Combinational electroporation and transplantation approach to studying gene functions in avian embryos

Gene transfection is an indispensable approach for studying gene function since it provides important information on gain-and/or loss-of-function.Chick embryos are also extensively employed for studying biological function since they are easily accessible and can be maintained alive after manipulation.The combination of both techniques presents a powerful approach to understanding how genes regulate embryo development.Furthermore,combining these approaches with tissue transplant techniques make even more attractive for elucidate gene function.Electroporation,employing parallelly fashioned electrodes,has been widely used in chick embryos.However,experimenters have been frustrated by unsuccessfully transfection in some embryonic tissue of interest because the electrodes were improperly positioned.We presently demonstrated the different patterns of organizing and positioning the electrodes,in combination with tissue transplantation,to efficiently and specifically transfect the chick embryonic head,trunk neural tube,heart tube,somites and neural crest cells with the GFP reporter gene.

Protein O-GlcNAcylation homeostasis regulates facultative heterochromatin to fine-tune sog-Dpp signaling during Drosophila early embryogenesis

Protein O-GlcNAcylation is a monosaccharide post-translational modification maintained by two evolutionarily conserved enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Mutations in human OGT have recently been associated with neurodevelopmental disorders, although the mechanisms linking O-GlcNAc homeostasis to neurodevelopment are not understood. Here, we investigate the effects of perturbing protein O-GlcNAcylation using transgenic Drosophila lines that overexpress a highly active OGA. We reveal that temporal reduction of protein O-GlcNAcylation in early embryos leads to reduced brain size and olfactory learning in adult Drosophila. Downregulation of O-GlcNAcylation induced by the exogenous OGA activity promotes nuclear foci formation of Polycomb-group protein Polyhomeotic and the accumulation of excess K27 trimethylation of histone H3 (H3K27me3) at the mid-blastula transition. These changes interfere with the zygotic expression of several neurodevelopmental genes, particularly short gastrulation (sog), a component of an evolutionarily conserved sog-Decapentaplegic (Dpp) signaling system required for neuroectoderm specification. Our findings highlight the importance of early embryonic O-GlcNAcylation homeostasis for the fidelity of facultative heterochromatin redeployment and initial cell fate commitment of neuronal lineages, suggesting a possible mechanism underpinning OGT-associated intellectual disability.

Endoderm contributes to endocardial composition during cardiogenesis

Heart formation commences from a single heart tube,which fuses from bilateral primordial heart fields.The developing heart tube is composed of outer-layer myocardial cells and inner-layer endocardial cells.Several distinct populations of precardiac cells contribute to cardiac morphogenesis.However,it still remains not very clear about the lineage of endocardium at gastrulation stage.Thereby,this study focused on ascertaining the correlation between the hypoblast in gastrulation and endocardium during cardiogenesis.Firstly,the fusing heart tube morphologically is closed to endoderm-derived pharynx floor,implying the possibility that pharynx floor might be wrapped into the formation of endoderm.Secondly,HNK1 is expressed in hypoblast strongly at gastrula stage and subsequently appeared in endocardium of cardiogenesis.Moreover,fate map data displayed that DiI labeled hypoblast was also present in endocardium later on.One more evidence is chick-quail chimera of hypoblast transplantation,in which quail-hypoblast derivative could be identified inendocardium of cardiogenesis by QCPN antibody.In sum,our current data suggests that endoderm in gastrula contribute at least partly to the formation of endocardium of cardiogenesis.