温度敏感TRP离子通道的异源组装和功能研究
发布时间：2018-12-13 19:35
【摘要】：温度感觉作为人体的一种重要感觉系统,对于感受周围环境变化,躲避危险性伤害以及维持机体内环境的稳定都具有非常重要的意义。长期以来对温度感觉机理的研究一直停留于直观描述阶段。造成温度感觉研究长期落后的主要原因在于温度感受器分子一直没有被确定。温度敏感TRP(Transient Receptor Potential)离子通道被分子克隆并证实为主要的细胞温度感受器,为在分子水平上研究温度感觉的分子机制提供了实验基础。 作者前期的研究工作表明温度敏感TRPV通道亚基间可以交互杂合组装为新的通道,杂合通道的类型和其不同的亚基配比有关,分子组装过程中亚基配比为随机的,杂合通道的电导和门控动力学特性介于纯合母体通道之间。通道亚基间的异源组装有助于离子通道在不同组织,不同细胞类型以及不同生理状态下功能多样性的广泛扩展。近年来对于温度敏感TRP通道的异源组装研究也有大量报道,但杂合通道的功能方面我们却知之甚少。 在本课题主要研究了杂合TRPV1/TRPV3通道在热、化合物及电位激活过程中的通道功能特性。同时作者还将通道异源组装的研究扩展到其它温度敏感TRP通道蛋白亚基。结果表明杂合TRPV1/TRPV3通道对TRPV1的激活剂和拮抗剂也都非常敏感;杂合TRPV1/TRPV3通道具有特定的温度敏感性、激活温度阈值和由热引发的通道敏化现象。杂合通道门控特性的改变显然是由TRPV1和TRPV3通道亚基间的相互作用所造成的。温度敏感TRP通道异源组装的实验研究中,作者证明了在细胞中表达TRPV1-TRPV3的串联体能够形成单一的杂合TRPV1/TRPV3通道,通过内面向外(inside-out)的膜片钳记录模式可以从表达串联体的细胞中记录到单通道电流。甚至单通道电导也很一致地处于纯合TRPV1和TRPV3通道的电导之间。这个结果和作者前期发表的有关TRPV1和TRPV3共同表达于细胞中可以组成杂合通道的结果一致。而对其它温度敏感TRP离子通道的异源组装研究显示其蛋白亚基间不能形成杂合通道。 综上所述,本研究证明了杂合TRPV1/TRPV3通道在细胞中的异源组装以及杂合通道在热激活及化学激活过程中功能的扩展,杂合通道的形成和功能特性的扩展也许有助于机体更加精确地对各种感觉和痛觉的敏感性进行细微的调节,从而更好地保护机体躲避有害刺激。
[Abstract]:As an important sensory system of human body, temperature sense is of great significance to feel the changes of the surrounding environment, avoid dangerous damage and maintain the stability of the body's internal environment. For a long time, the research on temperature sensing mechanism has been in the stage of visual description. The main reason for the long lag in the study of temperature sensing is that the thermoreceptor molecules have not been identified. The thermo-sensitive TRP (Transient Receptor Potential) ion channel was cloned and confirmed as the main cellular thermoreceptor, which provides the experimental basis for studying the molecular mechanism of temperature sensing at the molecular level. The previous work of the authors showed that the cross-heterozygous subunits of temperature-sensitive TRPV channels could be assembled into new channels, and the types of heterozygotes were related to their different subunit ratios, and the ratio of subunits in the molecular assembly process was random. The conductivity and gated dynamics of hybrid channels are between homozygous parent channels. The heterologous assembly of channel subunits contributes to the extensive expansion of ion channels in different tissues, cell types and physiological states. In recent years, there have been a lot of reports on the heterologous assembly of temperature-sensitive TRP channels, but little is known about the function of heterozygotes. In this paper, the functional characteristics of heterozygous TRPV1/TRPV3 channels during heat, compound and potential activation were studied. At the same time, the study of channel heterologous assembly was extended to other temperature sensitive TRP channel protein subunits. The results showed that the heterozygous TRPV1/TRPV3 channels were also sensitive to the activators and antagonists of TRPV1, and the heterozygous TRPV1/TRPV3 channels had specific temperature sensitivity, activation threshold and heat-induced channel sensitization. The change of heterozygous channel gating is obviously caused by the interaction between TRPV1 and TRPV3 channel subunits. In an experimental study of the heterologous assembly of thermo-sensitive TRP channels, the authors have demonstrated that a single heterozygous TRPV1/TRPV3 channel can be formed by the cascade expressing TRPV1-TRPV3 in cells. A single channel current can be recorded from the cells expressing the tandem through the patch-clamp recording mode of the inner outward (inside-out). Even the single channel conductance is very consistent between the homozygous TRPV1 and the TRPV3 channel conductance. This result is consistent with the results previously published by the authors that TRPV1 and TRPV3 co-express in cells to form heterozygous channels. Heterologous assembly of other temperature-sensitive TRP ion channels showed that heterozygous channels could not be formed between their protein subunits. In conclusion, this study demonstrated the heterogenous assembly of heterozygous TRPV1/TRPV3 channels in cells and the expansion of heterozygous channels in the processes of thermal and chemical activation. The formation of heterozygous channels and the expansion of functional characteristics may help the body to regulate the sensitivity of various senses and pain sensations more accurately and thus better protect the body from harmful stimuli.
【学位授予单位】：大连理工大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R339.1;Q43
本文编号：2377095
[Abstract]:As an important sensory system of human body, temperature sense is of great significance to feel the changes of the surrounding environment, avoid dangerous damage and maintain the stability of the body's internal environment. For a long time, the research on temperature sensing mechanism has been in the stage of visual description. The main reason for the long lag in the study of temperature sensing is that the thermoreceptor molecules have not been identified. The thermo-sensitive TRP (Transient Receptor Potential) ion channel was cloned and confirmed as the main cellular thermoreceptor, which provides the experimental basis for studying the molecular mechanism of temperature sensing at the molecular level. The previous work of the authors showed that the cross-heterozygous subunits of temperature-sensitive TRPV channels could be assembled into new channels, and the types of heterozygotes were related to their different subunit ratios, and the ratio of subunits in the molecular assembly process was random. The conductivity and gated dynamics of hybrid channels are between homozygous parent channels. The heterologous assembly of channel subunits contributes to the extensive expansion of ion channels in different tissues, cell types and physiological states. In recent years, there have been a lot of reports on the heterologous assembly of temperature-sensitive TRP channels, but little is known about the function of heterozygotes. In this paper, the functional characteristics of heterozygous TRPV1/TRPV3 channels during heat, compound and potential activation were studied. At the same time, the study of channel heterologous assembly was extended to other temperature sensitive TRP channel protein subunits. The results showed that the heterozygous TRPV1/TRPV3 channels were also sensitive to the activators and antagonists of TRPV1, and the heterozygous TRPV1/TRPV3 channels had specific temperature sensitivity, activation threshold and heat-induced channel sensitization. The change of heterozygous channel gating is obviously caused by the interaction between TRPV1 and TRPV3 channel subunits. In an experimental study of the heterologous assembly of thermo-sensitive TRP channels, the authors have demonstrated that a single heterozygous TRPV1/TRPV3 channel can be formed by the cascade expressing TRPV1-TRPV3 in cells. A single channel current can be recorded from the cells expressing the tandem through the patch-clamp recording mode of the inner outward (inside-out). Even the single channel conductance is very consistent between the homozygous TRPV1 and the TRPV3 channel conductance. This result is consistent with the results previously published by the authors that TRPV1 and TRPV3 co-express in cells to form heterozygous channels. Heterologous assembly of other temperature-sensitive TRP ion channels showed that heterozygous channels could not be formed between their protein subunits. In conclusion, this study demonstrated the heterogenous assembly of heterozygous TRPV1/TRPV3 channels in cells and the expansion of heterozygous channels in the processes of thermal and chemical activation. The formation of heterozygous channels and the expansion of functional characteristics may help the body to regulate the sensitivity of various senses and pain sensations more accurately and thus better protect the body from harmful stimuli.
【学位授予单位】：大连理工大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R339.1;Q43
【引证文献】
相关期刊论文 前1条
1 陈晓博;曹鹏;于锋;王大为;;温度敏感瞬时受体电位通道参与化疗引起的外周神经病变的研究进展[J];药学与临床研究;2014年02期
，本文编号：2377095
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