Several abnormalities in complement genes reportedly contribute to atypical hemolytic uremic syndrome (aHUS), but incomplete penetrance suggests that additional factors are necessary for the disease to manifest. Here, we sought to describe genotype-phenotype correlations among patients with combined mutations, defined as mutations in more than one complement gene. We screened 795 patients with aHUS and identified single mutations in 41% and combined mutations in 3%. Only 8%-10% of patients with mutations in CFH, C3, or CFB had combined mutations, whereas approximately 25% of patients with mutations in MCP or CFI had combined mutations. The concomitant presence of CFH and MCP risk haplotypes significantly increased disease penetrance in combined mutated carriers, with 73% penetrance among carriers with two risk haplotypes compared with 36% penetrance among carriers with zero or one risk haplotype. Among patients with CFH or CFI mutations, the presence of mutations in other genes did not modify prognosis; in contrast, 50% of patients with combined MCP mutation developed end stage renal failure within 3 years from onset compared with 19% of patients with an isolated MCP mutation. Patients with combined mutations achieved remission with plasma treatment similar to patients with single mutations. Kidney transplant outcomes were worse, however, for patients with combined MCP mutation compared with an isolated MCP mutation. In summary, these data suggest that genotyping for the risk haplotypes in CFH and MCP may help predict the risk of developing aHUS in unaffected carriers of mutations. Furthermore, screening patients with aHUS for all known disease-associated genes may inform decisions about kidney transplantation.

Dense deposit disease (DDD) is a severe renal disease characterized by accumulation of electron-dense material in the mesangium and glomerular basement membrane. Previously, DDD has been associated with deficiency of factor H (fH), a plasma regulator of the alternative pathway (AP) of complement activation, and studies in animal models have linked pathogenesis to the massive complement factor 3 (C3) activation caused by this deficiency. Here, we identified a unique DDD pedigree that associates disease with a mutation in the C3 gene. Mutant C3 923ΔDG , which lacks 2 amino acids, could not be cleaved to C3b by the AP C3-convertase and was therefore the predominant circulating C3 protein in the patients. However, upon activation to C3b by proteases, or to C3(H 2 O) by spontaneous thioester hydrolysis, C3 923ΔDG generated an active AP C3-convertase that was regulated normally by decay accelerating factor (DAF) but was resistant to decay by fH. Moreover, activated C3b 923ΔDG and C3(H 2 O) 923ΔDG were resistant to proteolysis by factor I (fI) in the presence of fH, but were efficiently inactivated in the presence of membrane cofactor protein (MCP). These characteristics cause a fluid phase-restricted AP dysregulation in the patients that continuously activated and consumed C3 produced by the normal C3 allele. These findings expose structural requirements in C3 that are critical for recognition of the substrate C3 by the AP C3-convertase and for the regulatory activities of fH, DAF, and MCP, all of which have implications for therapeutic developments. IntroductionComplement is a major component of innate immunity, with crucial roles in microbial killing, apoptotic cell clearance, immune complex handling, and modulation of adaptive immune responses. Complement is activated by 3 independent activation pathways: the classical pathway (CP), the lectin pathway (LP), and the alternative pathway (AP). The critical steps in complement activation are the formation of unstable protease complexes, named complement factor 3-convertases (C3-convertases; specifically, C3bBb for AP and C4b2a for CP and LP), and the cleavage of C3 by the convertases to generate C3b. Convertase-generated C3b can form more AP C3-convertase, providing exponential amplification to the initial activation. Binding of C3b to the C3-convertases generates the C5-convertases with the capacity to bind and cleave C5, initiating formation of the lytic membrane attack complex (MAC). In contrast to the CP and the LP, whose activation is triggered by immune complexes and bacterial mannose groups, respectively, the AP is intrinsically activated. Spontaneous activation of C3 in plasma occurs through the tick-over mecha-

Atypical hemolytic uremic syndrome (aHUS) and C3 glomerulopathy (C3G) are associated with dysregulation and overactivation of the complement alternative pathway. Typically, gene analysis for aHUS and C3G is undertaken in small patient numbers, yet it is unclear which genes most frequently predispose to aHUS or C3G. Accordingly, we performed a six-center analysis of 610 rare genetic variants in 13 mostly complement genes (, ,, ,, ,, ,, ,, , and) from >3500 patients with aHUS and C3G. We report 371 novel rare variants (RVs) for aHUS and 82 for C3G. Our new interactive Database of Complement Gene Variants was used to extract allele frequency data for these 13 genes using the Exome Aggregation Consortium server as the reference genome. For aHUS, significantly more protein-altering rare variation was found in five genes ,, ,, and than in the Exome Aggregation Consortium (allele frequency< 0.01%), thus correlating these with aHUS. For C3G, an association was only found for RVs in and the N-terminal C3b-binding or C-terminal nonsurface-associated regions of In conclusion, the RV analyses showed nonrandom distributions over the affected proteins, and different distributions were observed between aHUS and C3G that clarify their phenotypes.

IgA nephropathy (IgAN), a frequent cause of chronic kidney disease worldwide, is characterized by mesangial deposition of galactose-deficient IgA1-containing immune complexes. Complement involvement in IgAN pathogenesis is suggested by the glomerular deposition of complement components and the strong protection from IgAN development conferred by the deletion of the CFHR3 and CFHR1 genes (Δ). Here we searched for correlations between clinical progression and levels of factor H (FH) and FH-related protein 1 (FHR-1) using well-characterized patient cohorts consisting of 112 patients with IgAN, 46 with non-complement-related autosomal dominant polycystic kidney disease (ADPKD), and 76 control individuals. Patients with either IgAN or ADPKD presented normal FH but abnormally elevated FHR-1 levels and FHR-1/FH ratios compared to control individuals. Highest FHR-1 levels and FHR-1/FH ratios are found in patients with IgAN with disease progression and in patients with ADPKD who have reached chronic kidney disease, suggesting that renal function impairment elevates the FHR-1/FH ratio, which may increase FHR-1/FH competition for activated C3 fragments. Interestingly, Δ homozygotes are protected from IgAN, but not from ADPKD, and we found five IgAN patients with low FH carrying CFH or CFI pathogenic variants. These data support a decreased FH activity in IgAN due to increased FHR-1/FH competition or pathogenic CFH variants. They also suggest that alternative pathway complement activation in patients with IgAN, initially triggered by galactose-deficient IgA1-containing immune complexes, may exacerbate in a vicious circle as renal function deterioration increase FHR-1 levels. Thus, a role of FHR-1 in IgAN pathogenesis is to compete with complement regulation by FH.

Background and objectives Atypical hemolytic uremic syndrome is characterized by vascular endothelial damage caused by complement dysregulation. Consistently, complement inhibition therapies are highly effective in most patients with atypical hemolytic uremic syndrome. Recently, it was shown that a significant percentage of patients with early-onset atypical hemolytic uremic syndrome carry mutations in diacylglycerol kinase-«, an intracellular protein with no obvious role in complement. These data support an alternative, complement-independent mechanism leading to thrombotic microangiopathy that has implications for treatment of early-onset atypical hemolytic uremic syndrome. To get additional insights into this new form of atypical hemolytic uremic syndrome, the diacylglycerol kinase-« gene in a cohort with atypical hemolytic uremic syndrome was analyzed.

Dois trechos de Floresta Estacional Semidecodual, em distintos estádios sucessionais (inicial e maduro), da região de Viçosa, MG, foram estudados com o objetivo de quantificar a produção anual e o conteúdo de N, P, K, Ca e Mg da serapilheira, bem como caracterizar a dinâmica de decomposição e liberação dos nutrientes desse compartimento e a eficiência anual de utilização dos nutrientes. A queda anual de serapilheira foi de 6.310 kg.ha-1 no trecho de floresta em seu estádio inicial e de 8.819 kg.ha-1 no de floresta madura. O conteúdo de nutrientes foi de 137 e 180 kg.ha-1 de N, 5 e 8 kg.ha-1 de P; 17 e 45 kg.ha-1 de K; 89 e 179 kg.ha-1 de Ca; e 21 e 26 kg.ha-1 de Mg, nos trechos de floresta nos estádios inicial e maduro, respectivamente. A quantidade média de serapilheira acumulada sobre o solo totalizou 4.647 kg.ha-1 no trecho de floresta inicial e 7.006 kg.ha-1 na floresta madura. A estimativa média da taxa instantânea de decomposição (K) foi de 1,36 no trecho de floresta inicial e de 1,26 na floresta madura, sendo o tempo médio de renovação da serapilheira de 270 e 288 dias, respectivamente. A menor produção de serapilheira na floresta inicial reflete, em parte, a estrutura menos desenvolvida desse trecho de floresta em estádio inicial de sucessão, com produção de serapilheira de qualidade inferior à da floresta madura, no entanto com renovação mais rápida e utilização mais eficiente dos nutrientes.

HighlightsMutations in C3 have been associated with aHUS and other glomerulopathies.aHUS-associated C3 mutants R592W, R161W, and I1157T impair regulation by MCP, but not by FH.EM analysis provides the structural basis for the functional impairment of the R161W and I1157T mutants.Data supports aHUS-associated C3 mutations selectively affect complement regulation on surfaces.

Com o objetivo de se avaliar as exigências nutricionais e o efeito da omissão de nutrientes no crescimento de mudas de cedro australiano (Toona ciliata M. Roem var. australis), foi realizado um experimento em casa de vegetação. Utilizou-se como substrato um Latossolo Vermelho Distroférrico, com baixa disponibilidade de nutrientes, utilizando-se 15 tratamentos, sob a técnica do elemento faltante. Foram aplicados dois tratamentos completos (um com fornecimento de N, P, K, S, B, Cu, Zn e aplicação de calcário e outro com o fornecimento de N, P, K, Ca, Mg, S, B, Cu e Zn, sem a aplicação de calcário), além da omissão de cada nutriente (-N, -P, -K, -Ca, -Mg, -S, -B, -Cu e -Zn), um tratamento com omissão conjunta de B, Cu e Zn, um tratamento com aplicação apenas de calcário, um tratamento com aplicação de N, P, K, S, B, Cu e Zn, sem aplicação de calcário e uma testemunha absoluta (solo natural). Foram avaliadas as seguintes características: altura, diâmetro, matéria seca da parte aérea e das raízes, e teor de nutrientes na matéria seca da parte aérea, ao final de 150 dias. As plantas de cedro australiano apresentam elevada exigência nutricional, sendo que os nutrientes P, N, S, Ca, K, Mg e Cu, nessa ordem, foram limitantes ao crescimento das plantas. A omissão de B e Zn não afetou o crescimento das plantas. A aplicação de calcário foi essencial para o desenvolvimento das plantas de cedro australiano. O aparecimento dos primeiros sintomas de deficiência foi decorrente da omissão de: S, calcário e N.