Introns are essential for growth-regulated expression of the mouse thymidylate synthase gene.

The thymidylate synthase (TS) gene is expressed at much higher levels in proliferating cells than in quiescent cells. We have been studying the sequences that are important for regulating the mouse TS gene. We previously showed that DNA sequences upstream of the essential promoter elements as well as downstream of the ATG codon are both necessary (but neither is sufficient) for normal regulation in growth-stimulated cells. In the present study, we examined the possible roles of the coding region, polyadenylation signal, and introns as downstream regulatory elements. Minigenes consisting of 1 kb of the TS 5'-flanking region, the coding region (with or without various introns at their normal locations), and polyadenylation signals from the TS gene, the human beta-globin gene, and the bovine growth hormone gene were stably transfected into wild-type mouse 3T6 cells. Minigenes that contained introns 5 and 6, 1 and 2, or 1 alone were regulated regardless of which polyadenylation signal was included. A minigene that contained an internally deleted version of intron 1 was also regulated in response to growth stimulation. However, when all introns were omitted, there was little if any change in the level of minigene expression as cells progressed from G1 through S phase. These observations indicate that TS introns contain sequences that are necessary for normal growth-regulated expression of the mouse TS gene. These sequences appear to be associated with sequences that are important for splicing and to function in cooperation with upstream regulatory elements to bring about normal S-phase-specific expression.     

Stimulation of gene expression by introns: conversion of an inhibitory intron to a stimulatory intron by alteration of the splice donor sequence.

Efficient expression of many mammalian genes depends on the presence of at least one intron. We previously showed that addition of almost any of the introns from the mouse thymidylate synthase (TS) gene to an intronless TS minigene led to a large increase in expression. However, addition of intron 4 led to a reduction in minigene expression. The goal of the present study was to determine why TS intron 4 was unable to stimulate expression. Insertion of intron 4 into an intron-dependent derivative of the ribosomal protein L32 gene did not lead to a significant increase in expression, suggesting that its inability to stimulate expression was due to sequences within the intron. Deleting most of the interior of intron 4, improving the putative branch point, removing purines from the pyrimidine stretch at the 3' end of the intron, or removing possible alternative splice acceptor or donor sites within the intron each had little effect on the level of expression. However, when the splice donor sequence of intron 4 was modified so that it was perfectly complementary to U1 snRNA, the modified intron 4 stimulated expression approximately 6-fold. When the splice donor site of TS intron 1 (a stimulatory intron) was changed to that of TS intron 4, the modified intron 1 was spliced very inefficiently and lost the ability to stimulate mRNA production. Our observations support the idea that introns can stimulate gene expression by a process that depends directly on the splicing reaction.     

激光诱变小麦后代材料籽粒贮藏蛋白编码基因变异的研究

本研究采用SDS—PAGE分析了冀麦七号、冀麦18和冀早15三个品种激光后代籽粒贮藏蛋白的变异情况。结果表明,高分子量麦谷蛋白和醇溶蛋白亚基变异较小,其控制位点为Glu—1和Gl—3或Gli—1;而低分子量部分变异较大,其控制位点多在Glu—3、Gli—2和Gli—3。这些变异具有一定的随机性。有关激光诱变对小麦面粉品质的影响有待进一步研究。     

As above,not so below: Long-term dynamics of net primary production across a dryland transition zone

Drylands are key contributors to interannual variation in the terrestrial carbon sink, which has been attributed primarily to broad-scale climatic anomalies that disproportionately affect net primary production (NPP) in these ecosystems. Current knowledge around the patterns and controls of NPP is based largely on measurements of aboveground net primary production (ANPP), particularly in the context of altered precipitation regimes. Limited evidence suggests belowground net primary production (BNPP), a major input to the terrestrial carbon pool, may respond differently than ANPP to precipitation, as well as other drivers of environmental change, such as nitrogen deposition and fire. Yet long-term measurements of BNPP are rare, contributing to uncertainty in carbon cycle assessments. Here, we used 16 years of annual NPP measurements to investigate responses of ANPP and BNPP to several environmental change drivers across a grassland–shrubland transition zone in the northern Chihuahuan Desert. ANPP was positively correlated with annual precipitation across this landscape; however, this relationship was weaker within sites. BNPP, on the other hand, was weakly correlated with precipitation only in Chihuahuan Desert shrubland. Although NPP generally exhibited similar trends among sites, temporal correlations between ANPP and BNPP within sites were weak. We found chronic nitrogen enrichment stimulated ANPP, whereas a one-time prescribed burn reduced ANPP for nearly a decade. Surprisingly, BNPP was largely unaffected by these factors. Together, our results suggest that BNPP is driven by a different set of controls than ANPP. Furthermore, our findings imply belowground production cannot be inferred from aboveground measurements in dryland ecosystems. Improving understanding around the patterns and controls of dryland NPP at interannual to decadal scales is fundamentally important because of their measurable impact on the global carbon cycle. This study underscores the need for more long-term measurements of BNPP to improve assessments of the terrestrial carbon sink, particularly in the context of ongoing environmental change.     

Are brood sex ratios adaptive?—The effect of experimentally altered brood sex ratio on nestling growth,mortality and recruitment

Brood sex ratios (BSRs) have often been found to be nonrandom in respect of parental and environmental quality, and many hypotheses suggest that nonrandom sex ratios can be adaptive. To specifically test the adaptive value of biased BSRs, it is crucial to disentangle the consequences of BSR and maternal effects. In multiparous species, this requires cross-fostering experiments where foster parents rear offspring originating from multiple broods, and where the interactive effect of original and manipulated BSR on fitness components is tested. To our knowledge, our study on collared flycatchers (Ficedula albicollis) is the first that meets these requirements. In this species, where BSRs had previously been shown to be related to parental characteristics, we altered the original BSR of the parents shortly after hatching by cross-fostering nestlings among trios of broods and examined the effects on growth, mortality and recruitment of the nestlings. We found that original and experimental BSR, as well as the interaction of the two, were unrelated to the fitness components considered. Nestling growth was related only to background variables, namely brood size and hatching rank. Nestling mortality was related only to hatching asynchrony. Our results therefore do not support that the observed BSRs are adaptive in our study population. However, we cannot exclude the possibility of direct effects of experimentally altered BSRs on parental fitness, which should be evaluated in the future. In addition, studies similar to ours are required on various species to get a clearer picture of the adaptive value of nonrandom BSRs.     

反义寡聚硫代磷酸脱氧核苷抑制体外细胞培养系统中乙肝抗原表达的研究

设计合成了两个分别互补于乙肝病毒２．１ｋｂ ｍＲＮＡ起始区（片段Ａ）和增强子区（片段Ｂ）的硫代磷酸的ＤＮＡ片段,在经克隆ＨＢＶ ＤＮＡ转染ＨｅｐＧ２细胞建立的ＨＢＶ短暂表达系统及稳定产生ＨＢＶ的２２１５细胞中研究二者对ＨＢｓＡｇ及ＨＢｅＡｇ表达的抑制作用。结果表明反义寡聚物能不同程序抑制乙肝抗原表达,并与剂量呈一定正相关。在ＨｅｐＧ２细胞ＨＢＶ短暂表达系统中,６μｍｏｌ／Ｌ浓度时,片段Ａ、Ｂ对ＨＢ     

人工繁殖恒河猴的血清前清蛋白、转铁蛋白、苹果酸脱氢酶和醇脱氢酶遗传多态性研究

用薄层聚丙烯酰胺凝胶电泳方法分析118头人繁殖恒河猴血清四种蛋白质和同工酶遗传基因位点的多态性,结果表明,除醇脱氢酶(ADH)为单态外,其余三个基因位点均表现多态,前清蛋白(PA)可分为AA、AB、AC、AD和BB、CC,EE七种表型,各基因的频率为A 0.85,B 0.072,C 0.042,D 0.009,E 0.034转铁蛋白(Tf)可分为CC、DD、EE、FFGG、CD、CE、CG、CH、DE、DF、DG、DH、EF、EG、EH、FG十七种表型,墓因频率为C 0.064,D 0.380,E 0.188,F 0.111,G 0.244,H 0.014,苹果酸脱氢酶(MDH)可分MDH)1-1和MDH2-1两种表型,基因频率为MDH~10.958和MDH~20.042。     

Dieckol,a natural polyphenolic drug,inhibits the proliferation and migration of colon cancer cells by inhibiting PI3K,AKT, and mTOR phosphorylation

This study investigated that dieckol (DKL), a natural drug, inhibits colon cancer cell proliferation and migration by inhibiting phosphoinositide-3-kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) phosphorylation in HCT-116 cells. The cells were treated with DKL in various concentrations (32 and 50 μM) for 24 h and then analyzed for various experiments. MTT (tetrazolium bromide) and crystal violet assay investigated DKL-mediated cytotoxicity. Dichlorodihydrofluorescein diacetate staining was used to assess the reactive oxygen species (ROS) measurement, and apoptotic changes were studied by dual acridine orange and ethidium bromide staining. Protein expression of cell survival, cell cycle, proliferation, and apoptosis protein was evaluated by western blot analysis. Results indicated that DKL produces significant cytotoxicity in HCT-116, and the half-maximal inhibitory concentration was found to be 32 μM for 24-h incubation. Moreover, effective production of ROS and enhanced apoptotic signs were observed upon DKL treatment in HCT-116. DKL induces the expression of phosphorylated PI3K, AKT, and mToR-associated enhanced expression of cyclin-D1, proliferating cell nuclear antigen, cyclin-dependent kinase (CDK)-4, CDK-6, and Bcl-2 in HCT-116. In addition, proapoptotic proteins such as Bax, caspase-9, and caspase-3 were significantly enhanced by DKL treatment in HCT-116. Hence, DKL has been considered a chemotherapeutic drug by impeding the expression of PI3K-, AKT-, and mTOR-mediated inhibition of proliferation and cell cycle-regulating proteins.