Goethe University Frankfurt, Medical Clinic 1, Biomedical Research Laboratory, University Hospital, Frankfurt, Germany.
German Translational Cancer Network (DKTK), Frankfurt, Germany.
Protein Sci. 2024 Jan;33(1):e4840. doi: 10.1002/pro.4840.
Autophagy is a highly conserved cellular process that allows degradation of large macromolecules. p62/SQSTM1 is a key adaptor protein that interacts both with material to be degraded and with LC3 at the autophagosome, enabling degradation of cargos such as protein aggregates, lipid droplets and damaged organelles by selective autophagy. Dysregulation of autophagy contributes to the pathogenesis of many diseases. In this study, we investigated if the interaction of p62/SQSTM1 with LC3B could be regulated. We purified full-length p62/SQSTM1 and established an in vitro assay that measures the interaction with LC3B. We used the assay to determine the role of the different domains of p62/SQSTM1 in the interaction with LC3B. We identified a mechanism of regulation of p62/SQSTM1 where the ZZ and the PB1 domains regulate the exposure of the LIR-sequence to enable or inhibit the interaction with LC3B. A mutation to mimic the phosphorylation of a site on the ZZ domain leads to increased interaction with LC3B. Also, a small compound that binds to the ZZ domain enhances interaction with LC3B. Dysregulation of these mechanisms in p62/SQSTM1 could have implications for diseases where autophagy is affected. In conclusion, our study highlights the regulated nature of p62/SQSTM1 and its ability to modulate the interaction with LC3B through a LIR-sequence Accessibility Mechanism (LAM). Furthermore, our findings suggest the potential for pharmacological modulation of the exposure of LIR, paving the way for future therapeutic strategies.
自噬是一种高度保守的细胞过程，允许降解大分子物质。p62/SQSTM1 是一种关键的衔接蛋白，它与待降解的物质以及自噬体中的 LC3 相互作用，使蛋白聚集体、脂滴和受损细胞器等货物通过选择性自噬降解。自噬的失调与许多疾病的发病机制有关。在这项研究中，我们研究了 p62/SQSTM1 与 LC3B 的相互作用是否可以受到调节。我们纯化了全长的 p62/SQSTM1，并建立了一种体外测定方法来测量与 LC3B 的相互作用。我们使用该测定法来确定 p62/SQSTM1 的不同结构域在与 LC3B 的相互作用中的作用。我们确定了一种 p62/SQSTM1 调节机制，其中 ZZ 和 PB1 结构域调节 LIR 序列的暴露，以允许或抑制与 LC3B 的相互作用。模拟 ZZ 结构域上一个位点磷酸化的突变导致与 LC3B 的相互作用增加。此外，一种结合 ZZ 结构域的小分子化合物增强了与 LC3B 的相互作用。p62/SQSTM1 中这些机制的失调可能对自噬受到影响的疾病有影响。总之，我们的研究强调了 p62/SQSTM1 的调节性质及其通过 LIR 序列可及性机制（LAM）调节与 LC3B 相互作用的能力。此外，我们的发现表明，通过药理学调节 LIR 的暴露可能为未来的治疗策略铺平道路。
