PATERNAL TRANSCRIPTS FOR GLUCOSE-6-PHOSPHATE-DEHYDROGENASE AND ADENOSINE-DEAMINASE ARE FIRST DETECTABLE IN THE HUMAN PREIMPLANTATION EMBRYOAT THE 3-CELL TO 4-CELL STAGE

The transition between dependence on maternal transcripts and proteins inherited in the oocyte and embryonic gene expression in the human pr eimplantation embryo occurs at the four-to eight-cell stage. Recently, studies using reverse transcriptase polymerase chain reaction (RT-PCR ) have detected paternal transcripts for the Y-linked genes, ZFY and S RY, and the myotonic dystrophy associated protein kinase gene, DK, as early as the late pronucleate one-cell stage. However, expression at t he protein level has not been demonstrated and its function at these e arly stages is unknown. Using coding sequence polymorphisms to disting uish maternal and paternal transcripts, we have examined the transcrip tion of two ubiquitously expressed genes: X-linked glucose-6-phosphate dehydrogenase (G6PD) and adenosine deaminase (ADA). Both G6PD and ADA are housekeeping genes with TATA-less promoters which, because of the ir roles in metabolism and ubiquitous expression, may provide a more r eliable indication of the timing of activation of the embryonic genome . They also each have biallelic polymorphisms with a high heterozygosi ty ratio which can be detected by restriction digestion. Couples under going in vitro fertilization (IVF) were screened for these polymorphis ms. Individual spare oocytes and embryos at different stages of preimp lantation development were analyzed by RT-PCR and appropriate restrict ion digestion in those cases in which the male partner carried a diffe rent allele to the female partner. In addition, since only female embr yos inherit the paternal allele of X-linked G6PD, cDNA was also analyz ed for ZFX/ZFY transcripts to identify the sex of each embryo. One hun dred and twenty three individual oocytes and embryos were analyzed by RT-PCR and restriction digestion to detect the paternal transcripts fr om the polymorphic alleles. Maternal transcripts for G6PD, ADA, and ZF X were detected in all oocytes and embryos and at all stages. Followin g restriction digestion, paternal G6PD and ZFY transcripts were first detected at the four-cell stage and paternal ADA transcripts in an emb ryo at the three-cell stage coinciding with the onset of dependency on transcription from the embryonic genome. This approach should be wide ly applicable to other genes since similar polymorphisms exist in the coding regions of many genes. (C) 1997 Wiley-Liss, Inc.