Daniels, Robert; (1998) Gene expression in human preimplantation embryos. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Gene transcription during early embryonic development of the human has been investigated using single cell RT-PCR techniques for several house-keeping genes (APRT, adenosine phosphoribosyl transferase; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HPRT, hypoxanthine phosphorybosyl transferase) and tissue-specific genes (α-globin; β-globin; MPK, myotonin protein kinase; SNRPN, small nuclear ribonucleoprotein polypeptide N; WNT5B and WNT11). Analysis of the CpG content of the genes studied so far in human preimplantation development reveals a positive correlation between the presence of a CpG island in a gene and its expression in early embryos. The expression and methylation of the gene, XIST (X-Inactive Specific Transcript), which plays an important role in X-inactivation in female mammals, has been further studied. In mice, the Xist gene is imprinted in that it is expressed from the paternal allele in preimplantation development. However, in the human, XIST transcripts are present in male (maternal X only) as well as female preimplantation embryos at the 8-cell stage, suggesting that early XIST transcription does not initiate and regulate the onset of X-inactivation. Paternal X-inactivation may therefore correlate with a gametic XIST DNA imprint perpetuated to the blastocyst stage. It is possible that differential methylation may be the basis of imprinted X-inactivation in the human as in the mouse. Using methylation-sensitive-restriction-enzyme PCR, three differentially methylated CpG sites in the promoter region of the human XIST gene have been identified in human somatic cells. Results so far indicate that these sites are unmethylated in both human sperm and oocytes and the imprint mechanism remains unknown. In order to facilitate further research on gene expression in early human development, cDNA libraries have been created from single human preimplantation embryos. These libraries will enable rapid analysis of the stage-specific expression of a wide range of genes without requiring use of human preimplantation embryos themselves.

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