Hoch B, Wobus A M, Krause E G, Karczewski P
Max Delbrück Center for Molecular Medicine, D-13092 Berlin-Buch, Germany.
J Cell Biochem. 2000 Aug 2;79(2):293-300.
delta-isoforms of the Ca(2+)/calmodulin-dependent protein kinase type II (CaMKII) are considered to substantially influence cardiac functions. However, no data exist on the expression of these isoforms in the mouse heart. We analyzed the transcript pattern of non-neuronally expressed delta-isoforms in heart and skeletal muscle of adult mice by RT-PCR. For members of the delta-CaMKII subclass with both variable domains (subclass II), weak transcriptional expression of isoforms delta(2) and delta(3) was found in the heart. In skeletal muscle no delta(3)-specific transcript was detectable. In cardiac tissue, stronger signals result from amplifications of delta(9) and from members of the subclass I lacking the second variable domain. Western blotting was performed using a subclass II-specific antibody. In murine cardiac and skeletal muscle tissue a delta-CaMKII protein pattern was obtained similar to that described for rat. To gain insight into the expression of delta-CaMKII during the earliest steps of cardiogenic differentiation, we analyzed the transcript pattern of murine embryonic stem cell-derived cardiomyocytes in various differentiation stages. Reproducible RT-PCR signals could be obtained for delta(6) and delta(10), both belonging to the delta-CaMKII subclass I. Transcripts for delta(6) were ubiquitously expressed, whereas transcripts for delta(10) were detectable in increasing amounts after 7-10 days of the onset of cardiogenic differentiation. Our results point to a differentiation-dependent expression of the two delta-CaMKII subclasses, and also to differences in the expression of individual members of subclass I during the early stages of cardiogenic differentiation.
钙/钙调蛋白依赖性蛋白激酶II型(CaMKII)的δ亚型被认为对心脏功能有重大影响。然而,关于这些亚型在小鼠心脏中的表达尚无数据。我们通过逆转录聚合酶链反应(RT-PCR)分析了成年小鼠心脏和骨骼肌中非神经元表达的δ亚型的转录模式。对于具有可变结构域的δ-CaMKII亚类成员(亚类II),在心脏中发现亚型δ(2)和δ(3)的转录表达较弱。在骨骼肌中未检测到δ(3)特异性转录本。在心脏组织中,δ(9)的扩增以及缺乏第二个可变结构域的亚类I成员产生更强的信号。使用亚类II特异性抗体进行蛋白质印迹分析。在小鼠心脏和骨骼肌组织中获得的δ-CaMKII蛋白模式与大鼠中描述的相似。为了深入了解δ-CaMKII在心脏发生分化最早阶段的表达情况,我们分析了处于不同分化阶段的小鼠胚胎干细胞衍生心肌细胞的转录模式。对于属于δ-CaMKII亚类I的δ(6)和δ(10),可以获得可重复的RT-PCR信号。δ(6)的转录本普遍表达,而δ(10)的转录本在心脏发生分化开始7 - 10天后可检测到的量逐渐增加。我们的结果表明两个δ-CaMKII亚类的表达依赖于分化,并且在心脏发生分化早期亚类I的各个成员的表达也存在差异。