DNA-measuring Wadjet SMC ATPases restrict smaller circular plasmids by DNA cleavage

Structural maintenance of chromosomes (SMC) complexes fold DNA by loop extrusion to support chromosome segregation and genome maintenance. Wadjet systems (JetABCD/MksBEFG/EptABCD) are derivative SMC complexes with roles in bacterial immunity against selfish DNA. Here, we show that JetABCD restricts circular plasmids with an upper size limit of about 100 kb, while a linear plasmid evades restriction. Purified JetABCD complexes cleave circular DNA molecules regardless of DNA helical topology; cleavage is DNA sequence-nonspecific and depends on the SMC ATPase. A cryo-EM structure reveals a distinct JetABC dimer-of-dimers geometry with the two SMC dimers facing in opposite direction—rather than the same as observed with MukBEF. We hypothesize that JetABCD is a DNA-shape-specific endonuclease and propose the ‘total extrusion model’ for DNA cleavage exclusively when extrusion of an entire plasmid has been completed by a JetABCD complex. Total extrusion cannot be achieved on the larger chromosome, explaining how self-DNA may evade processing.染色体结构维持（Structural maintenance of chromosomes, SMC）复合物通过环挤出（loop extrusion）过程折叠DNA，以支持染色体分离与基因组维持。沃杰特系统（Wadjet systems，JetABCD/MksBEFG/EptABCD）是一类衍生的SMC复合物，在细菌抵御自私DNA的免疫防御中发挥功能。本研究证实，JetABCD可限制性抑制大小上限约100 kb的环状质粒，而线性质粒则可逃逸其抑制作用。纯化的JetABCD复合物能够切割环状DNA分子，该过程不受DNA螺旋拓扑结构影响，且无DNA序列特异性，并依赖SMC ATP酶的活性。冷冻电镜结构显示，JetABC呈现独特的二聚体-二聚体空间构象：两个SMC二聚体的朝向彼此相反，这与MukBEF复合物中观察到的同向二聚体排布截然不同。我们提出假说，认为JetABCD是一类DNA形状特异性核酸内切酶，并提出“完全环挤出模型”：仅当JetABCD复合物完成对整个质粒的环挤出时，才会介导DNA切割。由于大型宿主染色体无法实现完全环挤出，这解释了自身DNA如何逃逸JetABCD的加工过程。