糖原自噬(glycophagy)对于维持哺乳动物细胞葡萄糖稳态和生理至关重要。STBD1可以作为自噬受体,通过特异性识别糖原和相关的关键自噬因子来介导糖吞噬,但对机制知之甚少。这里,我们系统地表征了STBD1与糖原和相关糖的相互作用,并用麦芽四糖确定STBD1CBM20结构域的晶体结构,揭示了一种独特的结合模式,涉及STBD1CBM20用于识别糖原的两个不同的寡糖结合位点。此外,我们证明了STBD1的LC3相互作用区(LIR)基序可以选择性地与六个哺乳动物ATG8家族成员结合。通过解决STBD1LIR/GABARAPL1复合物结构,我们阐明了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,我们的基于细胞的分析显示,STBD1LIR/GABARAPL1相互作用和STBD1CBM20完整的两个寡糖结合位点对于细胞中STBD1,GABARAPL1和糖原的有效结合是必不可少的.最后,通过质谱,生物化学,和结构建模分析,我们揭示了STBD1可以通过其LIR直接绑定到RB1CC1的Claw域,从而招募关键的自噬启动因子RB1CC1。总之,我们的发现提供了对糖原识别的机械见解,ATG8家族蛋白,STBD1和RB1CC1,揭示了STBD1介导的糖吞噬的潜在工作机制。
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.