CONTROL OF CALCIUM AND PHOSPHORYLATION-REGULATED SIGNALLING PATHWAYS

钙调素依赖性蛋白磷酸酶的分子研究， 钙调神经磷酸酶（CN），已经确定了3个哺乳动物基因的催化（A） 亚基，并确定其在人类染色体定位。 许多门中A亚基基因的特征表明哺乳动物 这些形式起源于一个祖先系，包括丝状真菌 （脉孢菌属，曲霉属），并表明基因分歧发生后， 鸟类/爬行动物分支点。调节（B）亚基的两个基因 其中一个与A亚基的同种型共表达 在睾丸发育过程中，这两个亚基可能构成一个 与精子鞭毛相关的同工酶。生化研究 在细菌和昆虫细胞中表达的重组蛋白， B亚单位同种型可以与不同的 A亚基和真菌催化亚基。这些数据表明 B亚基相互作用结构域的功能保守性， 在绘图研究中被描绘出来。CN的重要生物学作用 是建立在T细胞调节上的。在转染了 A亚基的组成型活性突变体，白细胞介素（IL）-2 启动子被佛波醇酯协同激活，这表明 CN在T细胞受体介导的调节IL-1的信号传导事件中的作用 2基因转录。这些细胞也能抵抗 免疫抑制剂，FK-506和环孢菌素A，与最近的 发现CN与细胞溶质药物受体形成复合物 因此，CN是这些药物在T细胞中的主要靶标。 细胞从小鼠心肌细胞中克隆了钙调素调节的磷酸二酯酶（PDE）基因， PCR扩增小脑浦肯野细胞mRNA 细胞层原位杂交和北方印迹分析表明， 高表达的PDE mRNA在纹状体，这表明在控制作用 多巴胺介导的环磷酸腺苷途径。肽微测序是 完成了一个新的36 kDa钙调素结合蛋白分离自布赖恩， 显示与任何的黑曲霉蛋白质都没有同源性Western blot分析 抗肽抗体在脑、脾和 肾

Molecular studies of the calmodulin (CaM)-dependent protein phosphatase, calcineurin (CN), have identified 3 mammalian genes for the catalytic (A) subunit and determined their chromosomal localization in humans. Characterization of A subunit genes in many phyla indicate that mammalian forms arose from an ancestral line that includes the filamentous fungi (Neurospora, Aspergillus) and suggests that gene divergence occurred after the avian/reptilian branch point. Two genes for the regulatory (B) subunit were cloned, one of which is co-expressed with an isoform of the A subunit during testicular development; these two subunits may comprise an isoenzyme that is associated with the sperm flagellum. Biochemical studies with recombinant proteins, expressed in bacteria and in insect cells, show that the B subunit isoforms can associate interchangeably with different A subunits and with a fungal catalytic subunit. These data indicate functional conservation of the B subunit interaction domain, which has now been delineated in mapping studies. An important biological role for CN was established in T-cell regulation. In human cells transfected with a constitutively-active mutant of the A subunit, the interleukin (IL)-2 promoter was activated synergistically by phorbol esters; this suggests a role for CN in T-cell receptor-mediated signaling events that regulate IL- 2 gene transcription. These cells also were resistant to the immunosuppressants, FK-506 and cyclosporin A, in agreement with recent findings that CN forms complexes with the cytosolic drug receptors ("immunophilins"); thus, CN is the primary target of these drugs in T- cells. A CaM-regulated phosphodiesterase (PDE) was cloned from murine brain by PCR amplification of mRNA from microdissected cerebellar Purkinje cell layers. In situ hybridization and Northern blot analysis indicate high expression of PDE mRNA in the striatum, suggesting a role in control of dopamine-mediated cyclic AMP pathways. Peptide microsequencing was completed on a novel 36 kDa CaM-binding protein isolated from brian, showing no homology to any mammlaina proteins. Western blot analysis with anti-peptide antibodies showed highest amounts in brain, spleen and kidney.