Ubiquitylation is a widespread post-translational protein modification in eukaryotes and marks bacteria that invade the cytosol as cargo for antibacterial autophagy^1,2,3. The identity of the ubiquitylated substrate on bacteria is unknown. Here we show that the ubiquitin coat on Salmonella that invade the cytosol is formed through the ubiquitylation of a non-proteinaceous substrate, the lipid A moiety of bacterial lipopolysaccharide (LPS), by the E3 ubiquitin ligase ring finger protein 213 (RNF213). RNF213 is a risk factor for moyamoya disease^4,5, which is a progressive stenosis of the supraclinoid internal carotid artery that causes stroke (especially in children)^6,7. RNF213 restricts the proliferation of cytosolic Salmonella and is essential for the generation of the bacterial ubiquitin coat, both directly (through the ubiquitylation of LPS) and indirectly (through the recruitment of LUBAC, which is a downstream E3 ligase that adds M1-linked ubiquitin chains onto pre-existing ubiquitin coats⁸). In cells that lack RNF213, bacteria do not attract ubiquitin-dependent autophagy receptors or induce antibacterial autophagy. The ubiquitylation of LPS on Salmonella that invade the cytosol requires the dynein-like core of RNF213, but not its RING domain. Instead, ubiquitylation of LPS relies on an RZ finger in the E3 shell. We conclude that ubiquitylation extends beyond protein substrates and that ubiquitylation of LPS triggers cell-autonomous immunity, and we postulate that non-proteinaceous substances other than LPS may also become ubiquitylated.

泛素化是真核生物中广泛存在的翻译后蛋白质修饰，将侵入胞质溶胶的细菌标记为抗菌自噬的货物^1,2,3。细菌上泛素化底物的身份未知。在这里，我们表明侵入胞质溶胶的沙门氏菌上的泛素外壳是通过 E3 泛素连接酶无名指蛋白 213 (RNF213) 泛素化非蛋白质底物（细菌脂多糖 (LPS) 的脂质 A 部分）形成的。RNF213 是烟雾病的危险因素^4,5，这是一种导致中风（尤其是儿童）的颈内动脉床突上进行性狭窄^6,7。RNF213 限制细胞溶质的增殖沙门氏菌对细菌泛素外壳的生成至关重要，直接（通过 LPS 的泛素化）和间接（通过 LUBAC 的募集，LUBAC 是一种下游 E3 连接酶，可将 M1 连接的泛素链添加到预先存在的泛素外壳上⁸ ). 在缺乏 RNF213 的细胞中，细菌不会吸引泛素依赖性自噬受体或诱导抗菌自噬。LPS 对沙门氏菌的泛素化侵入胞质溶胶需要 RNF213 的动力蛋白样核心，但不需要其 RING 域。相反，LPS 的泛素化依赖于 E3 外壳中的 RZ 指状物。我们得出结论，泛素化延伸到蛋白质底物之外，LPS 的泛素化触发细胞自主免疫，我们假设 LPS 以外的非蛋白质物质也可能泛素化。