The WDR11 complex is a receptor for acidic-cluster-containing cargo proteins.

Vesicle trafficking is a fundamental process that allows for the sorting and transport of specific proteins (i.e., "cargoes") to different compartments of eukaryotic cells. Cargo recognition primarily occurs through coats and the associated proteins at the donor membrane. However, it remains unclear whether cargoes can also be selected at other stages of vesicle trafficking to further enhance the fidelity of the process. The WDR11-FAM91A1 complex functions downstream of the clathrin-associated AP-1 complex to facilitate protein transport from endosomes to the TGN. Here, we report the cryo-EM structure of human WDR11-FAM91A1 complex. WDR11 directly and specifically recognizes a subset of acidic clusters, which we term super acidic clusters (SACs). WDR11 complex assembly and its binding to SAC-containing proteins are indispensable for the trafficking of SAC-containing proteins and proper neuronal development in zebrafish. Our studies thus uncover that cargo proteins could be recognized in a sequence-specific manner downstream of a protein coat.

FAM91A1-TBC1D23 complex structure reveals human genetic variations susceptible for PCH.

Pontocerebellar hypoplasia (PCH) is a group of rare neurodevelopmental disorders with limited diagnostic and therapeutic options. Mutations in WDR11, a subunit of the FAM91A1 complex, have been found in patients with PCH-like symptoms; however, definitive evidence that the mutations are causal is still lacking. Here, we show that depletion of FAM91A1 results in developmental defects in zebrafish similar to that of TBC1D23, an established PCH gene. FAM91A1 and TBC1D23 directly interact with each other and cooperate to regulate endosome-to-Golgi trafficking of KIAA0319L, a protein known to regulate axonal growth. Crystal structure of the FAM91A1-TBC1D23 complex reveals that TBC1D23 binds to a conserved surface on FAM91A1 by assuming a Z-shaped conformation. More importantly, the interaction between FAM91A1 and TBC1D23 can be used to predict the risk of certain TBC1D23-associated mutations to PCH. Collectively, our study provides a molecular basis for the interaction between TBC1D23 and FAM91A1 and suggests that disrupted endosomal trafficking underlies multiple PCH subtypes.

Senescent renal tubular epithelial cells activate fibroblasts by secreting Shh to promote the progression of diabetic kidney disease.

Introduction: Diabetic kidney disease (DKD) is one of the complications of diabetes; however, the pathogenesis is not yet clear. A recent study has shown that senescence is associated with the course of DKD. In the present study, we explored whether senescent renal tubular cells promote renal tubulointerstitial fibrosis by secreting Sonic hedgehog (Shh) which mediates fibroblast activation and proliferation in DKD. Methods: A 36-week-old db/db mice model and the renal tubular epithelial cells were cultured in high glucose (HG, 60 mmol/L) medium for in vivo and in vitro experiments. Results: Compared to db/m mice, blood glucose, microalbuminuria, serum creatinine, urea nitrogen, and UACR (microalbuminuria/urine creatinine) were markedly increased in db/db mice. Collagen III, monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) were also increased in db/db mice kidneys, suggesting fibrosis and inflammation in the organ. Moreover, the detection of SA-ß-galactosidase (SA-ß-Gal) showed that the activity of SA-ß-Gal in the cytoplasm of renal tubular epithelial cells increased, and the cell cycle inhibition of the expression of senescence-related gene cell cycle inhibitor p16 INK4A protein and p21 protein increased, indicating that renal fibrosis in db/db mice was accompanied by cell senescence. Furthermore, Shh is highly expressed in the injured renal tubules and in the kidney tissue of db/db mice, as detected by enzyme-linked immunosorbent assay (ELISA). The results of immunofluorescence staining showed increased positive staining for Shh in renal tubular epithelial cells of db/db mice and decreased positive staining for Lamin B1, but increased positive staining for γH2A.X in cells with high Shh expression; similar results were obtained in vitro. In addition, HG stimulated renal tubular epithelial cells to secrete Shh in the supernatant of the medium. D-gal treatment of renal tubular epithelial cells increased the protein levels of Shh and p21. We also found enhanced activation and proliferation of fibroblasts cultured with the supernatant of renal tubular epithelial cells stimulated by HG medium but the proliferative effect was significantly diminished when co-cultured with cyclopamine (CPN), an inhibitor of the Shh pathway. Discussion: In conclusion, HG induces renal tubular epithelial cell senescence, and the secretion of senescence-associated proteins and Shh mediates inflammatory responses and fibroblast activation and proliferation, ultimately leading to renal fibrosis.

Phosphorylation of SNX27 by MAPK11/14 links cellular stress-signaling pathways with endocytic recycling.

Endocytosed proteins can be delivered to lysosomes for degradation or recycled to either the trans-Golgi network or the plasma membrane. It remains poorly understood how the recycling versus degradation of cargoes is determined. Here, we show that multiple extracellular stimuli, including starvation, LPS, IL-6, and EGF treatment, can strongly inhibit endocytic recycling of multiple cargoes through the activation of MAPK11/14. The stress-induced kinases in turn directly phosphorylate SNX27, a key regulator of endocytic recycling, at serine 51 (Ser51). Phosphorylation of SNX27 at Ser51 alters the conformation of its cargo-binding pocket and decreases the interaction between SNX27 and cargo proteins, thereby inhibiting endocytic recycling. Our study indicates that endocytic recycling is highly dynamic and can crosstalk with cellular stress-signaling pathways. Suppression of endocytic recycling and enhancement of receptor lysosomal degradation serve as new mechanisms for cells to cope with stress and save energy.