The Dual-Pseudotyped Lentiviral Vector with VSV-G and Sendai Virus HN Enhances Infection Efficiency through the Synergistic Effect of the Envelope Proteins.

A gene delivery system utilizing lentiviral vectors (LVs) requires high transduction efficiency for successful application in human gene therapy. Pseudotyping allows viral tropism to be expanded, widening the usage of LVs. While vesicular stomatitis virus G (VSV-G) single-pseudotyped LVs are commonly used, dual-pseudotyping is less frequently employed because of its increased complexity. In this study, we examined the potential of phenotypically mixed heterologous dual-pseudotyped LVs with VSV-G and Sendai virus hemagglutinin-neuraminidase (SeV-HN) glycoproteins, termed V/HN-LV. Our findings demonstrated the significantly improved transduction efficiency of V/HN-LV in various cell lines of mice, cynomolgus monkeys, and humans compared with LV pseudotyped with VSV-G alone. Notably, V/HN-LV showed higher transduction efficiency in human cells, including hematopoietic stem cells. The efficient incorporation of wild-type SeV-HN into V/HN-LV depended on VSV-G. SeV-HN removed sialic acid from VSV-G, and the desialylation of VSV-G increased V/HN-LV infectivity. Furthermore, V/HN-LV acquired the ability to recognize sialic acid, particularly N-acetylneuraminic acid on the host cell, enhancing LV infectivity. Overall, VSV-G and SeV-HN synergistically improve LV transduction efficiency and broaden its tropism, indicating their potential use in gene delivery.

Essential Roles of Exocyst Complex Component 3-like 2 on Cardiovascular Development in Mice.

Angiogenesis is a process to generate new blood vessels from pre-existing vessels and to maintain vessels, and plays critical roles in normal development and disease. However, the molecular mechanisms underlying angiogenesis are not fully understood. This study examined the roles of exocyst complex component (Exoc) 3-like 2 (Exoc3l2) during development in mice. We found that Exoc3l1, Exoc3l2, Exoc3l3 and Exoc3l4 are expressed abundantly in endothelial cells at embryonic day 8.5. The generation of Exoc3l2 knock-out (KO) mice showed that disruption of Exoc3l2 resulted in lethal in utero. Substantial numbers of Exoc3l2 KO embryos exhibited hemorrhaging. Deletion of Exoc3l2 using Tie2-Cre transgenic mice demonstrated that Exoc3l2 in hematopoietic and endothelial lineages was responsible for the phenotype. Taken together, these findings reveal that Exoc3l2 is essential for cardiovascular and brain development in mice.

Esrrb function is required for proper primordial germ cell development in presomite stage mouse embryos.

Estrogen related receptor beta (Esrrb) is an orphan nuclear receptor that is required for self-renewal and pluripotency in mouse embryonic stem (ES) cells. However, in the early post-implantation mouse embryo, Esrrb is specifically expressed in the extraembryonic ectoderm (ExE) and plays a crucial role in trophoblast development. Previous studies showed that Esrrb is also required to maintain trophoblast stem (TS) cells, the in vitro stem cell model of the early trophoblast lineage. In order to identify regulatory targets of Esrrb in vivo, we performed microarray analysis of Esrrb-null versus wild-type post-implantation ExE, and identified 30 genes down-regulated in Esrrb-mutants. Among them is Bmp4, which is produced by the ExE and known to be critical for primordial germ cell (PGC) specification in vivo. We further identified an enhancer region bound by Esrrb at the Bmp4 locus by performing Esrrb ChIP-seq and luciferase reporter assay using TS cells. Finally, we established a knockout mouse line in which the enhancer region was deleted using CRISPR/Cas9 technology. Both Esrrb-null embryos and enhancer knockout embryos expressed lower levels of Bmp4 in the ExE, and had reduced numbers of PGCs. These results suggested that Esrrb functions as an upstream factor of Bmp4 in the ExE, regulating proper PGC development in mice.