Connexin45 expression is preferentially associated with the ventricular conduction system in mouse and rat heart.
Connexin45 expression is preferentially associated with the ventricular conduction system in mouse and rat heart.
复制标题
Connexin45 表达优先与小鼠和大鼠心脏的心室传导系统相关。
作者：
S. Coppen;E. Dupont;S. Rothery;N. Severs
心肌细胞通过缝隙连接电连接，缝隙连接是由连接蛋白组成的低电阻细胞间通道簇。不同连接蛋白类型的数量和空间分布的变化与心脏电生理特性的区域分化有关。尽管独立研究已经证明，连接蛋白43在工作心室心肌中含量丰富，并且连接蛋白40优先在许多物种的房室传导系统中表达，但有关连接蛋白45在心脏中的空间分布的信息仅限于使用获得的数据针对单一肽序列产生的抗体。在本研究中，我们报告了一种新的抗连接蛋白45抗体的生产和表征及其在小鼠和大鼠心肌中连接蛋白45表达的研究中的应用。亲和纯化的抗血清，在豚鼠中提出的残基354至367的人连接蛋白45，识别一个单一的45-kD带的HeLa细胞转染表达连接蛋白45的蛋白质印迹，并给出点状免疫标记的细胞边界，冷冻断裂细胞化学证明代表间隙连接。在小鼠和大鼠心脏组织的北方印迹上仅检测到低水平的连接蛋白45 mRNA，并且连接蛋白45蛋白水平低于Western印迹的检测限。免疫标记的心室组织的共聚焦显微镜显示，工作心肌的大部分是免疫阴性的连接蛋白45。一个明确定义的区域包含连接蛋白45表达的细胞，然而，定位于内皮细胞表面，与连接蛋白40表达的传导系统的肌细胞重叠。由于这些结果与连接蛋白45广泛分布于工作心室肌细胞的流行观点形成对比，我们比较了与商业供应的抗连接蛋白45抗血清针对先前研究中使用的相同肽产生的免疫标记模式。商业连接蛋白45抗血清在整个心室肌中广泛标记，但这种标记被连接蛋白43序列的6个氨基酸肽匹配部分抑制，表明商业连接蛋白45抗血清与组织中的连接蛋白43发生交叉反应。这种交叉反应性的进一步证据来自对连接蛋白43转染细胞的观察，其用商业抗连接蛋白45抗血清给出阳性免疫标记。我们的一个特定的协会connexin 45与connexin 40表达的心肌细胞在大鼠和小鼠心室的演示提出了可能性，connexin 45有助于调制的电生理特性在心室传导系统，并强调需要重新评估connexin 45在其他物种的分布和作用。
Cardiac myocytes are electrically coupled by gap junctions, clusters of low-resistance intercellular channels composed of connexins. Variations in the quantities and spatial distribution of different connexin types have been implicated in regional differentiation of electrophysiological properties in the heart. Although independent studies have demonstrated that connexin43 is abundant in working ventricular myocardium and that connexin40 is preferentially expressed in the atrioventricular conduction system of a number of species, information on the spatial distribution of connexin45 in the heart is limited to data obtained using an antibody raised to a single peptide sequence. In the present study, we report on the production and characterization of a new anti-connexin45 antibody and its application to the investigation of connexin45 expression in mouse and rat myocardium. The affinity-purified antiserum, raised in guinea pig to residues 354 to 367 of human connexin45, recognized a single 45-kD band on Western blots of HeLa cells transfected to express connexin45 and gave punctate immunolabeling at the cell borders, demonstrated by freeze-fracture cytochemistry to represent gap junctions. Only low levels of connexin45 mRNA were detected on Northern blots of mouse and rat cardiac tissues, and connexin45 protein levels were below the limit of detection on Western blots. Confocal microscopy of immunolabeled ventricular tissue revealed that the major part of the working myocardium was immunonegative for connexin45. A clearly defined zone containing connexin45-expressing cells was, however, localized to the endocardial surface, overlapping with connexin40-expressing myocytes of the conduction system. As these results contrast with the prevailing view that connexin45 is widely distributed in working ventricular myocytes, we compared the immunolabeling pattern obtained with a commercially supplied anti-connexin45 antiserum raised against the same peptide that was used in previous studies. The commercial connexin45 antiserum gave widespread labeling throughout the ventricular myocardium, but this labeling was inhibited by a six-amino acid peptide matching part of the connexin43 sequence, indicating cross-reaction of the commercial connexin45 antiserum with connexin43 in the tissue. Further evidence for such cross-reactivity came from observations on connexin43-transfected cells, which gave positive immunolabeling with the commercial anti-connexin45 antiserum. Our demonstration of a specific association of connexin45 with connexin40-expressing myocytes in rat and mouse ventricle raises the possibility that connexin45 contributes to the modulation of electrophysiological properties in the ventricular conduction system and highlights the need for reappraisal of the distribution and role of connexin45 in other species.
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影响因子：
3.3
作者：
BRUZZONE, R;HAEFLIGER, JA;PAUL, DL
通讯作者：
PAUL, DL
间隙连接蛋白 connexin45 的表征。
DOI：
10.1007/bf00232672
发表时间：
1994
期刊：
The Journal of membrane biology
影响因子：
--
作者：
Laing,JG;Westphale,EM;Engelmann,GL;Beyer,EC
通讯作者：
Beyer,EC
影响因子：
20.1
作者：
DARROW, BJ;LAING, JG;BEYER, EC
通讯作者：
BEYER, EC
影响因子：
20.1
作者：
KANTER, HL;LAING, JG;SAFFITZ, JE
通讯作者：
SAFFITZ, JE
DOI：
10.1016/0735-1097(94)90879-6
发表时间：
1994-10-01
影响因子：
24
作者：
DAVIS, LM;KANTER, HL;SAFFITZ, JE
通讯作者：
SAFFITZ, JE