State Key Laboratory of Chemical Biology, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200032, China.
School of Physical Science and Technology, ShanghaiTech University, Shanghai 201210, China.
Proc Natl Acad Sci U S A. 2024 Sep 10;121(37):e2402817121. doi: 10.1073/pnas.2402817121. Epub 2024 Sep 5.
Autophagy of glycogen (glycophagy) is crucial for the maintenance of cellular glucose homeostasis and physiology in mammals. STBD1 can serve as an autophagy receptor to mediate glycophagy by specifically recognizing glycogen and relevant key autophagic factors, but with poorly understood mechanisms. Here, we systematically characterize the interactions of STBD1 with glycogen and related saccharides, and determine the crystal structure of the STBD1 CBM20 domain with maltotetraose, uncovering a unique binding mode involving two different oligosaccharide-binding sites adopted by STBD1 CBM20 for recognizing glycogen. In addition, we demonstrate that the LC3-interacting region (LIR) motif of STBD1 can selectively bind to six mammalian ATG8 family members. We elucidate the detailed molecular mechanism underlying the selective interactions of STBD1 with ATG8 family proteins by solving the STBD1 LIR/GABARAPL1 complex structure. Importantly, our cell-based assays reveal that both the STBD1 LIR/GABARAPL1 interaction and the intact two oligosaccharide binding sites of STBD1 CBM20 are essential for the effective association of STBD1, GABARAPL1, and glycogen in cells. Finally, through mass spectrometry, biochemical, and structural modeling analyses, we unveil that STBD1 can directly bind to the Claw domain of RB1CC1 through its LIR, thereby recruiting the key autophagy initiation factor RB1CC1. In all, our findings provide mechanistic insights into the recognitions of glycogen, ATG8 family proteins, and RB1CC1 by STBD1 and shed light on the potential working mechanism of STBD1-mediated glycophagy.
自噬糖原（glycophagy）对于维持哺乳动物细胞内葡萄糖稳态和生理功能至关重要。STBD1 可以作为自噬受体，通过特异性识别糖原和相关关键自噬因子来介导 glycophagy，但具体机制尚不清楚。在这里，我们系统地研究了 STBD1 与糖原和相关糖的相互作用，并确定了 STBD1 CBM20 结构域与麦芽四糖的晶体结构，揭示了 STBD1 CBM20 采用的独特结合模式，涉及两个不同的寡糖结合位点，用于识别糖原。此外，我们证明了 STBD1 的 LC3 相互作用区域（LIR）基序可以选择性地结合六种哺乳动物 ATG8 家族成员。我们通过解析 STBD1 LIR/GABARAPL1 复合物结构，阐明了 STBD1 与 ATG8 家族蛋白选择性相互作用的详细分子机制。重要的是，我们的细胞实验揭示了 STBD1 LIR/GABARAPL1 相互作用和 STBD1 CBM20 的完整两个寡糖结合位点对于 STBD1、GABARAPL1 和糖原在细胞内有效结合都是必不可少的。最后，通过质谱、生化和结构建模分析，我们揭示了 STBD1 可以通过其 LIR 直接与 RB1CC1 的 Claw 结构域结合，从而招募关键的自噬起始因子 RB1CC1。总之，我们的研究结果为 STBD1 识别糖原、ATG8 家族蛋白和 RB1CC1 的机制提供了深入的见解，并揭示了 STBD1 介导的 glycophagy 的潜在工作机制。
