112 Identification of Solute Carrier Family 5 Member 9 (SLC5A9) as Biomarker of In Vivo and In Vitro Oocyte Developmental Competence

Abstract

The transcriptome associated with the ability of oocytes to support embryogenesis and development remains unclear. The aim of this study was to compare the mRNA expression pattern between oocytes collected from fertile heifers (H) and repeat breeders (RB) using RNAseq. Oocytes were collected by ovum pick-up from Italian Simmental (IS) and Holstein Friesian (HF) heifers that became pregnant at the following oestrus, and from cows of the same breeds classified as RB after they failed to become pregnant for a minimum of 3 consecutive AI. Oocytes from each animal (5 for HF_RB, 3 each for all other groups) were pooled and sequenced using Illumina HiSEqn 2000 (Illumina Inc., San Diego, CA, USA). Principal component analysis showed that HF oocytes clustered away from those of IS, irrespective of the fertility status. However, within breeds, H and RB always clustered apart from each other. This suggests that differences in the transcriptional blueprint between breeds was more prominent than differences related to fertility. However, these were still strong when considering breeds separately, therefore we treated breeds as independent groups and performed differential expression analysis on H v. RB in IS and HF. DESEqn 2 identified 135 differentially expressed genes when we compared H and RB in IS and 2124 in HF. This difference in number of DE genes between the 2 breeds reflects the fact that the variance between H and RB transcriptional patterns in HF is larger than in IS. We looked for genes that were differentially and consistently expressed between H and RB in both breeds and identified 10 genes: histone H2B, exo/endonuclease G, solute carrier family 5 member 9, BH3 interacting domain death agonist, TBK1 binding protein 1, purine-rich element binding protein A, tRNA-yW synthesising protein 1 homologue, Jun oncogene, FRA10AC1 homologue, and SKI oncogene. To validate their potential use as biomarkers, we performed a preliminary in vitro validation by RT-qPCR. As described in the literature, oocytes collected from ovaries with more than 10 follicles of 2 to 6 mm were classified as high quality, while those collected from ovaries with fewer than 10 follicles were classified as low quality (Gandolfi et al. 1997 Theriogenology 48, 1153-1160). Gene expression analysis confirmed SLC5A9, a membrane-bound protein involved in sodium-dependent transport of glucose, as a potential biomarker to discriminate between a competent and incompetent oocyte. Altogether, our data provide a detailed overview of the mRNA pattern defining the differences between a competent and incompetent oocyte in vivo, and identified a new potential marker able to differentiate fertile and infertile oocyte both in vivo and in vitro.

Study supported by PRIN 08–09, EU-Quantomics no. 222664 and Carraresi Foundation.