Nucleotide and protein sequences for dog masticatory tropomyosin identify a novel Tpm4 gene product.
Nucleotide and protein sequences for dog masticatory tropomyosin identify a novel Tpm4 gene product.
复制标题
DOI：
10.1007/s10974-015-9425-1
复制
发表时间：
2015-10
影响因子：
2.7
通讯作者：
Reiser PJ
中科院分区：
文献类型：
--
作者：
Brundage EA;Biesiadecki BJ;Reiser PJ
几种脊椎动物物种（包括食肉动物）的下颌闭合肌表达肌球蛋白重链的独特的“咀嚼”同种型，沿着在大多数其它肌肉中不表达的其它肌原纤维蛋白的同种型。一般认为，这些肌肉中的肌原纤维同种型的补充用于捕获活猎物、分解坚韧的植物材料和防御性咬的高力量产生。一个独特的异构体原肌球蛋白（Tpm）的报告，表达在猫下颌闭合肌肉，基于二维凝胶流动性，肽图谱，和免疫组织化学。本研究的目的是获得这种独特的Tpm亚型的蛋白质和基因序列信息。咬肌（也是下颌闭合肌）、胫骨肌（主要是快缩纤维）和深外侧腓肠肌（主要是慢缩纤维）的样本均来自成年犬。克隆表达的Tpm同种型，测序产生与基因组预测的横纹肌Tpm 1.1 St相同的cDNA（a，B，B，a）（历史上称为αTpm）、Tpm 2.2 St（a，B，B，a）（βTpm）和Tpm 3.12 St（a，B，B，a）（cTpm）亚型（命名反映了主要的组织表达（“St”-横纹肌）和外显子剪接模式），以及在下颌闭合肌中观察到的新的284个氨基酸的同种型，其与Tpm 4基因（δTpm）家族的基因组预测产物相同。新的同种型被命名为Tpm 4.3St（a，B，B，a）。与其他骨骼Tpm亚型相比，在狗咬肌中表达的肌原纤维Tpm亚型在含有6 M尿素的凝胶上表现出独特的电泳迁移率。为了验证克隆的Tpm 4.3亚型是在狗咬肌中表达的Tpm，E.大肠杆菌表达的Tpm4.3在尿素存在下进行纯化。结果表明，Tpm 4.3具有相同的电泳迁移率的独特的狗咬肌Tpm异构体和不同的迁移率的肌肉Tpm 1.1，Tpm 2.2和Tpm 3.12异构体。我们的结论是，独特的Tpm亚型在狗咬肌是Tpm 4基因的产物，该基因的284个氨基酸的蛋白质产物代表了一种新的肌原纤维Tpm亚型从未观察到在横纹肌中表达。
Jaw-closing muscles of several vertebrate species, including members of Carnivora, express a unique, “masticatory”, isoform of myosin heavy chain, along with isoforms of other myofibrillar proteins that are not expressed in most other muscles. It is generally believed that the complement of myofibrillar isoforms in these muscles serves high force generation for capturing live prey, breaking down tough plant material and defensive biting. A unique isoform of tropomyosin (Tpm) was reported to be expressed in cat jaw-closing muscle, based upon two-dimensional gel mobility, peptide mapping, and immunohistochemistry. The objective of this study was to obtain protein and gene sequence information for this unique Tpm isoform. Samples of masseter (also a jaw-closing muscle), tibialis (with predominantly fast-twitch fibers), and the deep lateral gastrocnemius (predominantly slow-twitch fibers) were obtained from adult dogs. Expressed Tpm isoforms were cloned and sequencing yielded cDNAs that were identical to genomic predicted striated muscle Tpm1.1St(a,b,b,a) (historically referred to as αTpm), Tpm2.2St(a,b,b,a) (βTpm) and Tpm3.12St(a,b,b,a) (cTpm) isoforms (nomenclature reflects predominant tissue expression (“St”—striated muscle) and exon splicing pattern), as well as a novel 284 amino acid isoform observed in jaw-closing muscle that is identical to a genomic predicted product of the Tpm4 gene (δTpm) family. The novel isoform is designated as Tpm4.3St(a,b,b,a). The myofibrillar Tpm isoform expressed in dog masseter exhibits a unique electrophoretic mobility on gels containing 6 M urea, compared to other skeletal Tpm isoforms. To validate that the cloned Tpm4.3 isoform is the Tpm expressed in dog masseter, E. coli-expressed Tpm4.3 was electrophoresed in the presence of urea. Results demonstrate that Tpm4.3 has identical electrophoretic mobility to the unique dog masseter Tpm isoform and is of different mobility from that of muscle Tpm1.1, Tpm2.2 and Tpm3.12 isoforms. We conclude that the unique Tpm isoform in dog masseter is a product of the Tpm4 gene and that the 284 amino acid protein product of this gene represents a novel myofibrillar Tpm isoform never before observed to be expressed in striated muscle.