New inducible genetic method reveals critical roles of GABA in the control of feeding and metabolism
New inducible genetic method reveals critical roles of GABA in the control of feeding and metabolism
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DOI：
10.1073/pnas.1602049113
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发表时间：
2016-03-29
影响因子：
11.1
通讯作者：
Wu, Qi
中科院分区：
文献类型：
--
作者：
Meng, Fantao;Han, Yong;Wu, Qi
目前可用的诱导型Cre/loxP系统，尽管它们在基因操作中具有相当大的实用性，但在某些情况下存在缺陷，例如不令人满意的重组率和对生理和行为的有害影响。为了克服这些局限性，我们设计了一种新的，诱导基因靶向系统，通过引入一个框内的无义突变的编码序列的Cre重组酶（nsCre）。从nsCre转基因转录的突变体mRNA可以在无义抑制子如氨基糖苷类的存在下有效地翻译成全长功能性Cre重组酶。在一个概念验证模型中，GABA信号从下丘脑神经元表达刺豚鼠相关肽（AgRP）的基因失活后4天内与合成氨基糖苷类药物治疗。年轻成年小鼠AgRP神经元中GABA合成的破坏导致体重急剧减轻，这是由于食物摄入减少和能量消耗增加;它们还表现出葡萄糖耐受不良。相反，年龄较大的小鼠与遗传失活的GABA信号的AgRP神经元只有短暂的减少喂养和体重，他们的能量消耗和葡萄糖耐量不受影响。这些结果表明，来自AgRP神经元的GABA能信号通过年龄依赖性机制在控制摄食和代谢中起关键作用。这种新的遗传技术将增强目前用于阐明许多生理和神经过程机制的工具。
Currently available inducible Cre/loxP systems, despite their considerable utility in gene manipulation, have pitfalls in certain scenarios, such as unsatisfactory recombination rates and deleterious effects on physiology and behavior. To overcome these limitations, we designed a new, inducible gene-targeting system by introducing an in-frame nonsense mutation into the coding sequence of Cre recombinase (nsCre). Mutant mRNAs transcribed from nsCre transgene can be efficiently translated into full-length, functional Cre recombinase in the presence of nonsense suppressors such as aminoglycosides. In a proof-of-concept model, GABA signaling from hypothalamic neurons expressing agouti-related peptide (AgRP) was genetically inactivated within 4 d after treatment with a synthetic aminoglycoside. Disruption of GABA synthesis in AgRP neurons in young adult mice led to a dramatic loss of body weight due to reduced food intake and elevated energy expenditure; they also manifested glucose intolerance. In contrast, older mice with genetic inactivation of GABA signaling by AgRP neurons had only transient reduction of feeding and body weight; their energy expenditure and glucose tolerance were unaffected. These results indicate that GABAergic signaling from AgRP neurons plays a key role in the control of feeding and metabolism through an age-dependent mechanism. This new genetic technique will augment current tools used to elucidate mechanisms underlying many physiological and neurological processes.