Immunohistochemical studies of hearts from the lesser spotted dogfish, Scyliorhinus canicula (Chondrichthyes) revealed that the pan-myosin heavy chain (pan-MyHC) antibody MF20 homogeneously labels all the myocardium, while the pan-MyHC antibody A4.1025 labels the myocardium of the inflow (sinus venosus and atrium) but not the outflow (ventricle and conus arteriosus) cardiac segments, as opposed to other vertebrates. We hypothesized that the conventional pattern of cardiac MyHC isoform distribution present in most vertebrates, i.e. MYH6 in the inflow and MYH7 in the outflow segments, has evolved from a primitive pattern that persists in Chondrichthyes. In order to test this hypothesis, we conducted protein detection techniques to identify the MyHC isoforms expressed in adult dogfish cardiac segments and to assess the pan-MyHC antibodies reactivity against the cardiac segments of representative species from different vertebrate groups. Western and slot blot results confirmed the specificity of MF20 and A4.1025 for MyHC in dogfish and their differential reactivity against distinct myocardial segments. HPLC-ESI-MS/MS and ESI-Quadrupole-Orbitrap revealed abundance of MYH6 and MYH2 in the inflow and of MYH7 and MYH7B in the outflow segments. Immunoprecipitation showed higher affinity of A4.1025 for MYH2 and MYH6 than for MYH7 and almost no affinity for MYH7B. Immunohistochemistry showed that A4.1025 signals are restricted to the inflow myocardial segments of elasmobranchs, homogeneous in all myocardial segments of teleosts and acipenseriforms, and low in the ventricle of polypteriforms. The cardiac inflow and outflow segments of the dogfish show predominance of fast- and slow-twitch MyHC isoforms respectively, what can be considered a synapomorphy of gnathostomes. The myocardium of the dogfish contains two isomyosins (MYH2 and MYH7B) not expressed in the adult heart of other vertebrates. We propose that these isomyosins lost their function in cardiac contraction during the evolution of gnathostomes, the later acquiring a regulatory role in myogenesis through its intronic miRNA. Loss of MYH2 and MYH7B expression in the heart possibly occurred before the origin of Osteichthyes, being the latter reacquired in polypteriforms. We raise the hypothesis that the slow tonic MYH7B facilitates the peristaltic contraction of the conus arteriosus of fish with a primitive cardiac anatomical design and of the vertebrate embryo.

中文翻译：

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有颌脊椎动物心脏节段肌球蛋白重链亚型的差异表达及其进化意义

对小斑角鲨 (Chondrichthyes) 心脏的免疫组织化学研究表明，泛肌球蛋白重链 (pan-MyHC) 抗体 MF20 均匀标记所有心肌，而泛 MyHC 抗体 A4.1025 标记与其他脊椎动物相反，流入（静脉窦和心房）而不是流出（心室和动脉圆锥）心脏部分。我们假设大多数脊椎动物中存在的心脏 MyHC 亚型分布的常规模式，即流入段中的 MYH6 和流出段中的 MYH7，已经从软骨鱼中持续存在的原始模式演变而来。为了检验这个假设，我们进行了蛋白质检测技术，以鉴定成年角鲨心脏节段中表达的 MyHC 同种型，并评估泛 MyHC 抗体对来自不同脊椎动物群体的代表性物种的心脏节段的反应性。Western 和槽印迹结果证实了 MF20 和 A4.1025 对角鲨中 MyHC 的特异性及其对不同心肌节段的不同反应性。HPLC-ESI-MS/MS 和 ESI-Quadrupole-Orbitrap 揭示了流入段中 MYH6 和 MYH2 的丰度以及流出段中的 MYH7 和 MYH7B 的丰度。免疫沉淀显示 A4.1025 对 MYH2 和 MYH6 的亲和力高于对 MYH7 的亲和力，而对 MYH7B 几乎没有亲和力。免疫组织化学显示 A4.1025 信号仅限于弹性分支的流入心肌节段，在硬骨鱼和鲟形的所有心肌节段中均质，在鳍状肢的心室中低。角鲨的心脏流入和流出段分别显示出快速和慢速抽搐的 MyHC 亚型的优势，这可以被认为是 gnathostomes 的突触。角鲨的心肌含有两种在其他脊椎动物的成年心脏中不表达的同肌球蛋白（MYH2 和 MYH7B）。我们提出这些同肌球蛋白在有颚动物进化过程中失去了它们在心脏收缩中的功能，后者通过其内含子 miRNA 在肌发生中获得调节作用。心脏中 MYH2 和 MYH7B 表达的缺失可能发生在骨鱼的起源之前，后者在多翅目中重新获得。