Bowlby M R, Fadool D A, Holmes T C, Levitan I B
Department of Biochemistry and Volen Center for Complex Systems, Brandeis University, Waltham, Massachusetts 02254, USA.
J Gen Physiol. 1997 Nov;110(5):601-10. doi: 10.1085/jgp.110.5.601.
The voltage-dependent potassium channel, Kv1.3, is modulated by the epidermal growth factor receptor (EGFr) and the insulin receptor tyrosine kinases. When the EGFr and Kv1.3 are coexpressed in HEK 293 cells, acute treatment of the cells with EGF during a patch recording can suppress the Kv1.3 current within tens of minutes. This effect appears to be due to tyrosine phosphorylation of the channel, as it is blocked by treatment with the tyrosine kinase inhibitor erbstatin, or by mutation of the tyrosine at channel amino acid position 479 to phenylalanine. Previous work has shown that there is a large increase in the tyrosine phosphorylation of Kv1.3 when it is coexpressed with the EGFr. Pretreatment of EGFr and Kv1.3 cotransfected cells with EGF before patch recording also results in a decrease in peak Kv1.3 current. Furthermore, pretreatment of cotransfected cells with an antibody to the EGFr ligand binding domain (alpha-EGFr), which blocks receptor dimerization and tyrosine kinase activation, blocks the EGFr-mediated suppression of Kv1.3 current. Insulin treatment during patch recording also causes an inhibition of Kv1.3 current after tens of minutes, while pretreatment for 18 h produces almost total suppression of current. In addition to depressing peak Kv1.3 current, EGF treatment produces a speeding of C-type inactivation, while pretreatment with the alpha-EGFr slows C-type inactivation. In contrast, insulin does not influence C-type inactivation kinetics. Mutational analysis indicates that the EGF-induced modulation of the inactivation rate occurs by a mechanism different from that of the EGF-induced decrease in peak current. Thus, receptor tyrosine kinases differentially modulate the current magnitude and kinetics of a voltage-dependent potassium channel.
电压依赖性钾通道Kv1.3受表皮生长因子受体（EGFr）和胰岛素受体酪氨酸激酶的调节。当EGFr和Kv1.3在HEK 293细胞中共表达时，在膜片钳记录过程中用表皮生长因子（EGF）急性处理细胞，可在数十分钟内抑制Kv1.3电流。这种效应似乎是由于通道的酪氨酸磷酸化，因为它可被酪氨酸激酶抑制剂埃博霉素处理或通道氨基酸位置479处的酪氨酸突变为苯丙氨酸所阻断。先前的研究表明，当Kv1.3与EGFr共表达时，其酪氨酸磷酸化会大幅增加。在膜片钳记录前用EGF对EGFr和Kv1.3共转染的细胞进行预处理，也会导致Kv1.3电流峰值降低。此外，用针对EGFr配体结合域的抗体（α-EGFr）对共转染细胞进行预处理，可阻断受体二聚化和酪氨酸激酶激活，从而阻断EGFr介导的对Kv1.3电流的抑制。在膜片钳记录过程中用胰岛素处理也会在数十分钟后抑制Kv1.3电流，而预处理18小时则几乎完全抑制电流。除了降低Kv1.3电流峰值外，EGF处理还会加快C型失活，而用α-EGFr预处理则会减慢C型失活。相比之下，胰岛素不影响C型失活动力学。突变分析表明，EGF诱导的失活速率调节机制与EGF诱导的电流峰值降低机制不同。因此，受体酪氨酸激酶对电压依赖性钾通道的电流幅度和动力学有不同的调节作用。
