Xue-Shan Ma

Atomic-scale mapping of dipole frustration at 90 degrees charged domain walls in ferroelectric PbTiO3 films

The atomic-scale structural and electric parameters of the 90° domain-walls in tetragonal ferroelectrics are of technological importance for exploring the ferroelectric switching behaviors and various domain-wall-related novel functions. We have grown epitaxial PbTiO3/SrTiO3 multilayer films in which the electric dipoles at 90° domain-walls of ferroelectric PbTiO3 are characterized by means of aberration-corrected scanning transmission electron microscopy. Besides the well-accepted head-to-tail 90° uncharged domain-walls, we have identified not only head-to-head positively charged but also tail-to-tail negatively charged domain-walls. The widths, polarization distributions, and strains across these charged domain-walls are mapped quantitatively at atomic scale, where remarkable difference between these domain-walls is presented. This study is expected to provide fundamental information for understanding numerous novel domain-wall phenomena in ferroelectrics.

Different fates of oocytes with DNA double-strand breaks in vitro and in vivo

In female mice, despite the presence of slight DNA double-strand breaks (DSBs), fully grown oocytes are able to undergo meiosis resumption as indicated by germinal vesicle breakdown (GVBD); however, severe DNA DSBs do reduce and delay entry into M phase through activation of the DNA damage checkpoint. But little is known about the effect of severe DNA DSBs on the spindle assembly checkpoint (SAC) during oocyte maturation. We showed that nearly no first polar body (PB1) was extruded at 12 h of in vitro maturation (IVM) in severe DNA DSBs oocytes, and the limited number of oocytes with PB1 were actually at telophase. However, about 60% of the severe DNA DSBs oocytes which underwent GVBD at 2 h of IVM released a PB1 at 18 h of IVM and these oocytes did reach the second metaphase (MII) stage. Chromosome spread at MI and MII stages showed that chromosomes fragmented after GVBD in severe DNA DSBs oocytes. The delayed PB1 extrusion was due to the disrupted attachment of microtubules to kinetochores and activation of the SAC. At the same time, misaligned chromosome fragments became obvious at the first metaphase (MI) in severe DNA DSBs oocytes. These data implied that the inactivation of SAC during the metaphase-anaphase transition of first meiosis was independent of chromosome integrity. Next, we induced DNA DSBs in vivo, and found that the number of superovulated oocytes per mouse was significantly reduced; moreover, this treatment increased the percentage of apoptotic oocytes. These results suggest that DNA DSBs oocytes undergo apoptosis in vivo.

Laser microbeam-induced DNA damage inhibits cell division in fertilized eggs and early embryos

DNA double-strand breaks are caused by both intracellular physiological processes and environmental stress. In this study, we used laser microbeam cut (abbreviated microcut or cut), which allows specific DNA damage in the pronucleus of a fertilized egg and in individual blastomere(s) of an early embryo, to investigate the response of early embryos to DNA double-strand breaks. Line type γH2AX foci were detected in the cut region, while Chk2 phosphorylation staining was observed in the whole nuclear region of the cut pronuclei or blastomeres. Zygotes with cut male or female pronucleus showed poor developmental capability: the percentage of cleavage embryos was significantly decreased, and the embryos failed to complete further development to blastocysts. The cut blastomeres in 2-cell, 4-cell, and 8-cell embryos ceased cleavage, and they failed to incorporate into compacted morulae, but instead underwent apoptosis and cell death at the blastocyst stage; the uncut part of embryos could develop to blastocysts, with a reduced percentage or decreased cell number. When both blastomeres of the 2-cell embryos were cut by laser microbeam, cell death occurred 24 h earlier, suggesting important functions of the uncut blastomere in delaying cell death of the cut blastomere. Taken together, we conclude that microbeam-induced DNA damage in early embryos causes compromised development, and that embryos may have their own mechanisms to exclude DNA-damaged blastomeres from participating in further development.

Atomic mapping of Ruddlesden-Popper faults in transparent conducting BaSnO3-based thin films

Doped BaSnO3 has arisen many interests recently as one of the promising transparent conducting oxides for future applications. Understanding the microstructural characteristics are crucial for the exploration of relevant devices. In this paper, we investigated the microstructural features of 0.001% La doped BaSnO3 thin film using both conventional and aberration corrected transmission electron microscopes. Contrast analysis shows high densities of Ruddlesden-Popper faults in the film, which are on {100} planes with translational displacements of 1/2a < 111 > . Atomic EELS element mappings reveal that the Ruddlesden-Popper faults are Ba-O layer terminated, and two kinds of kink structures at the Ruddlesden-Popper faults with different element distributions are also demonstrated. Quantitative analysis on lattice distortions of the Ruddlesden-Popper faults illustrates that the local lattice spacing poses a huge increment of 36%, indicating that large strains exist around the Ruddlesden-Popper faults in the film.

LKB1 acts as a critical gatekeeper of ovarian primordial follicle pool.

Liver Kinase b1 (LKB1/STK11)is a tumor suppressor responsible for the Peutz-Jeghers syndrome, an autosomal-dominant, cancer-prone disorder in which patients develop neoplasms in several organs, including the oviduct, ovary, and cervix. Besides, the C allele of a SNP in the Lkb1 gene impedes the likelihood of ovulation in polycystic ovary syndrome (PCOS) in women treated with metformin, a known LKB1-AMPK activator. It is very likely that LKB1 plays roles in female fertility. To identify the physiological functions of LKB1 in the mouse ovary, we selectively disrupted LKB1 in oocytes by the Cre-LoxP conditional knockout system and found that Lkb1fl/fl; Gdf9-Cre mice were severely subfertile with significantly enlarged ovaries compared to Lkb1fl/fl mice. Interestingly, without Lkb1 expression in oocytes from the primordial follicle stage, the entire primordial follicle pool was activated but failed to mature and ovulate, subsequently causing premature ovarian failure (POF). Further investigation demonstrated that elevated mTOR signaling regulated by an AKT-independent LKB1-AMPK pathway was responsible for the excessive follicle activation and growth. Our findings reveal the role of LKB1 as an indispensable gatekeeper for the primordial follicle pool, offer new functional understanding for the tumor suppressor genes in reproductive organs, and might also provide valuable information for understanding POF and infertility.

SIRT1, 2, 3 protect mouse oocytes from postovulatory aging

The quality of metaphase II oocytes will undergo a time-dependent deterioration following ovulation as the result of the oocyte aging process. In this study, we determined that the expression of sirtuin family members (SIRT1, 2, 3) was dramatically reduced in mouse oocytes aged in vivo or in vitro. Increased intracellular ROS was observed when SIRT1, 2, 3 activity was inhibited. Increased frequency of spindle defects and disturbed distribution of mitochondria were also observed in MII oocytes aged in vitro after treatment with Nicotinamide (NAM), indicating that inhibition of SIRT1, 2, 3 may accelerate postovulatory oocyte aging. Interestingly, when MII oocytes were exposed to caffeine, the decline of SIRT1, 2, 3 mRNA levels was delayed and the aging-associated defective phenotypes could be improved. The results suggest that the SIRT1, 2, 3 pathway may play a potential protective role against postovulatory oocyte aging by controlling ROS generation.

Cyclin B3 controls anaphase onset independent of spindle assembly checkpoint in meiotic oocytes

Cyclin B3 is a relatively new member of the cyclin family whose functions are little known. We found that depletion of cyclin B3 inhibited metaphase-anaphase transition as indicated by a well-sustained MI spindle and cyclin B1 expression in meiotic oocytes after extended culture. This effect was independent of spindle assembly checkpoint activity, since both Bub3 and BubR1 signals were not observed at kinetochores in MI-arrested cells. The metaphase I arrest was not rescued by either Mad2 knockdown or cdc20 overexpression, but it was rescued by securin RNAi. We conclude that cyclin B3 controls the metaphase-anaphase transition by activating APC/Ccdc20 in meiotic oocytes, a process that does not rely on SAC activity.

The Dynamics and Regulatory Mechanism of Pronuclear H3k9me2 Asymmetry in Mouse Zygotes

H3K9 methylation is an important histone modification that is correlated with gene transcription repression. The asymmetric H3K9 dimethylation (H3K9me2) pattern between paternal and maternal genomes is generated soon after fertilization. In the present study, we carefully determined the dynamics of H3K9me2 changes in mouse zygotes, and investigated the regulatory mechanisms. The results indicated that histone methyltransferase G9a, but not GLP, was involved in the regulation of asymmetric H3K9me2, and G9a was the methyltransferase that induced the appearance of H3K9me2 in the male pronucleus of the zygote treated with cycloheximide. We found that there were two distinct mechanisms that regulate H3K9me2 in the male pronucleus. Before 8 h of in vitro fertilization (IVF), a mechanism exists that inhibits the association of G9a with the H3K9 sites. After 10 h of IVF the inhibition of G9a activity depends on yet unknown novel protein(s) synthesis. The two mechanisms of transfer take place between 8–10 h of IVF, and the novel protein failed to inhibit G9a activity in time, resulting in the appearance of a low level de novo H3K9me2 in the male pronucleus.

Geminin Deletion in Mouse Oocytes Results in Impaired Embryo Development and Reduced Fertility

Geminin is an important regulator of DNA replication and cell differentiation, but its role in female reproduction remains uncertain. Maternal geminin does not regulate oocyte meiotic maturation but does control accurate DNA replication. Geminin deletion in oocytes results in impaired embryo development and reduced fertility.

Cep55 regulates spindle organization and cell cycle progression in meiotic oocyte

Cep55 is a relatively novel member of the centrosomal protein family. Here, we show that Cep55 is expressed in mouse oocytes from the germinal vesicle (GV) to metaphase II (MII) stages. Immuostaining and confocal microscopy as well as time lapse live imaging after injection of mRNA encoding fusion protein of Cep55 and GFP identified that Cep55 was localized to the meiotic spindle, especially to the spindle poles at metaphase, while it was concentrated at the midbody in telophase in meiotic oocytes. Knockdown of Cep55 by specific siRNA injection caused the dissociation of γ-tubulin from the spindle poles, resulting in severely defective spindles and misaligned chromosomes, leading to metaphase I arrest and failure of first polar body (PB1) extrusion. Correspondingly, cyclin B accumulation and spindle assembly checkpoint (SAC) activation were observed in Cep55 knockdown oocytes. Our results suggest that Cep55 may act as an MTOC-associated protein regulating spindle organization, and thus cell cycle progression during mouse oocyte meiotic maturation.