Regulation of a calcium-dependent protein kinase in response to osmotic stress

Fan et al. explore how ubiquitin-dependent proteolysis regulates the levels of active kinase in plant responses to osmotic stress.

Wei Fan and Zixing Li

School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai 200240, China

Background: Plants experience osmotic stress during drought conditions and in soils with high salt levels. Osmotic stress triggers a rapid rise in intracellular calcium ion (Ca2+) concentrations in plants. Upon the binding of Ca2+, the calcium-dependent protein kinase (CPK) undergoes a conformational change, releasing the kinase active site from the repression of an autoinhibitory junction. CPKs become active to phosphorylate their downstream targets to transduce osmotic stress signals. CPK4 plays a positive role in plant osmotic stress responses.

Question: How are active CPK4 protein levels dynamically and precisely regulated in response to hyperosmotic stress?

Findings: We found that salt/mannitol-induced osmotic stress promotes CPK4 protein accumulation by disrupting 26S proteasome-mediated degradation of CPK4. The U-BOX E3 ligase PUB44 interacts with and mediates the ubiquitination of CPK4, which results in the proteasomal degradation of CPK4. PUB44 exhibits a CPK4-dependent negative role in plant osmotic stress responses. In addition, we found that Ca-binding and kinase activation decrease the ubiquitination of CPK4 and enhance CPK4 protein stabilization.

Next steps: We plan to investigate how the binding of Ca2+ to CPK determines the ubiquitination and degradation of CPK4.


Wei Fan, Xiliang Liao, Yanqiu Tan, Xiruo Wang, Julian I. Schroeder, Zixing Li. (2023). Arabidopsis PLANT U-BOX44 down-regulates osmotic stress signaling by mediating Ca2+-DEPENDENT PROTEIN KINASE4 degradation.

背景回顾:渗透胁迫条件下,植物胞质自由钙离子浓度会快速上升。钙依赖的蛋白激酶(calcium-dependent protein kinase,CPK)结合细胞内钙离子,蛋白构象发生改变,释放自抑制域对激酶活性的抑制,并通过自磷酸化或其他激酶的磷酸化而激活,激活的钙依赖的蛋白激酶磷酸化下游底物蛋白,激发植物渗透胁迫信号,使植物响应渗透胁迫。CPK4在植物响应干旱胁迫时作为正调因子发挥重要作用,促使植物细胞调节自身生理活动以响应环境渗透胁迫


研究发现:我们鉴定到一个U-BOX类型的E3泛素连接酶PUB44,能够识别并泛素化修饰CPK4 促其通过26S蛋白酶体途径降解,调节细胞内CPK4的蛋白丰度。渗透胁迫信号下,CPK4结合钙离子,蛋白发生构象改变,减弱PUB44泛素连接酶对CPK4蛋白的泛素化修饰程度。同时,渗透胁迫信号还会抑制PUB44蛋白的泛素酶活性,进一步降低CPK4蛋白的泛素化修饰水平。通过这些方式,植物细胞在正常环境下维持一定CPK4蛋白的丰度,并在胁迫条件下快速增加CPK4蛋白的含量,激活细胞适应性生理活动来响应渗透胁迫