Naturally occurring genetic variants of human caspase-1 differ considerably in structure and the ability to activate interleukin-1ß.

Caspase-1 (Interleukin-1 Converting Enzyme, ICE) is a proinflammatory enzyme that plays pivotal roles in innate immunity and many inflammatory conditions such as periodic fever syndromes and gout. Inflammation is often mediated by enzymatic activation of interleukin (IL)-1ß and IL-18. We detected seven naturally occurring human CASP1 variants with different effects on protein structure, expression, and enzymatic activity. Most mutations destabilized the caspase-1 dimer interface as revealed by crystal structure analysis and homology modeling followed by molecular dynamics simulations. All variants demonstrated decreased or absent enzymatic and IL-1ß releasing activity in vitro, in a cell transfection model, and as low as 25% of normal ex vivo in a whole blood assay of samples taken from subjects with variant CASP1, a subset of whom suffered from unclassified autoinflammation. We conclude that decreased enzymatic activity of caspase-1 is compatible with normal life and does not prevent moderate and severe autoinflammation.

Soluble BAFF levels inversely correlate with peripheral B cell numbers and the expression of BAFF receptors.

The TNF family member protein BAFF/BLyS is essential for B cell survival and plays an important role in regulating class switch recombination as well as in the selection of autoreactive B cells. In humans, increased concentrations of soluble BAFF are found in different pathological conditions, which may be as diverse as autoimmune diseases, B cell malignancies, and primary Ab deficiencies (PAD). Because the mechanisms that regulate BAFF levels are not well understood, we newly developed a set of mAbs against human BAFF to study the parameters that determine the concentrations of soluble BAFF in circulation. Patients with PAD, including severe functional B cell defects such as BTK, BAFF-R, or TACI deficiency, were found to have higher BAFF levels than asplenic individuals, patients after anti-CD20 B cell depletion, chronic lymphocytic leukemia patients, or healthy donors. In a comparable manner, mice constitutively expressing human BAFF were found to have higher concentrations of BAFF in the absence than in the presence of B cells. Therefore, our data strongly suggest that BAFF steady-state concentrations mainly depend on the number of B cells as well as on the expression of BAFF-binding receptors. Because most patients with PAD have high levels of circulating BAFF, the increase in BAFF concentrations cannot compensate defects in B cell development and function.

Relevance of biallelic versus monoallelic TNFRSF13B mutations in distinguishing disease-causing from risk-increasing TNFRSF13B variants in antibody deficiency syndromes.

TNFRSF13B encodes transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI), a B cell- specific tumor necrosis factor (TNF) receptor superfamily member. Both biallelic and monoallelic TNFRSF13B mutations were identified in patients with common variable immunodeficiency disorders. The genetic complexity and variable clinical presentation of TACI deficiency prompted us to evaluate the genetic, immunologic, and clinical condition in 50 individuals with TNFRSF13B alterations, following screening of 564 unrelated patients with hypogammaglobulinemia. We identified 13 new sequence variants. The most frequent TNFRSF13B variants (C104R and A181E; n=39; 6.9%) were also present in a heterozygous state in 2% of 675 controls. All patients with biallelic mutations had hypogammaglobulinemia and nearly all showed impaired binding to a proliferation-inducing ligand (APRIL). However, the majority (n=41; 82%) of the pa-tients carried monoallelic changes in TNFRSF13B. Presence of a heterozygous mutation was associated with antibody deficiency (P< .001, relative risk 3.6). Heterozygosity for the most common mutation, C104R, was associated with disease (P< .001, relative risk 4.2). Furthermore, heterozygosity for C104R was associated with low numbers of IgD(-)CD27(+) B cells (P= .019), benign lymphoproliferation (P< .001), and autoimmune complications (P= .001). These associations indicate that C104R heterozygosity increases the risk for common variable immunodeficiency disorders and influences clinical presentation.

Screening of functional and positional candidate genes in families with common variable immunodeficiency.

BACKGROUND: Common variable immunodeficiency (CVID) comprises a heterogeneous group of primary antibody deficiencies with complex clinical and immunological phenotypes. The recent discovery that some CVID patients show monogenic defects in the genes encoding ICOS, TACI or CD19 prompted us to investigate several functional candidate genes in individuals with CVID. RESULTS: The exonic, protein coding regions of the genes encoding: APRIL, BCMA, IL10, IL10Ralpha, IL10Rbeta, IL21, IL21R, and CCL18, were analyzed primarily in familial CVID cases, who showed evidence of genetic linkage to the respective candidate gene loci and CVID families with a recessive pattern of inheritance. Two novel SNPs were identified in exon 5 and exon 8 of the IL21R gene, which segregated with the disease phenotype in one CVID family. Eleven additional SNPs in the genes encoding BCMA, APRIL, IL10, IL10Ralpha, IL21 and IL21R were observed at similar frequencies as in healthy donors. CONCLUSION: We were unable to identify obvious disease causing mutations in the protein coding regions of the analyzed genes in the studied cohort.

Sequence analysis of TNFRSF13b, encoding TACI, in patients with systemic lupus erythematosus.

B cell activating factor belonging to the TNF family (BAFF) and a proliferation inducing ligand (APRIL), and their receptors BAFF receptor (BAFFR), B cell maturation antigen (BCMA), and transmembrane activator and CAML interactor (TACI) are involved in the regulation of B cell homeostasis and differentiation. BAFF overexpression leads to systemic lupus erythematosus (SLE) in mice and elevated BAFF levels have been observed in human SLE and mouse models for SLE. Furthermore, genetic inactivation of TACI in mice results in a SLE-like phenotype. Based on our recent finding that TACI is mutated in patients with common variable immunodeficiency, of whom more than 30% suffer from autoimmune conditions, we analyzed TACI in humans with SLE. Sequence analysis of TNFRSF13b/TACI in 119 unrelated SLE patients revealed four variants: R20C in exon 1, R72H in exon 3, the silent variation c.327 G > A in exon 3, and A181E in exon 4. No significant association with any of these variants was found, when compared to the frequencies of the variants in a healthy control cohort. Furthermore, the mutated alleles R20C and R72H did not segregate with the SLE phenotype in familial cases of SLE. Thus, our evaluation of the coding region of TNFRSF13b/TACI did not reveal any deleterious or disease-associated mutations.