African-Americans have an increased risk of developing chronic and end-stage kidney disease, with much of it attributed to two common genetic variants in the APOL1 gene, termed G1 and G2. Direct evidence demonstrating that these APOL1 risk alleles are pathogenic is still lacking as the APOL1 gene is only present in some primates and humans; thus experimental proof of causality of these risk alleles for renal disease has been challenging. Here, we generated mice with podocyte-specific inducible expression of the APOL1 reference allele (termed G0) or each of the risk alleles (G1 or G2). We show that mice with podocyte-specific expression of either APOL1 risk allele, but not the G0 allele, develop functional (albuminuria, azotemia), structural (foot process effacement and glomerulosclerosis) and molecular (gene expression) changes that closely resemble the human kidney disease. Disease development was cell-type specific, and likely reversible, and the severity correlated with the level of expression of the risk allele. We further found that expression of the APOL1 risk alleles interferes with endosomal trafficking and blocks autophagic flux, leading ultimately to inflammatory-mediated podocyte death and glomerular scarring. In summary, this is the first in vivo demonstration that expression of APOL1 risk alleles are causal for altered podocyte function and glomerular disease.

CD40 is a TNF receptor superfamily member that provides activation signals in antigen-presenting cells such as B cells, macrophages, and dendritic cells. Multimerization of CD40 by its ligand initiates signaling by recruiting TNF receptor-associated factors (TRAFs) to the CD40 cytoplasmic domain. Recombinant human TRAF proteins overexpressed in insect cells were biochemically characterized and used to finely map TRAF binding regions in the human CD40 cytoplasmic domain. TRAF1, TRAF2, TRAF3, and TRAF6, but not TRAF4 or TRAF5, bound directly to the CD40 cytoplasmic domain. CD40 interactions with TRAF2 and TRAF3 were stronger than the interactions with TRAF1 and TRAF6. Full-length TRAF3 and TRAF5 formed hetero-oligomers, presumably through their predicted isoleucine zippers. TRAF3-TRAF5 hetero-oligomers interacted with CD40, indicating that TRAF5 can be indirectly recruited to the CD40 cytoplasmic domain. Overlapping peptides synthesized on cellulose membranes were used to map each TRAF interaction region. TRAF1, TRAF2, and TRAF3 interacted with the same region. The recognition site for TRAF6 was a nonoverlapping membrane proximal region. Using peptides with progressive deletions, a minimal TRAF1, TRAF2, and TRAF3 binding region was mapped to the PVQET sequence in the CD40 cytoplasmic domain. The minimal region for TRAF6 binding was the sequence QEPQEINF. These studies demonstrate that the CD40 cytoplasmic domain contains two nonoverlapping TRAF binding regions and suggest that TRAF1, TRAF2, and TRAF3 could bind competitively to one site. Relative affinities and competition of individual and hetero-oligomeric TRAF proteins for CD40 binding sites may contribute to receptor specificity and cell-type selectivity in CD40-dependent signaling.

Chronic kidney disease (CKD), a condition when the kidneys are unable to clear waste products, affects 700 million people globally. Genome-wide association (GWA) studies identified sequence variants for CKD; however, the biological basis of GWAS remains poorly understood. To address this issue, we created an expression quantitative trait loci (eQTL) atlas for the glomerular and tubular compartments of the human kidney. Integrating the CKD GWAS with eQTL, single-cell RNA sequencing and regulatory region maps, we identified novel genes for CKD. Putative causal genes were enriched for proximal tubule expression and endo-lysosomal function, where DAB2, an adaptor protein in the TGFβ pathway, formed a central node. Functional experiments confirmed that reducing Dab2 expression in renal tubules protected mice from CKD. In conclusion, compartment-specific eQTL analysis is an important avenue for the identification of novel genes and cellular pathways involved in CKD development and thus potential new opportunities for its treatment.

Tumor necrosis factor receptor superfamily members convey signals that promote diverse cellular responses. Receptor trimerization by extracellular ligands initiates signaling by recruiting members of the tumor necrosis factor receptor-associated factor (TRAF) family of adapter proteins to the receptor cytoplasmic domains. We report the 2.4-Å crystal structure of a 22-kDa, receptor-binding fragment of TRAF2 complexed with a functionally defined peptide from the cytoplasmic domain of the CD40 receptor. TRAF2 forms a mushroom-shaped trimer consisting of a coiled coil and a unique ␤-sandwich domain. Both domains mediate trimerization. The CD40 peptide binds in an extended conformation with every side chain in contact with a complementary groove on the rim of each TRAF monomer. The spacing between the CD40 binding sites on TRAF2 supports an elegant signaling mechanism in which trimeric, extracellular ligands preorganize the receptors to simultaneously recognize three sites on the TRAF trimer.