Reproductive isolation between sympatric sister species,Mussaenda kwangtungensis and M. pubescens var. alba

Reproductive isolation de fi nes the biological species concept and plays a key role in the formation and maintenance of species. The relative contributions of different isolating stages has been suggested to be closely associated with phylogenetic relatedness. Few studies have focused on the relative contributions of pre- versus postzygotic mechanisms, and even fewer have been conducted under strict phylogenetic frameworks. Pre- and post-zygotic reproductive isolation stages have been investigated in the sister species Mussaenda kwangtungensis and M. pubescens var. alba. The two species have partly overlapping distribution ranges and fl owering times, while the principal pollinators differed strikingly for them, demonstrating strong pre-zygotic isolations. Natural hybrids were detected by simple sequence repeat markers and their maternal parents were identi fi ed based on chloroplast gene sequences. Five out of 81 individuals were suggested to be hybrids that fall into the categories F2, BC1, and BC2 by theNew Hybrids analysis. Interspeci fi c crossings resulted in signi fi cantly reduced fruit set and seed germination rates.Phylogenetic analysis revealed short Kimura-2-parameter distance between M. kwangtungensis and M. pubescens var.alba. These fi ndings strongly supported the hypothesis that for species with a closer phylogenetic relationship, prezygotic isolation plays an important part in limiting gene exchange in sympatric areas.     

CRISPR/Cas9-mediated gene editing in human tripronuclear zygotes

Genome editing tools such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated system (Cas) have been widely used to modify genes in model systems including animal zygotes and human cells, and hold tremendous promise for both basic research and clinical applications. To date, a serious knowledge gap remains in our understanding of DNA repair mechanisms in human early embryos, and in the efficiency and potential off-target effects of using technologies such as CRISPR/Cas9 in human pre-implantation embryos. In this report, we used tripronuclear (3PN) zygotes to further investigate CRISPR/Cas9-mediated gene editing in human cells. We found that CRISPR/Cas9 could effectively cleave the endogenous β-globin gene (HBB). However, the efficiency of homologous recombination directed repair (HDR) of HBB was low and the edited embryos were mosaic. Off-target cleavage was also apparent in these 3PN zygotes as revealed by the T7E1 assay and whole-exome sequencing. Furthermore, the endogenous delta-globin gene (HBD), which is homologous to HBB, competed with exogenous donor oligos to act as the repair template, leading to untoward mutations. Our data also indicated that repair of the HBB locus in these embryos occurred preferentially through the non-crossover HDR pathway. Taken together, our work highlights the pressing need to further improve the fidelity and specificity of the CRISPR/Cas9 platform, a prerequisite for any clinical applications of CRSIPR/Cas9-mediated editing.     

Determination of baseline susceptibility to Cry1Ab protein for Asian corn borer (Lep., Crambidae)

Abstract: Although transgenic Bacillus thuringiensis (Bt) corn can provide a new tool for control of the Asian corn borer (ACB), Ostrinia furnacalis (Guenée), concern has been raised regarding the possibility of the target insect evolving resistance to the Bt protein under intensive selection pressure from Bt corn. Therefore, it is necessary to establish baseline data to enable detection of changes in susceptibility in field populations after prolonged exposure to Bt corn. Susceptibility to purified Cry1Ab protein from Bt was determined for 10 populations of ACB from the major corn‐growing regions of China, ranging geographically from Heilongjiang Province in the northeast to Shaanxi Province in the east‐central part. Neonate ACB were exposed to semi‐artificial diet incorporated with increasing Cry1Ab protein concentrations, and mortality and growth inhibition were evaluated after 7 days. The range of LC₅₀ (50% lethal concentration) among the populations was 0.10 to 0.81 μg/g (Cry1Ab protein/diet). Differences (P < 0.05) in susceptibility among the populations were significant. LC₅₀s generated from the Huanghuaihai Summer Corn Region were higher than those from the Spring Corn Regions. Bt was one of the significant natural biomortality factors of overwintering generation ACB. There was a significant correlation between percentage of the larvae infected with Bt and their LC₅₀ values to Cry1Ab protein in geographic distinct populations (r = 0.7350*, d.f. = 8, _r0.05 = 0.632). Based on the background of Bt formulations used for corn insect pests control in these areas, these differences were not caused by prior exposure to Bt insecticides. Instead, the small differences likely reflect natural Bt selection pressure. Because the variation in susceptibility to Cry1Ab was small (<10‐fold), the ACB apparently is susceptible to Cry1Ab across its range within China.     

GFP-mTn融合蛋白对蓝猪耳生活细胞微丝骨架的标记

用农杆菌介导法将嵌合基因GFP-mTn(mTn是微丝结合蛋白Talin的微丝结合域,可以显示活体细胞中微丝的结构)导入蓝猪耳。经激光共聚焦显微镜观察了转基因植株的各种不同组织中融合蛋白的表达和分布情况。在叶片的表皮细胞、保卫细胞、根部的皮层细胞中有融合蛋白的不同程度表达。但仅在保卫细胞中微丝标记状况良好,显示基因表达的组织特异性。经光诱导处于开放态的气孔的保卫细胞微丝呈网状结构,在细胞内无规则分布;经黑暗诱导处于关闭态的气孔保卫细胞中微丝束沿保卫细胞纵轴排列,呈卷曲状分布,并观察到螺旋和环状的微丝结构。在转基因植株的其他部位,例如茎表皮细胞、根毛细胞和花粉粒中,未检测到目的基因的表达。本研究获得的转基因植株为研究气孔运动过程中微丝动态变化提供了有用的材料。     

ADMA induces monocyte adhesion via activation of chemokine receptors in cultured THP-1 cells

Asymmetric dimethylarginine (ADMA), an endogenous NOS inhibitor, is also an important inflammatory factor contributing to the development of atherosclerosis (AS). The present study was to test the effect of ADMA on angiotensin (Ang) II-induced monocytic adhesion. Human monocytoid cells (THP-1) or isolated peripheral blood monocyte cells (PBMCs) were incubated with Ang II (10⁻⁶ M) or exogenous ADMA (30 μM) for 4 or 24 h in the absence or presence of losartan or antioxidant PDTC. In cultured THP-1 cells, Ang II (10⁻⁶ M) for 24 h elevated the level of ADMA in the medium, upregulated the protein expression of protein arginine methyltransferase (PRMT) and decreased the activity of dimethylarginine dimethylaminohydrolase (DDAH). Both of Ang II and ADMA increased monocytic adhesion to human umbilical vein endothelial cells (HUVECs), elevated the levels of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-8 and tumor necrosis factor (TNF)-α and upregulated CCR₂ and CXCR₂ mRNA expression, concomitantly with increase in reactive oxygen species (ROS) generation and activation of nuclear factor (NF)-κB. Pretreatment with losartan (10 μM) or PDTC (10 μM) abolished the effects mediated by Ang II or ADMA. In isolated PBMCs from healthy individuals, ADMA upregulated the expression of CXCR₂ mRNA, which was attenuated by losartan (10 μM), however, ADMA had no effect on surface protein expression of CCR₂. The present results suggest that ADMA may be involved in monocytic adhesion induced by Ang II via activation of chemokine receptors by ROS/NF-κB pathway.     

A major QTL controlling seed dormancy and pre-harvest sprouting resistance on chromosome 4A in a Chinese wheat landrace

Wheat pre-harvest sprouting (PHS) can cause significant reduction in yield and end-use quality of wheat grains in many wheat-growing areas worldwide. To identify a quantitative trait locus (QTL) for PHS resistance in wheat, seed dormancy and sprouting of matured spikes were investigated in a population of 162 recombinant inbred lines (RILs) derived from a cross between the white PHS-resistant Chinese landrace Totoumai A and the white PHS-susceptible cultivar Siyang 936. Following screening of 1,125 SSR primers, 236 were found to be polymorphic between parents, and were used to screen the mapping population. Both seed dormancy and PHS of matured spikes were evaluated by the percentage of germinated kernels under controlled moist conditions. Twelve SSR markers associated with both PHS and seed dormancy were located on the long arm of chromosome 4A. One QTL for both seed dormancy and PHS resistance was detected on chromosome 4AL. Two SSR markers, Xbarc 170 and Xgwm 397, are 9.14 cM apart, and flanked the QTL that explained 28.3% of the phenotypic variation for seed dormancy and 30.6% for PHS resistance. This QTL most likely contributed to both long seed dormancy period and enhanced PHS resistance. Therefore, this QTL is most likely responsible for both seed dormancy and PHS resistance. The SSR markers linked to the QTL can be used for marker-assisted selection of PHS-resistant white wheat cultivars. Shi-Bin Cai and Cui-Xia Chen contributed equally to this work.     

Formation of a new receptor-operated channel by heteromeric assembly of TRPP2 and TRPC1 subunits

Although several protein-protein interactions have been reported between transient receptor potential (TRP) channels, they are all known to occur exclusively between members of the same group. The only intergroup interaction described so far is that of TRPP2 and TRPC1; however, the significance of this interaction is unknown. Here, we show that TRPP2 and TRPC1 assemble to form a channel with a unique constellation of new and TRPP2/TRPC1-specific properties. TRPP2/TRPC1 is activated in response to G-protein-coupled receptor activation and shows a pattern of single-channel conductance, amiloride sensitivity and ion permeability distinct from that of TRPP2 or TRPC1 alone. Native TRPP2/TRPC1 activity is shown in kidney cells by complementary gain-of-function and loss-of-function experiments, and its existence under physiological conditions is supported by colocalization at the primary cilium and by co-immunoprecipitation from kidney membranes. Identification of the heteromultimeric TRPP2/TRPC1 channel has implications in mechanosensation and cilium-based Ca(2+) signalling.