The transcriptome of human oocytes

Abstract

The identification of genes and deduced pathways from the mature human oocyte can help us better understand oogenesis, folliculogenesis, fertilization, and embryonic development. Human metaphase II oocytes were used within minutes after removal from the ovary, and its transcriptome was compared with a reference sample consisting of a mixture of total RNA from 10 different normal human tissues not including the ovary. RNA amplification was performed by using a unique protocol. Affymetrix Human Genome U133 Plus 2.0 GeneChip arrays were used for hybridizations. Compared with reference samples, there were 5,331 transcripts significantly up-regulated and 7,074 transcripts significantly down-regulated in the oocyte. Of the oocyte up-regulated probe sets, 1,430 have unknown function. A core group of 66 transcripts was identified by intersecting significantly up-regulated genes of the human oocyte with those from the mouse oocyte and from human and mouse embryonic stem cells. GeneChip array results were validated using RT-PCR in a selected set of oocyte-specific genes. Within the up-regulated probe sets, the top overrepresented categories were related to RNA and protein metabolism, followed by DNA metabolism and chromatin modification. This report provides a comprehensive expression baseline of genes expressed in in vivo matured human oocytes. Further understanding of the biological role of these genes may expand our knowledge on meiotic cell cycle, fertilization, chromatin remodeling, lineage commitment, pluripotency, tissue regeneration, and morphogenesis.

Fig. 1.

Summary of CRL RNA amplification protocol. (A) Flow Chart of the CRL amplification protocol. (B) Representative plot of gene intensities comparing the CRL and Ambion amplification methods using 20 ng and 1 μg of total RNA, respectively.

Fig. 2.

RT–PCR verification of the GeneChip array result. Loading orders of the gel were as following: M, 100 bp molecular weight standards with sizes as indicated on the left margin; OCT4, POU domain, class 5, transcription factor 1; STELLA, DPPA3, developmental pluripotency-associated 3; ESG1, embryonal stem cell-specific gene 1; VASA, DEAD box RNA helicase; GDF9, growth differentiation factor 9; ZP1, zona pellucida glycoprotein 1; H1FOO, H1 histone family, member O, oocyte-specific; CDH3, cadherin 3, type 1, P-cadherin (placental); TUBB4Q, β-tubulin; ACTB, β-actin; and negative control with no DNA template.

Fig. 3.

TGF-β signaling pathway. Genes shown in red are differentially up-regulated in human oocytes.

Fig. 4.

Venn diagrams showing the intersection between differentially up-regulated genes in the human (HU OC) and mouse oocytes (MO OC) (1,587 transcripts were found to be in common in both species) (A); HU OC and hESCs (388 transcripts were found to be common in both cell types) (B); MO OC and mESCs (591 transcripts were found to be common in both cell types) (C); and HU OC/hESC and MO OC/mESC (78 transcripts were found to be common in all four cell types (D).