Terminal regions of Drosophila embryos are patterned by signaling through ERK, which is genetically deregulated in multiple human diseases. Quantitative studies of terminal patterning have been recently used to investigate gain-of-function variants of human MEK1, encoding the MEK kinase that directly activates ERK by dual phosphorylation. Unexpectedly, several mutations reduced ERK activation by extracellular signals, possibly through a negative feedback triggered by signal-independent activity of the mutant variants. Here we present experimental evidence supporting this model. Using a MEK variant that combines a mutation within the negative regulatory region with alanine substitutions in the activation loop, we prove that pathogenic variants indeed acquire signal-independent kinase activity. We also demonstrate that signal-dependent activation of these variants is independent of KSR, a conserved adaptor that is indispensable for activation of normal MEK. Finally, we show that attenuation of ERK activation by extracellular signals stems from transcriptional induction of Mkp3, a dual specificity phosphatase that deactivates ERK by dephosphorylation. These findings in the Drosophila embryo highlight its power for investigating diverse effects of human disease mutations.

果蝇的末端区域胚胎是通过 ERK 信号形成的，ERK 在多种人类疾病中基因失调。末端模式的定量研究最近已被用于研究人类 MEK1 的功能获得变体，编码通过双重磷酸化直接激活 ERK 的 MEK 激酶。出乎意料的是，一些突变降低了细胞外信号对 ERK 的激活，这可能是通过突变变体的信号独立活动触发的负反馈。在这里，我们提供了支持该模型的实验证据。使用将负调节区内的突变与激活环中的丙氨酸取代相结合的 MEK 变体，我们证明了致病性变体确实获得了信号独立的激酶活性。我们还证明这些变体的信号依赖性激活与 KSR 无关，KSR 是一种保守的适配器，对于正常 MEK 的激活必不可少。最后，我们表明细胞外信号对 ERK 激活的减弱源于 Mkp3 的转录诱导，Mkp3 是一种通过去磷酸化使 ERK 失活的双重特异性磷酸酶。这些发现在果蝇胚胎突出了其研究人类疾病突变的多种影响的能力。