Changes in sub-cellular localisation of trophoblast and inner cell mass specific transcription factors during bovine preimplantation development

Abstract

BackgroundPreimplantation bovine development is emerging as an attractive experimental model, yet little is known about the mechanisms underlying trophoblast (TE)/inner cell mass (ICM) segregation in cattle. To gain an insight into these processes we have studied protein and mRNA distribution during the crucial stages of bovine development. Protein distribution of lineage specific markers OCT4, NANOG, CDX2 were analysed in 5-cell, 8–16 cell, morula and blastocyst stage embryos. ICM/TE mRNA levels were compared in hatched blastocysts and included: OCT4, NANOG, FN-1, KLF4, c-MYC, REX1, CDX2, KRT-18 and GATA6.ResultsAt the mRNA level the observed distribution patterns agree with the mouse model. CDX2 and OCT4 proteins were first detected in 5-cell stage embryos. NANOG appeared at the morula stage and was located in the cytoplasm forming characteristic rings around the nuclei. Changes in sub-cellular localisation of OCT4, NANOG and CDX2 were noted from the 8–16 cell onwards. CDX2 initially co-localised with OCT4, but at the blastocyst stage a clear lineage segregation could be observed. Interestingly, we have observed in a small proportion of embryos (2%) that CDX2 immunolabelling overlapped with mitotic chromosomes.ConclusionsCell fate specification in cattle become evident earlier than presently anticipated – around the time of bovine embryonic genome activation. There is an intriguing possibility that for proper lineage determination certain transcription factors (such as CDX2) may need to occupy specific regions of chromatin prior to its activation in the interphase nucleus. Our observation suggests a possible role of CDX2 in the process of epigenetic regulation of embryonic cell fate.

CDX2 is essential for cell proliferation and polarity in porcine blastocysts

“…In contrast, porcine CDX2 protein is initially observed in some TE cells right after cavitation at E5 and remains exclusively in the TE thereafter. Interestingly, the time point for porcine CDX2 protein expression is very similar to that in bovine (Goissis and Cibelli, 2014;Madeja et al, 2013) and human (Niakan and Eggan, 2013) embryos, in which CDX2 protein becomes detectable after blastocyst formation. Similarly, prolonged OCT4 expression in the TE is also observed in human (Cauffman et al, 2005) and bovine (van Eijk et al, 1999) embryos.…”

“…Different time points for CDX2 accumulation in mice and other mammals might account for the different phenotypes of CDX2-depleted mouse and pig embryos. Therefore, the results from porcine studies may be more pertinent to our understanding of CDX2 function in embryonic development in large animals, including cows and primates, which also show the same temporal pattern of CDX2 accumulation (Goissis and Cibelli, 2014;Madeja et al, 2013;Niakan and Eggan, 2013). However, more studies in various strains are needed to clarify the functional significance of CDX2 in mouse cleavage-stage embryos (Wu and Schöler, 2011).…”

“…In mice, both Cdx2-knockout and -knockdown blastocysts exhibit a hatching failure, decreased cell number, abnormal TE cell junctions/polarity and a disturbed TE GRN (Blij et al, 2012;Jedrusik et al, 2010Jedrusik et al, , 2015Nishioka et al, 2008;Ralston and Rossant, 2008;Strumpf et al, 2005;Wu et al, 2010). In primates (Sritanaudomchai et al, 2009) and cattle (Goissis and Cibelli, 2014;Madeja et al, 2013), CDX2 knockdown results in a hatching failure and deficient cell proliferation in blastocysts. Consistently, our study also showed that CDX2 knockdown compromises TE cell polarization and junctions.…”

Bone Morphogenetic Proteins in Preimplantation Embryos

“…Subsequent cell divisions lead to symmetric or asymmetric distribution of polarity information, based on the angle of cell division. Polar and apolar cells would then differ in their developmental potential (Lee, Choi, et al, 2014;Madeja et al, 2013;Yamanaka et al, 2006). In farm animals, i.e., cows and pigs, the "inside-outside" hypothesis is not fulfilled as morula compaction, formation of tight junctions in the outer cells and ICM allocation occur independently from each other (Kirchhof et al, 2000;Van Soom et al, 1997).…”

Lessons learned from functional assessment of pluripotency-associated transcription factors during early embryogenesis and embryonic stem cells

“…Both CMYC and NMYC show similar expression patterns during mouse preimplantation development and are functionally redundant for pluripotency (Smith et al, 2010). The expression of CMYC has also been analyzed in preimplantation embryos of ruminants species (Miles et al, 2012;Madeja et al, 2013;Singh et al, 2014;Silva et al, 2017), but its functional roles in such species remain to be described. Dosage-sensitive sex reversal, adrenal hypoplasia critical region, chromosome X, gene 1 (DAX1) DAX1 (NR0B1) was detected in oocytes and during all embryonic development of several species (Clipsham et al, 2004;Stickels et al, 2015;Moura et al, 2017;Silva et al, 2017).…”