Activated Ras requires autophagy to maintain oxidative metabolism and tumorigenesis

Autophagy is a catabolic pathway used by cells to support metabolism in response to starvation and to clear damaged proteins and organelles in response to stress. We report here that expression of a H-ras(V12) or K-ras(V12) oncogene up-regulates basal autophagy, which is required for tumor cell survival in starvation and in tumorigenesis. In Ras-expressing cells, defective autophagosome formation or cargo delivery causes accumulation of abnormal mitochondria and reduced oxygen consumption. Autophagy defects also lead to tricarboxylic acid (TCA) cycle metabolite and energy depletion in starvation. As mitochondria sustain viability of Ras-expressing cells in starvation, autophagy is required to maintain the pool of functional mitochondria necessary to support growth of Ras-driven tumors. Human cancer cell lines bearing activating mutations in Ras commonly have high levels of basal autophagy, and, in a subset of these, down-regulating the expression of essential autophagy proteins impaired cell growth. As cancers with Ras mutations have a poor prognosis, this "autophagy addiction" suggests that targeting autophagy and mitochondrial metabolism are valuable new approaches to treat these aggressive cancers.     

Forebrain-specific knockout of B-raf kinase leads to deficits in hippocampal long-term potentiation, learning, and memory

Raf kinases are downstream effectors of Ras and upstream activators of the MEK-ERK cascade. Ras and MEK-ERK signaling play roles in learning and memory (L&M) and neural plasticity, but the roles of Raf kinases in L&M and plasticity are unclear. Among Raf isoforms, B-raf is preferentially expressed in the brain. To determine whether B-raf has a role in synaptic plasticity and L&M, we used the Cre-LoxP gene targeting system to derive forebrain excitatory neuron B-raf knockout mice. This conditional knockout resulted in deficits in ERK activation and hippocampal long-term potentiation (LTP) and impairments in hippocampus-dependent L&M, including spatial learning and contextual discrimination. Despite the widespread expression of B-raf, this mutation did not disrupt other forms of L&M, such as cued fear conditioning and conditioned taste aversion. Our findings demonstrate that B-raf plays a role in hippocampal ERK activation, synaptic plasticity, and L&M.     

P2Y1受体对缺血状态下星形胶质细胞产生胶质原纤维酸性蛋白与胶质细胞源性神经营养因子的影响及相关信号通路

目的研究P2Y1受体对缺血时星形胶质细胞产生胶质原纤维酸性蛋白（glial fibrillary acidic protein,GFAP）及胶质细胞源性神经营养因子（glial cell line—derived neurotrophic factor,GDNF）的影响及其相关信号通路。方法分别利用右侧大腑中动脉线拴阻塞及培养细胞缺氧无营养后恢复正常培养,造成体内、外缺血再灌注模型。用免疫荧光标记、实时定量RT—PCR、Western blotting、酶联免疫吸附试验观察P2Y1受体、GDNF定位,检测GFAP、GDNF及信号分子的表达变化。结果与单纯性缺血组比较,用选择性拈抗剂MRS2179阻断P2Y1受体后,可使体内、外星形胶质细胞产生的GFAP减少,同时使其产生GDNF增加。体外缺氧无营养并阻断P2Y1受体后：可使磷酸化蛋白激酶B（Akt）及cAMP反应元件结合蛋（cAMP response element binding protein,CREB）升高,而使磷酸化JAK2及STAT3（Ser727）降低;JAK2的抑制剂AG490在降低磷酸化STAT3（Ser727）的同时也降低GFAP表达水平;PI3-K的抑制剂LY294002可降低磷酸化的Akt及CREB;MEK1／2抑制剂U0126可同时降低磷酸化的JAK2、STAT3 （Ser727）、Akt及CREB。结论P2Y1受体参与短时性缺血时星形胶质细胞GFAP及GDNF的产生过程,相关信号途径分别为JAK2／STAT3和P13-K／AKT／CREB,并且两条途径存在串话。     

RASSF1A和APC基因启动子区甲基化与前列腺癌的关系

目的 研究Ras相关区域家族1A (Ras association domain family l isoform A,RASSF1A)及结肠腺瘤性息肉病(adenomatosis polyposis coli,APC)基因启动子区CpG甲基化及与前列腺癌(PCa)之间的关系,探索PCa早期诊断的检测方法.方法 收集60例PCa和40例前列腺增生(BPH)病例的组织标本及其相关的临床指标.应用亚硫酸氢盐修饰后测序法检测PCa组织及BPH组织中RASSF1A、APC基因启动子区CpG甲基化情况.结果 PCa组RASSF1A、APC基因CG位点甲基化率高于BPH组(60.8％ vs 14％,48.84％ vs 1.19％,P＜0.05);基因甲基化率与PSA、Gleason评分、病理分期和PCa危险分期关系密切(P＜0.01); RASSF1A及APC基因甲基化联合检测用于判断PCa及BPH的敏感度和特异度是95.74％和82.9％.结论 RASSFIA、APC基因启动子区甲基化与PCa发生及发展有关,其甲基化率的变化与PCa的危险分期关系密切.检测前列腺组织中相关基因甲基化状态,有望成为诊断早期PCa的一种方法.     

Fear learning and extinction are linked to neuronal plasticity through Rin1 signaling

The amygdala is known to have a crucial role in both the acquisition and extinction of conditioned fear, but the physiological changes and biochemical mechanisms underlying these forms of learning are only partly understood. The Ras effector Rin1 activates Abl tyrosine kinases and Rab5 GTPases and is highly expressed in mature neurons of the telencephalon including the amygdala, where it inhibits the acquisition of fear memories (Rin1^?/? mice show enhanced learning of conditioned fear). Here we report that Rin1^?/? mice exhibit profound deficits in both latent inhibition and fear extinction, suggesting a critical role for Rin1 in gating the acquisition and persistence of cue‐dependent fear conditioning. Surprisingly, we also find that depotentiation, a proposed cellular mechanism of extinction, is enhanced at lateral‐basolateral (LA‐BLA) amygdaloid synapses in Rin1^?/? mice. Inhibition of a single Rin1 downstream effector pathway, the Abl tyrosine kinases, led to reduced amygdaloid depotentiation, arguing that proper coordination of Abl and Rab5 pathways is critical for Rin1‐mediated effects on plasticity. While demonstrating a correlation between amygdala plasticity and fear learning, our findings argue against models proposing a direct causative relationship between amygdala depotentiation and fear extinction. Taken together, the behavior and physiology of Rin1^?/? mice provide new insights into the regulation of memory acquisition and maintenance. In addition, Rin1^?/? mice should prove useful as a model for pathologies marked by enhanced fear acquisition and retention, such as posttraumatic stress disorder. © 2009 Wiley‐Liss, Inc.