三个苹果NBS基因的鉴定及其对外源生长物质的响应

为了探讨苹果NBS基因的生理作用,从苹果中鉴定了NBS家族的3个基因,分别命名为MdNBS、MdTIR-NBS1和MdTIR-NBS-LRR1。这3个基因编码的蛋白质均含有NB-ARC结构域,MdTIR-NBS1和MdTIR-NBS-LRR1蛋白N端还有TIR结构域,MdTIR-NBS-LRR1蛋白在C端含有LRR结构。荧光定量PCR分析表明,这3个不同类型的NBS基因的表达具有明显不同的组织特异性,此外,MdNBS、MdTIR-NBS1和MdTIR-NBS-LRR1在‘嘎啦’苹果幼苗叶片中均受SA和MeJA诱导。ACC处理不能提高MdNBS和MdTIR-NBS1基因的表达,但能诱导MdTIR-NBS-LRR1基因表达。结果表明,MdNBS、MdTIR-NBS1和MdTIR-NBS-LRR1基因可能参与了苹果抗逆或抗病防御反应。     

N-Methyl-d-aspartate Receptor- and Calpain-mediated Proteolytic Cleavage of K+-Cl? Cotransporter-2 Impairs Spinal Chloride Homeostasis in Neuropathic Pain

Loss of synaptic inhibition by γ-aminobutyric acid and glycine due to potassium chloride cotransporter-2 (KCC2) down-regulation in the spinal cord is a critical mechanism of synaptic plasticity in neuropathic pain. Here we present novel evidence that peripheral nerve injury diminishes glycine-mediated inhibition and induces a depolarizing shift in the reversal potential of glycine-mediated currents (E_glycine) in spinal dorsal horn neurons. Blocking glutamate N-methyl-d-aspartate (NMDA) receptors normalizes synaptic inhibition, E_glycine, and KCC2 by nerve injury. Strikingly, nerve injury increases calcium-dependent calpain activity in the spinal cord that in turn causes KCC2 cleavage at the C terminus. Inhibiting calpain blocks KCC2 cleavage induced by nerve injury and NMDA, thereby normalizing E_glycine. Furthermore, calpain inhibition or silencing of μ-calpain at the spinal level reduces neuropathic pain. Thus, nerve injury promotes proteolytic cleavage of KCC2 through NMDA receptor-calpain activation, resulting in disruption of chloride homeostasis and diminished synaptic inhibition in the spinal cord. Targeting calpain may represent a new strategy for restoring KCC2 levels and tonic synaptic inhibition and for treating chronic neuropathic pain.     

MiR-130a 在猪皮下脂肪细胞分化中的调节作用

为研究miR-130a对猪皮下脂肪细胞分化的影响及可能机制,本试验分离猪皮下脂肪前体细胞,诱导分化为成熟脂肪细胞,检测脂肪细胞分化过程中脂滴变化及miR-130a及其可能靶基因TNF α和PPARγ的表达模式.同时合成miR-130a mimics及inhibitor 对细胞进行转染,并以乱序序列作为阴性对照（NC）.细胞转染24 h后进行诱导分化,连续诱导8 d,检测各处理细胞的聚脂情况及甘油三酯含量变化,荧光定量PCR检测脂肪细胞分化相关基因的表达变化.结果显示,猪皮下脂肪前体细胞分化过程中脂滴逐渐变大增多,miR-130a、TNF α和PPARγ的表达模式具有一定的相似性.转染结果显示,相对于对照组,miR 130a mimics转染组细胞脂滴减少变小,甘油三酯含量降低（P<0.05）,脂肪细胞分化相关基因LPL、PPARγ、adiponectin、FASN和葡萄糖转运相关基因GLUT1,GLUT4以及JNK通路上的PDE3B的表达均比对照组显著下调（P<0.01）;而miR-130a inhibitor转染组细胞则脂滴增多,甘油三酯含量提高（P<0.05）,但大部分分化相关基因的表达与对照组无显著差异,提示miR-130a可能不只通过单一的靶基因影响脂肪细胞分化.其结果为后续深入研究miR-130a调节猪脂肪细胞分化的通路及机制奠定基础.     

Overexpression of Myostatin2 in zebrafish reduces the expression of dystrophin associated protein complex (DAPC) which leads to muscle dystrophy

Myostatin, a member of the TGF-β superfamily, is a potent negative regulator of skeletal muscle and growth. Previously, we reported Mstn1 from zebrafish and studied its influence on muscle development. In this study, we identified another form of Myostatin protein which is referred to as Mstn2. The size of Mstn2 cDNA is 1342 bp with 109 and 132 bp of 5′ and 3′-untranslated regions (UTRs), respectively. The coding region is 1101 bp encoding 367 amino acids. The identity between zebrafish Mstn1 and 2 is 66%. The phylogenetic tree revealed that the Mstn2 is an ancestral form of Mstn1. To study the functional aspects, we overexpressed mstn2 and noticed that embryos became less active and the juveniles with bent and curved phenotypes when compared to the control. The RT-PCR and in situ hybridization showed concurrent reduction of dystrophin associated protein complex (DAPC). In cryosection and in situ hybridization, we observed the disintegration of somites, lack of transverse myoseptum and loss of muscle integrity due to the failure of muscle attachment in mstn2 overexpressed embryos. Immunohistochemistry and western blot showed that there was a reduction of dystrophin, dystroglycan and sarcoglycan at translational level in overexpressed embryos. Taken together, these results indicate the suitability of zebrafish as an excellent animal model and our data provide the first in vivo evidence of muscle attachment failure by the overexpression of mstn2 and it leads to muscle loss which results in muscle dystrophy that may contribute to Duchenne syndrome and other muscle related diseases. A. Anusha Amali and Cliff Ji-Fan Lin contributed equally.     

Mutation Patterns at Dinucleotide Microsatellite Loci in Humans

Microsatellites are a major type of molecular markers in genetics studies. Their mutational dynamics are not clear. We investigated the patterns and characteristics of 97 mutation events unambiguously identified, from 53 multigenerational pedigrees with 630 subjects, at 362 autosomal dinucleotide microsatellite loci. A size-dependent mutation bias (in which long alleles are biased toward contraction, whereas short alleles are biased toward expansion) is observed. There is a statistically significant negative relationship between the magnitude (repeat numbers changed during mutation) and direction (contraction or expansion) of mutations and standardized allele size. Contrasting with earlier findings in humans, most mutation events (63%) in our study are multistep events that involve changes of more than one repeat unit. There was no correlation between mutation rate and recombination rate. Our data indicate that mutational dynamics at microsatellite loci are more complicated than the generalized stepwise mutation models.     

4种金龟甲的寄主偏好性及对不同植物挥发物的EAG反应

为了揭示华北大黑鳃金龟、暗黑鳃金龟、小黄鳃金龟和福婆鳃金龟对不同植物的寄主偏好性,本研究调查了四种金龟甲在6种寄主植物桃树、梨树、榆树、山楂、丁香和金银木上的发生情况。同时采用动态顶空吸附法收集7种植物挥发物,利用触角电位EAG技术,测定四种金龟甲雌、雄虫对不同寄主植物玉兰、柿树、金银木、榆树、芙蓉葵、丁香及非寄主植物银杏的挥发物的EAG反应。结果显示,金龟甲对不同寄主植物的偏好性有显著差异,华北大黑鳃金龟的偏好寄主为梨树,暗黑鳃金龟的偏好寄主为榆树、山楂和梨树,而小黄鳃金龟和福婆鳃金龟的偏好寄主均为丁香。四种金龟甲均对其共同寄主植物榆树挥发物的EAG反应最大,另外,小黄鳃金龟和福婆鳃金龟对丁香挥发物也有较大的EAG反应。对同一植物挥发物,华北大黑鳃金龟雌虫对部分植物挥发物的EAG反应比雄虫大;而其它3种鳃金龟雌、雄虫对所测试的植物挥发物的EAG反应均无显著性差异。上述结果说明金龟甲EAG反应的大小与其寄主偏好性一致。     

灵昆岛东滩潮间带大型底栖动物功能群及营养等级构成

2003年11月至2004年8月,在温州湾的灵昆岛东滩针对大型底栖动物群落进行了取样调查,利用调查数据进行功能群和营养等级构成研究。共得到大型底栖动物33种,隶属7门8纲21科,主要为软体动物、节肢动物甲壳类和环节动物多毛类。其中,高潮带25种,中潮带30种,低潮带14种。Margalef物种丰富度指数和Shannon—Weiner多样性指数均表现为：高潮带〉中潮带〉低潮带;Pielou均匀度指数和Simpson优势度指数表现为：高潮带〈中潮带〈低潮带。定性取样得到的浮游生物食者4种,植食者12种,肉食者8种,杂食者7种,碎屑食者2种,定量取样得到的分别为：3,8,5,3种和2种。定量取样获得数据中各个功能群种类数和密度季节间差异均不显著,不考虑季节变化因素肉食者和植食者密度在潮带间存在差异,杂食者和植食者种类数在潮带间存在差异,其他功能群密度和种类数数据在潮带间的差异不显著。研究显示了功能群的生境梯度变化,高潮带主导的功能群为肉食者,植食者和浮游生物食者;中潮带主导的功能群为肉食者和浮游生物食者;低潮带主导的功能群为肉食者,这些变化反映了不同潮带营养等级构成和食物来源的差异,由于生境中能提供的植物沿着高潮带,中潮带和低潮带方向递减,至低潮带未发现植食者;且由于中潮带的生境最为复杂,其功能群的构成也最为复杂;3个潮带的碎屑食者数量比较稳定。大型底栖动物群落与其食物来源一起共有4个营养等级,其中大型底栖动物占据了第2、3、4级,第1级主要碎屑、浮游生物和植物,第2级包括了碎屑食者、浮游生物食者和植食者,第3级主要是杂食者,第4级是肉食者,各个营养级、食物来源之间构成一张复杂的食物网。