Massive dysregulation of genes involved in cell signaling and placental development in cloned cattle conceptus and maternal endometrium

Abstract

A major unresolved issue in the cloning of mammals by somatic cell nuclear transfer (SCNT) is the mechanism by which the process fails after embryos are transferred to the uterus of recipients before or during the implantation window. We investigated this problem by using RNA sequencing (RNA-seq) to compare the transcriptomes in cattle conceptuses produced by SCNT and artificial insemination (AI) at day (d) 18 (preimplantation) and d 34 (postimplantation) of gestation. In addition, endometrium was profiled to identify the communication pathways that might be affected by the presence of a cloned conceptus, ultimately leading to mortality before or during the implantation window. At d 18, the effects on the transcriptome associated with SCNT were massive, involving more than 5,000 differentially expressed genes (DEGs). Among them are 121 genes that have embryonic lethal phenotypes in mice, cause defects in trophoblast and placental development, and/or affect conceptus survival in mice. In endometria at d 18, <0.4% of expressed genes were affected by the presence of a cloned conceptus, whereas at d 34, ∼36% and <0.7% of genes were differentially expressed in intercaruncular and caruncular tissues, respectively. Functional analysis of DEGs in placental and endometrial tissues suggests a major disruption of signaling between the cloned conceptus and the endometrium, particularly the intercaruncular tissue. Our results support a "bottleneck" model for cloned conceptus survival during the periimplantation period determined by gene expression levels in extraembryonic tissues and the endometrial response to altered signaling from clones. somatic cell nuclear transfer | conceptus | placentation | conceptus-maternal communication I n cattle, as in other mammals, exquisitely orchestrated physiological changes of the conceptus and uterus are necessary for a successful pregnancy. Synchronization of the complex events at the time of implantation relies on the timed release of molecular signals from the conceptus and the endometrium. Embryo-derived IFN-τ (IFNT) is the major signal of pregnancy in cattle, preventing luteolysis and regulating the expression of genes that are responsible for promoting local changes in the endometrium to accommodate the conceptus (1-3). In females, progesterone is the major driver of endometrial changes that prepare the uterus for conceptus implantation (4, 5). In addition to IFNT and progesterone, signaling between the bovine conceptus and the endometrium is bidirectional, and involves several pathways that work concomitantly (6) for the successful establishment of pregnancy.Independent studies have shown that the majority of embryonic losses in cattle occur during the period that spans embryo cleavage until the attachment of the blastocyst to the endometrium (7). The reasons for these losses remain unclear and likely result from several factors, including embryonic lethal genes (8, 9), environmental stressors (7), and endometrial condition (10). Cloning of cattle by somatic cell nuclear transfer ...

Fine-tuned adaptation of embryo-endometrium pairs at implantation revealed by gene regulatory networks Tailored conceptus-maternal communication at implantation

“…For instance, there were twice as many highly positive (r > 0.95) and approximately half the number of highly negative (r < -0.95) co-expressing connections between extraembryonic and caruncular tissues compared to extraembryonic and intercaruncular tissues. These results greatly expand previous findings that the conceptus triggers distinct molecular responses in caruncular and intercaruncular tissues (6, 26, 41, 42).…”

“…We analyzed the RNA-sequencing data that consisted of samples collected from five cattle pregnancies terminated at gestation day 18 (GSE74152 (26)). The conceptus was dissected, and transcriptome data was generated for extraembryonic tissue; whereas the endometrium was dissected into caruncular (gland-free) and intercaruncular (containing endometrial glands) areas, and transcriptome data was generated from both regions of the endometrium (Fig 1A).…”

“…In mammals and particularly in the bovine species, a large body of gene expression data was produced at various steps of early pregnancy derived from in vitro or in vivo produced embryos (6, 26, 27, 39), varied physiological status of the dam (40), and fertility classified heifers (29). Altogether, results based on groups analyses (conceptus or endometrium) have demonstrated different degrees of interactions between the conceptus and endometrium at the initial phases of implantation.…”

“…The cross-talk between the conceptus and the endometrium is associated with the expression and regulation of a wealth of genes in each entity (24, 25). The nature of the conceptus modifies gene expression of the endometrium in cattle (6, 26, 27) and decidualizing human endometrial stromal cells (28). Similarly, the endometrium from dams with different fertility potentials (29) or metabolic status (30) influences the gene expression of the conceptus.…”

Rewiring of gene expression in circulating white blood cells is associated with pregnancy outcome in heifers (Bos taurus)

“…Sequences were submitted to a custom build bioinformatics pipeline 74,75,76 . Reads were aligned to the bovine genome (ARS-UCD1.2 obtained from Ensembl 77 ) using HISAT2 78 .…”