Autoimmune responses against posttranslationally modified antigens are a hallmark of several autoimmune diseases. For example, antibodies against citrullinated protein antigens (ACPA) have shown their relevance for the prognosis and diagnosis of rheumatoid arthritis (RA), and have been implicated in disease pathogenesis. It is conceivable that other autoantibody systems, recognizing other posttranslationally modified proteins, are also present in RA. Here, we describe the presence of an autoantibody system that discriminates between citrulline- and homocitrulline-containing antigens in the sera of RA-patients. IgG antibodies recognizing carbamylated (homocitrulline-containing) antigens were present in sera of over 45% of RA-patients. Likewise, anticarbamylated protein (anti-CarP) IgA antibodies were observed in 43% of RA-sera. ACPA and anti-CarP antibodies are distinct autoantibodies because, in selected double-positive patients, the anti-CarP antibody binding to carbamylated antigens could be inhibited by carbamylated antigens, but not by control or citrullinated antigens. Similarly, ACPA-binding to citrullinated antigens could only be inhibited by citrullinated antigens. In line with this observation, 16% of ACPA-negative RA-patients, as measured by a standard ACPA assay, harbored IgG anti-CarP antibodies, whereas 30% of these patients tested positive for IgA anti-CarP antibodies. The presence of anti-CarP antibodies was predictive for a more severe disease course in ACPA-negative patients as measured by radiological progression. Taken together, these data show the presence of a unique autoantibody system recognizing carbamylated, but not citrullinated, protein antigens. These antibodies are predictive for a more severe clinical course in ACPA-negative RA-patients, indicating that anti-CarP antibodies are a unique and relevant serological marker for ACPA-negative RA.

RNA viruses encode an RNA-dependent RNA polymerase (RdRp) that catalyzes the synthesis of their RNA(s). In the case of positive-stranded RNA viruses belonging to the order Nidovirales, the RdRp resides in a replicase subunit that is unusually large. Bioinformatics analysis of this non-structural protein has now revealed a nidoviral signature domain (genetic marker) that is N-terminally adjacent to the RdRp and has no apparent homologs elsewhere. Based on its conservation profile, this domain is proposed to have nucleotidylation activity. We used recombinant non-structural protein 9 of the arterivirus equine arteritis virus (EAV) and different biochemical assays, including irreversible labeling with a GTP analog followed by a proteomics analysis, to demonstrate the manganese-dependent covalent binding of guanosine and uridine phosphates to a lysine/histidine residue. Most likely this was the invariant lysine of the newly identified domain, named nidovirus RdRp-associated nucleotidyltransferase (NiRAN), whose substitution with alanine severely diminished the described binding. Furthermore, this mutation crippled EAV and prevented the replication of severe acute respiratory syndrome coronavirus (SARS-CoV) in cell culture, indicating that NiRAN is essential for nidoviruses. Potential functions supported by NiRAN may include nucleic acid ligation, mRNA capping and protein-primed RNA synthesis, possibilities that remain to be explored in future studies.

SummaryThrough a network of progressively maturing vesicles, the endosomal system connects the cell’s interior with extracellular space. Intriguingly, this network exhibits a bilateral architecture, comprised of a relatively immobile perinuclear vesicle “cloud” and a highly dynamic peripheral contingent. How this spatiotemporal organization is achieved and what function(s) it curates is unclear. Here, we reveal the endoplasmic reticulum (ER)-located ubiquitin ligase Ring finger protein 26 (RNF26) as the global architect of the entire endosomal system, including the trans-Golgi network (TGN). To specify perinuclear vesicle coordinates, catalytically competent RNF26 recruits and ubiquitinates the scaffold p62/sequestosome 1 (p62/SQSTM1), in turn attracting ubiquitin-binding domains (UBDs) of various vesicle adaptors. Consequently, RNF26 restrains fast transport of diverse vesicles through a common molecular mechanism operating at the ER membrane, until the deubiquitinating enzyme USP15 opposes RNF26 activity to allow vesicle release into the cell’s periphery. By drawing the endosomal system’s architecture, RNF26 orchestrates endosomal maturation and trafficking of cargoes, including signaling receptors, in space and time.

Mutations in mitochondrial DNA (mtDNA) associate with various disease states. A few mtDNA mutations strongly associate with diabetes, with the most common mutation being the A3243G mutation in the mitochondrial DNA-encoded tRNA(Leu,UUR) gene. This article describes clinical characteristics of mitochondrial diabetes and its molecular diagnosis. Furthermore, it outlines recent developments in the pathophysiological and molecular mechanisms leading to a diabetic state. A gradual development of pancreatic ␤-cell dysfunction upon aging, rather than insulin resistance, is the main mechanism in developing glucose intolerance. Carriers of the A3243G mutation show during a hyperglycemic clamp at 10 mmol/l glucose a marked reduction in firstand second-phase insulin secretion compared with noncarriers. The molecular mechanism by which the A3243G mutation affects insulin secretion may involve an attenuation of cytosolic ADP/ATP levels leading to a resetting of the glucose sensor in the pancreatic ␤-cell, such as in maturity-onset diabetes of the young (MODY)-2 patients with mutations in glucokinase. Unlike in MODY2, which is a nonprogressive form of diabetes, mitochondrial diabetes does show a pronounced age-dependent deterioration of pancreatic function indicating involvement of additional processes. Furthermore, one would expect that all mtDNA mutations that affect ATP synthesis lead to diabetes. This is in contrast to clinical observations. The origin of the age-dependent deterioration of pancreatic function in carriers of the A3243G mutation and the contribution of ATP and other mitochondrion-derived factors such as reactive oxygen species to the development of diabetes is discussed. Diabetes 53 (Suppl. 1):S103-S109, 2004 D iabetes is a collection of diseases characterized by the presence of chronic hyperglycemia. Maintenance of normal glucose homeostasis involves the action of a glucose sensor in the pancreatic ␤-cell that detects an increase in blood glucose concentration and converts that into increased secretion of insulin. Increased circulating insulin concentrations suppress hepatic glucose output and stimulate glucose uptake by muscle and adipose tissue.Pathophysiological mechanisms leading to diabetes can involve an inappropriate secretion of insulin, insulin resistance of the liver, muscle and fat, or combined defects. The risk of an individual to develop diabetes involves a complex interaction between genetic and environmental factors. Gene variants that have been identified to contribute to the major forms of diabetes, such as autoimmune type 1 diabetes and metabolic syndrome-associated type 2 diabetes, are "low penetrance" variants that modulate the susceptibility of an individual to develop diabetes or that protect against the disease (1-3).A number of gene mutants have been identified in the past decade that represent high penetrance risk genes for diabetes. Carriers of these gene mutants have a nearly 100% chance to develop diabetes during their life span. These so-called monogenetic forms of diabetes com...

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