Ubiquitination is a reversible post-translational protein modification, which can be regulated by a variety of signal factors and widely involved in almost every important cellular process. The linear ubiquitin chain assembly complex (LUBAC), which consists of a catalytic subunit HOIP and two regulatory subunits HOIL-1L and SHARPIN, is the only currently identified E3 ligase complex capable of catalyzing linear poly-Ub chains on protein substrates. LUBAC can be recruited into multiple signaling pathways to regulate the transduction of upstream and downstream signaling pathways. The linear poly-Ub chains generated by LUBAC can be act as scaffolds to recruit effector proteins, thereby widely participating in NF-κB and other signaling pathways, and play pivotal roles in innate immunity, cell death and antibacterial selective autophagy (xenophagy). Shigella flexneri, a gram-negative bacterium, is the major culprit of bacterial shigellosis, and causes many deaths annually. For evading the host immune response during infection, Shigella flexneri secrets two highly similar E3 ligases, IpaH1.4 and IpaH2.5, to subvert the linear ubiquitin chain assembly complex (LUBAC) of host cells. However, the detailed molecular mechanism underpinning the subversion of LUBAC by IpaH1.4/2.5 remains elusive.

 

Recently, a research paper titled “Mechanistic insights into the subversion of the linear ubiquitin chain assembly complex by the E3 ligase IpaH1.4 of Shigella flexneri” (https://www.pnas.org/doi/full/10.1073/pnas.2116776119) was published on the PNAS journal by Professor PAN Lifeng’s group from Shanghai Institute of Organic Chemistry, CAS. In this paper, using various biochemical techniques, such as Nuclear Magnetic Resonance (NMR), Isothermal Titration Calorimetry (ITC), and Fast Protein Liquid Chromatography (FPLC), they discovered that, IpaH1.4 specifically recognizes HOIP and HOIL-1L through its LRR domain by binding to the HOIP RING1 domain and HOIL-1L UBL domain, respectively. They determined the crystal structures of IpaH1.4 LRR/HOIP RING1 complex, IpaH1.4 LRR/HOIL-1L UBL complex and HOIP RING1/UBE2L3 complex for the first time, elucidated the binding mechanisms of IpaH1.4 with HOIP and HOIL-1L, and unveiled that the recognition of HOIP by IpaH1.4 can inhibit the E2-binding of HOIP. Additionally, they demonstrated that the interaction of IpaH1.4 LRR with HOIP RING1 or HOIL-1L UBL is essential for the ubiquitination of HOIP or HOIL-1L in vitro as well as the suppression of NF-κB activation by IpaH1.4 in cells. In summary, this work elucidated that three different strategies were adopted by IpaH1.4 simultaneously to disrupt the function of LUBAC, thereby inhibiting the activation of NF-κB signaling pathway in host cells and antibacterial selective autophagy: 1) mediates the K48-linked ubiquitination of HOIP for proteasomal degradation; 2) inhibits the E3 activity of LUBAC by blocking the E2-binding of HOIP; 3) suppresses the capacity of HOIL-1L binding to HOIP and disturbs the stability of LUBAC, by acting on the UBL domain of HOIL-1L as well as mediating K48-linked ubiquitination and subsequent proteasomal degradation of HOIL-1L. In summary, this study revealed, for the first time, the molecular mechanism underlying the multiple tactics adopted by the E3 ligase IpaH1.4 of S. flexneri to hijack the LUBAC complex of host cells from biochemical and structural perspective, and provided a new potential target for drug development against S. flexneri infection in the future.

 

The associate researcher Dr. LIU Jianping, the PhD student WANG Yaru from Professor PAN’s group and the PhD student WANG Danni from Professor YAO’s group are the co-authors of this paper. This work was supported by grants from the National Natural Science Foundation of China (31470749, 21621002, 91753113, 21822705), Science and Technology Commission of Shanghai Municipality (17JC1405200), Strategic Priority Research Program of the Chinese Academy of Sciences (XDB20000000), and the start-up fund from the State Key Laboratory of Bioorganic and Natural Products Chemistry and Chinese Academy of Sciences.

 

 

Figure 1. Mechanistic insights into the subversion of the linear ubiquitin chain assembly complex by the E3 ligase IpaH1.4/2.5 of Shigella flexneri(Image by PAN Lifeng)

 

PAN Lifeng Ph.D.Professor

Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences

Ling Ling Road 345 Shanghai 200032 China

Tel: 0086-21-54925561

Email: panlf@sioc.ac.cn