells by densitometry analysis, we found that although in normal lymphocytes and transformed HeLa cells their profile was in an order of L1 > HERV-K10 > SVA, remarkably this was reversed in oocytes. To investigate whether de novo retrotransposition events occur and reverse transcriptases are expressed in the human oocyte, we introduced in GVs either a retrotransposition active human L1 or mouse reverse transcriptase deficient-VL30 retrotransposon tagged with an EGFP-based retrotransposition cassette. Interestingly, in both the cases, we observed EGFP-positive oocytes, associated with an abnormal morphology for L1 and granulation for VL30, and the retrotransposition events were confirmed by PCR. Our results: (i) show that L1, HERV-K10 and SVA retrotransposons are transcriptionally expressed and (ii) provide evidence, for the first time, for retrotransposition events occurring in the human oocyte. These findings suggest that both, network of retrotransposon transcripts and controlled retrotranspositions, might serve important functions required for oocyte development and fertilization while the uncontrolled ones might explain the onset of genetic disorders.
Georgiou
Ioannis
I
Laboratory of Reproductive Genetics, Medical School, University of Ioannina, Ioannina, Greece.
Noutsopoulos
Dimitrios
D
Dimitriadou
Eftychia
E
Markopoulos
Georgios
G
Apergi
Anastasia
A
Lazaros
Leandros
L
Vaxevanoglou
Terpsi
T
Pantos
Kostas
K
Syrrou
Maria
M
Tzavaras
Theodore
T
D000818:Animals; D001483:Base Sequence; D005786:Gene Expression Regulation; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D009865:Oocytes / Q000166:cytology / Q000378:metabolism*; D012313:RNA / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018626:Retroelements / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
C034477:RNA primers; D012333:RNA, Messenger; D018626:Retroelements; D012313:RNA