Digenic inheritance of human primary microcephaly delineates centrosomal and non-centrosomal pathways.

Primary microcephaly (PM) is characterized by a small head since birth and is vastly heterogeneous both genetically and phenotypically. While most cases are monogenic, genetic interactions between Aspm and Wdr62 have recently been described in a mouse model of PM. Here, we used two complementary, holistic in vivo approaches: high throughput DNA sequencing of multiple PM genes in human patients with PM, and genome-edited zebrafish modeling for the digenic inheritance of PM. Exomes of patients with PM showed a significant burden of variants in 75 PM genes, that persisted after removing monogenic causes of PM (e.g., biallelic pathogenic variants in CEP152). This observation was replicated in an independent cohort of patients with PM, where a PM gene panel showed in addition that the burden was carried by six centrosomal genes. Allelic frequencies were consistent with digenic inheritance. In zebrafish, non-centrosomal gene casc5 -/- produced a severe PM phenotype, that was not modified by centrosomal genes aspm or wdr62 invalidation. A digenic, quadriallelic PM phenotype was produced by aspm and wdr62. Our observations provide strong evidence for digenic inheritance of human PM, involving centrosomal genes. Absence of genetic interaction between casc5 and aspm or wdr62 further delineates centrosomal and non-centrosomal pathways in PM

Molecular basis of ClC-6 function and its impairment in human disease

ClC-6 is a late endosomal voltage-gated chloride-proton exchanger that is predominantly expressed in the nervous system. Mutated forms of ClC-6 are associated with severe neurological disease. However, the mechanistic role of ClC-6 in normal and pathological states remains largely unknown. Here, we present cryo-EM structures of ClC-6 that guided subsequent functional studies. Previously unrecognized ATP binding to cytosolic ClC-6 domains enhanced ion transport activity. Guided by a disease-causing mutation (p.Y553C), we identified an interaction network formed by Y553/F317/T520 as potential hotspot for disease-causing mutations. This was validated by the identification of a patient with a de novo pathogenic variant p.T520A. Extending these findings, we found contacts between intramembrane helices and connecting loops that modulate the voltage dependence of ClC-6 gating and constitute additional candidate regions for disease-associated gain-of-function mutations. Besides providing insights into the structure, function, and regulation of ClC-6, our work correctly predicts hotspots for CLCN6 mutations in neurodegenerative disorders

An exome-wide study of renal operational tolerance

BackgroundRenal operational tolerance is a rare and beneficial state of prolonged renal allograft function in the absence of immunosuppression. The underlying mechanisms are unknown. We hypothesized that tolerance might be driven by inherited protein coding genetic variants with large effect, at least in some patients.MethodsWe set up a European survey of over 218,000 renal transplant recipients and collected DNAs from 40 transplant recipients who maintained good allograft function without immunosuppression for at least 1 year. We performed an exome-wide association study comparing the distribution of moderate to high impact variants in 36 tolerant patients, selected for genetic homogeneity using principal component analysis, and 192 controls, using an optimal sequence-kernel association test adjusted for small samples.ResultsWe identified rare variants of HOMER2 (3/36, FDR 0.0387), IQCH (5/36, FDR 0.0362), and LCN2 (3/36, FDR 0.102) in 10 tolerant patients vs. 0 controls. One patient carried a variant in both HOMER2 and LCN2. Furthermore, the three genes showed an identical variant in two patients each. The three genes are expressed at the primary cilium, a key structure in immune responses.ConclusionRare protein coding variants are associated with operational tolerance in a sizable portion of patients. Our findings have important implications for a better understanding of immune tolerance in transplantation and other fields of medicine.ClinicalTrials.gov, identifier: NCT05124444