Phosphatase type 2A-dependent and -independent pathways for ATR phosphorylation of Chk1

ATM and Rad3-related (ATR) is a regulatory kinase that, when activated by hydroxyurea, UV, or human immunodeficiency virus-1 Vpr, causes cell cycle arrest through Chk1-Ser(345) phosphorylation. We demonstrate here that of these three agents only Vpr requires protein phosphatase type 2A (PP2A) to activate ATR for Chk1-Ser(345) phosphorylation. A requirement for PP2A by Vpr was first shown with the PP2A-specific inhibitor okadaic acid, which reduced Vpr-induced G(2) arrest and Cdk1-Tyr(15) phosphorylation. Using small interference RNA to down-regulate specific subunits of PP2A indicated that the catalytic beta-isoform PP2A(Cbeta) and the A regulatory alpha-isoform PP2A(Aalpha) are involved in the G(2) induction, and these downregulations decreased the Vpr-induced, ATR-dependent phosphorylations of Cdk1-Tyr(15) and Chk1-Ser(345). In contrast, the same down-regulations had no effect on hydroxyurea- or UV-activated ATR-dependent Chk1-Ser(345) phosphorylation. Vpr and hydroxyurea/UV all induce ATR-mediated gammaH2AX-Ser(139) phosphorylation and foci formation, but down-regulation of PP2A(Aalpha) or PP2A(Cbeta) did not decrease gammaH2AX-Ser(139) phosphorylation by any of these agents or foci formation by Vpr. Conversely, H2AX down-regulation had little effect on PP2A(Aalpha/Cbeta)-mediated G(2) arrest and Chk1-Ser(345) phosphorylation by Vpr. The expression of vpr increases the amount and phosphorylation of Claspin, an activator of Chk1 phosphorylation. Down-regulation of either PP2A(Cbeta) or PP2A(Aalpha) had little effect on Claspin phosphorylation, but the amount of Claspin was reduced. Claspin may then be one of the phosphoproteins through which PP2A(Aalpha/Cbeta) affects Chk1 phosphorylation when ATR is activated by human immunodeficiency virus-1 Vpr.     

Into the night: camera traps reveal nocturnal activity in a presumptive diurnal primate, Rhinopithecus brelichi

Most living primates exhibit a daytime or nighttime activity pattern. Strict diurnality is thought to be the rule among anthropoids except for owl monkeys. Here we report the diel activity pattern of an Asian colobine, the Guizhou snub-nosed monkey Rhinopithecus brelichi, based on a methodology that relied on using 24-h continuously operating camera traps. We conducted the study in Fanjingshan National Nature Reserve in Guizhou, China from March 22 to May 19 and from June 17 to October 14, 2011. After standardizing all time elements to a meridian-based time according to the geographic coordinates of the study site, we showed unequivocally that the monkeys, though predominantly diurnal, exhibited activity beyond daylight hours throughout the study. Specifically, their activity at night and during twilight periods suggests a complex interplay of behavioral adaptations, among others, to living in a temperate environment where day length and food resources fluctuate substantially across seasons. We contend that, under prevailing ecological conditions, so-called strictly diurnal primates may adjust their activity schedule opportunistically in order to increase energy intake. We also discuss the advantages of using camera traps in primate studies, and how the standardized use of meridian-based time by researchers would benefit comparisons of diel activity patterns among primates.     

Addition of Phenylboronic Acid to Malus domestica Pollen Tubes Alters Calcium Dynamics,Disrupts Actin Filaments and Affects Cell Wall Architecture

A key role of boron in plants is to cross-link the cell wall pectic polysaccharide rhamnogalacturonan-II (RG-II) through borate diester linkages. Phenylboronic acid (PBA) can form the same reversible ester bonds but cannot cross-link two molecules, so can be used as an antagonist to study the function of boron. This study aimed to evaluate the effect of PBA on apple (Malus domestica) pollen tube growth and the underlying regulatory mechanism. We observed that PBA caused an inhibition of pollen germination, tube growth and led to pollen tube morphological abnormalities. Fluorescent labeling, coupled with a scanning ion-selective electrode technique, revealed that PBA induced an increase in extracellular Ca²⁺ influx, thereby elevating the cytosolic Ca²⁺ concentration [Ca²⁺]c and disrupting the [Ca²⁺]c gradient, which is critical for pollen tube growth. Moreover the organization of actin filaments was severely perturbed by the PBA treatment. Immunolocalization studies and fluorescent labeling, together with Fourier-transform infrared analysis (FTIR) suggested that PBA caused an increase in the abundance of callose, de-esterified pectins and arabinogalactan proteins (AGPs) at the tip. However, it had no effect on the deposition of the wall polymers cellulose. These effects are similar to those of boron deficiency in roots and other organs, indicating that PBA can induce boron deficiency symptoms. The results provide new insights into the roles of boron in pollen tube development, which likely include regulating [Ca²⁺]c and the formation of the actin cytoskeleton, in addition to the synthesis and assembly of cell wall components.     

Protein interaction analysis of ST14 domains and their point and deletion mutants

ST14 (suppression of tumorigenicity 14) is a transmembrane serine protease that contains a serine protease catalytic (SP) domain, an SEA domain, two complement subcomponent C1r/s (CUB) domains, and four low density lipoprotein receptor class A domains. Glutathione S-transferase fusion proteins with SP, CUB, and low density lipoprotein receptor domains and their corresponding mutants were generated to analyze protein interactions with these domains. Modified glutathione S-transferase pull-down assays demonstrated the interaction between the SP domain and hepatocyte growth factor activator inhibitor-1. With the same method, a CUB domain-interacting protein was isolated and turned out to be the transmembrane protein with epidermal growth factor-like and two follistatin-like domains 1 (TMEFF1). Quantitative real time PCR revealed that the expression of the TMEFF1 gene was dependent on the transfection of the ST14 gene in the RKO cell line. Our results also suggested that ST14 and TMEFF1 were co-expressed in the human breast cancer cell line MCF7, human placenta, kidney, and liver tissues. Interestingly, these two genes were co-up-regulated in kidney tumors versus normal tissues, consistent with our results that showed the dependence of TMEFF1 expression on ST14 in RKO cells. Finally, homology modeling studies suggested that TMEFF1 might form a complex with ST14 by an interaction between epidermal growth factor and CUB domains.     

Overexpression of TRIM26 suppresses the proliferation,metastasis, and glycolysis in papillary thyroid carcinoma cells

Papillary thyroid carcinoma (PTC) is the common subtype of thyroid cancer, which is a common endocrine malignancy. Tripartite motif 26 (TRIM26) has been found to act as a tumor suppressor in several cancers. However, the functional roles of TRIM26 in PTC remain unknown. In this study, we examined the TRIM26 expression in PTC and evaluated the effects of TRIM26 on proliferation, metastasis, and glycolysis in PTC cells. The results proved that TRIM26 was significantly downregulated in PTC tissues and cell lines. TRIM26 overexpression inhibited cell proliferation, migration, and invasion in PTC cells. TRIM26 overexpression also suppressed the epithelial-to-mesenchymal transition process. Besides, overexpression of TRIM26 caused significant decrease in glucose uptake and lactate production in PTC cells. Further investigations revealed that TRIM26 overexpression inhibited the activation of PI3K/Akt pathway. Treatment with an activator (740Y-P) of the PI3K/AKT pathway reversed the antitumor effects of TRIM26 on PTC cells. These findings provided evidence that TRIM26 acted as a tumor suppressor in PTC.     

Influence of bacterial leaf blight on the photosynthetic characteristics of resistant and susceptible rice

The effect of crop disease on photosynthetic characteristics is important for disease control. Two varieties, Shenzhou 98 and Neiwuyou 8015 with resistance and susceptibility to bacterial leaf blight (BLB), respectively, were selected, and the responses of the net photosynthetic rate (P_N) to active photon flux density (PPFD) and intercellular carbon dioxide concentration (C_i), as well as chlorophyll fluorescence, pigments and stomatal resistance (SR), were measured. The results showed that BLB infection greatly decreased the maximum photosynthetic rate (P_max), light saturation point (LSP), carboxylation efficiency (CE), maximal fluorescence (F_m) and actual photochemical efficiency of PSII ( Φ _PSII) but increased the light compensation point (LCP) and dark respiratory rate (R_D), which suggested that the performance of rice photosynthesis was decreased by BLB infection. The BLB infection had a lower effect on resistant rice Shenzhou 98 than on susceptible rice Neiwuyou 8015. The reduction of pigment and increased SR caused by BLB infection may have resulted in the decline in the photosynthetic rate. Significant effects of the BLB infection were observed on chlorophyll fluorescence F_m and Φ_PSII in resistant and susceptible rice. These parameters may be useful for noninvasive monitoring of plant disease considering the negative effect caused by other stresses.     

Simultaneous Increases in Proliferation and Apoptosis of Vascular Smooth Muscle Cells Accelerate Diabetic Mouse Venous Atherosclerosis

Aims

This study was designed to demonstrate simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) leading to accelerated vein graft remodeling and to explore the underlying mechanisms.

Methods

Vein grafts were performed in non-diabetic and diabetic mice. The cultured quiescent VSMCs were subjected to mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs). Harvested vein grafts and treated VSMCs were used to detect cell proliferation, apoptosis, mitogen-activated protein kinases (MAPKs) activation and SM-α-actin expression.

Results

Significantly thicker vessel walls and greater increases in proliferation and apoptosis were observed in diabetic vein grafts than those in non-diabetic. Both SS and AGEs were found to induce different activation of three members of MAPKs and simultaneous increases in proliferation and apoptosis of VSMCs, and combined treatment with both had a synergistic effect. VSMCs with strong SM-α-actin expression represented more activated JNKs or p38MAPK, and cell apoptosis, while the cells with weak SM-α-actin expression demonstrated preferential activation of ERKs and cell proliferation. In contrast, inhibition of MAPKs signals triggered significant decreases in VSMC proliferation, and apoptosis. Treatment of the cells with RNA interference of receptor of AGEs (RAGE) also resulted in significant decreases in both proliferation and apoptosis.

Conclusions

Increased pressure-induced SS triggers simultaneous increases in proliferation and apoptosis of VSMCs in the vein grafts leading to vein arterializations, which can be synergistically accelerated by high glucose-induced AGEs resulting in vein graft atherosclerosis. Either SS or AGEs and their combination induce simultaneous increases in proliferation and apoptosis of VSMCs via different activation of three members of MAPKs resulting from different VSMC subtypes classified by SM-α-actin expression levels.     

栽培番茄的塿土和黄绵土水分和温度变化规律

用土柱试验,研究了栽培樱桃番茄(Lycopersicon esculentum var.cerasiforme Alef.)的塿土和黄绵土水分运移和温度变化规律,水分运移模型选用土壤中水分分布的动力学模型,土壤温度、空气温湿度变化选用正弦曲线模型。结果表明:塿土在各个不同深度的平均含水量均高于黄绵土,塿土的入渗速率高于黄绵土,同一深度塿土温度高于黄绵土,土壤温度随着深度的增加具有明显的滞后性;黄绵土中樱桃番茄的水分利用效率大于塿土,空气温湿度、土壤温度和土壤含水量相互影响。水分运移模型在土壤浅层处可以得到很好的拟合效果,在拟合方程的变量范围内,根据时间可以较准确的确定樱桃番茄盛果期土壤浅层含水量,对于进一步提高农业干旱防御能力、有效制定节水灌溉计划、提高水分利用效率提供了理论依据。