State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China.
Bio-X Institutes, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.
J Am Soc Nephrol. 2021 Aug;32(8):1946-1960. doi: 10.1681/ASN.2020111590.
Slit diaphragm is a specialized adhesion junction between the opposing podocytes, establishing the final filtration barrier to urinary protein loss. At the cytoplasmic insertion site of each slit diaphragm there is an electron-dense and protein-rich cellular compartment that is essential for slit diaphragm integrity and signal transduction. Mutations in genes that encode components of this membrane-less compartment have been associated with glomerular diseases. However, the molecular mechanism governing formation of compartmentalized slit diaphragm assembly remains elusive.
We systematically investigated the interactions between key components at slit diaphragm, such as MAGI2, Dendrin, and CD2AP, through a combination of biochemical, biophysical, and cell biologic approaches.
We demonstrated that MAGI2, a unique MAGUK family scaffold protein at slit diaphragm, can autonomously undergo liquid-liquid phase separation. Multivalent interactions among the MAGI2-Dendrin-CD2AP complex drive the formation of the highly dense slit diaphragm condensates at physiologic conditions. The reconstituted slit diaphragm condensates can effectively recruit Nephrin. A nephrotic syndrome-associated mutation of interfered with formation of the slit diaphragm condensates, thus leading to impaired enrichment of Nephrin.
Key components at slit diaphragm (, MAGI2 and its complex) can spontaneously undergo phase separation. The reconstituted slit diaphragm condensates can be enriched in adhesion molecules and cytoskeletal adaptor proteins. Therefore, the electron-dense slit diaphragm assembly might form via phase separation of core components of the slit diaphragm in podocytes.
裂孔隔膜是相邻足细胞之间的一种特殊黏附连接，建立了阻止尿液蛋白丢失的最终滤过屏障。在每个裂孔隔膜的细胞质插入部位，有一个电子致密且富含蛋白质的细胞区室，对于裂孔隔膜的完整性和信号转导至关重要。编码该无膜区室成分的基因突变与肾小球疾病有关。然而，控制分隔裂孔隔膜组装的分子机制仍然难以捉摸。
我们通过生化、生物物理和细胞生物学方法的组合，系统地研究了裂孔隔膜关键成分（如 MAGI2、Dendrin 和 CD2AP）之间的相互作用。
我们证明了 MAGI2，一种位于裂孔隔膜的独特 MAGUK 家族支架蛋白，能够自主进行液-液相分离。MAGI2-Dendrin-CD2AP 复合物之间的多价相互作用驱动了在生理条件下高度密集的裂孔隔膜凝聚的形成。重建的裂孔隔膜凝聚物可以有效地招募 Nephrin。一种肾病综合征相关的突变 干扰了裂孔隔膜凝聚的形成，从而导致 Nephrin 的富集受损。
裂孔隔膜（MAGI2 及其复合物）的关键成分可以自发进行相分离。重建的裂孔隔膜凝聚物可以富含黏附分子和细胞骨架衔接蛋白。因此，电子致密的裂孔隔膜组装可能通过足细胞中裂孔隔膜核心成分的相分离形成。
