DAZL And BOULE Promote Primordial Germ Cells Differentiation

According to the World Health Organization (WHO) assessment, infertility has been a threat to about 15 percent of reproductive age couples in the world, which means that one of every six couples undergo reproductive disorders. In our country, recent survey of infertility accounted for 10 percent of married couples. What’s more, as recent research data showed, the rate of increase is build-up. There are many reasons in male sterility disease, mainly two reasons involving the formation of germ cells and abnormal spermatogenesis. Until this year, clinical treatment for infertility major resort to assisted reproduction resolved, but for patients with certain disorders of sperm, it is still no way to help them get healthy baby. However, differentiation from pluripotency stem cells to germ cells in vitro may be an effective way to solve this problem after somatic reprogramming to iPSCs.Primordial germ cells (PGCs), the precursors of germ cells both males and females, represents a lineage characteristics. In a pluripotent state, PGCs have the ability to differentiate into sperm and egg cells. And what’s more, primordial germ cells can regain similar to embryonic germ cells (EGCs), which is in pluripotent state. Bone morphogenetic protein (BMP) from extraembryonic tissue to the proximal epiblast results in the specification of germ cells lineage at embryonic day 6.25 (E 6.25), by assigning a few epiblast cells to develop into PGCs. During PGCs development process, there are two main events, the first one is the change of epigenetic characteristics, including global DNA demethylation and methylation, imprinting erasure and re-established, the second is the re-expression of pluripotent gene, including Oct4, Sox2 and Nanog. Altogether, the important target for primordial germ cells is not only for clinical treatment but also for epigenetic research and other basic research.Currently, the study of international and domestic of PGCs has made remarkable achievements. Recently, Mitinori Saitou and his colleague have established a Blimp1-mVenus (BV) and Stella-ECFP (SC) double-labeled cell line, which can indicate sensitively the development of primordial germ cells from epiblast cells, and then primordial germ cell like cells were induced after six day’s culture BMP4 and BMP8a, further differentiation of haploid germ cells were get and then they had got mouse offspring through ICSI (intracytoplasmic sperm injection). On this basis, through overexpress Blimp1,Prdm14 and AP2γ, three key genes during PGCs differentiation from epiblast, they also get PGCLCs (PGC-like cells) and healthy offspring. However, the scarcity of primordial germ cells in the developing mammalian embryo hampers the study of germ cell development and the potential application in assisted reproduction.DAZ gene family, including DAZ, DAZL and BOULE, is a gene family which is specific expression in germ line. In mouse genome, there are only two gene, DAZL and BOULE, located in autosome. In the protein structure, DAZ gene family have the same conformation, including a RRM (RNA recognition motif) and at least one DAZ motif contains 24 amino acid residues. DAZ gene family binding 3’UTR of mRNA to active its expression. As natural deletions and knock-out experiments show, DAZL and BOULE are essential for germ cell differentiation.On the basic of integration of Blimp1, Prdm14 and AP2γ, contains Blimp1-mVenus and Stella-ECFP reporter gene, we established a stable mouse embryonic stem cell line which integrated both DAZL and BOULE genes. Through differentiation in vitro, we found that overexpression can promote PGCLCs specification, and also have a tendency to upregulated key genes, Blimp1 and Stella, expression. Altogether, DAZL and BOULE promote PGCLCs differentiation through upregulated key genes expression.