After reading many 2‐DE‐based articles featuring lists of the differentially expressed proteins, one starts experiencing a disturbing déjà vu. The same proteins seem to predominate regardless of the experiment, tissue or species. To quantify the occurrence of individual differentially expressed proteins in 2‐DE experiment reports, we compiled the identities of differentially expressed proteins identified in human, mouse, and rat tissues published in three recent volumes of Proteomics and calculated the appearance of the most predominant proteins in the dataset. The most frequently identified protein is a highly abundant glycolytic enzyme enolase 1, differentially expressed in nearly every third experiment on both human and rodent tissues. Heat‐shock protein 27 (HSP27) and heat‐shock protein 60 (HSP60) were differentially expressed in about 30 percent of human and rodent samples, respectively. Considering protein families as units, keratins and peroxiredoxins are the most frequently identified molecules, with at least one member of the group being differentially expressed in about 40 percent of all experiments. We suggest that the frequent identification of these proteins must be considered in the interpretation of any 2‐DE studies. We consider if these commonly observed changes represent common cellular stress responses or are a reflection of the technical limitations of 2‐DE.

Aims Iron replacement improves clinical status in iron‐deficient patients with heart failure (HF), but the pathophysiology is poorly understood. Iron is essential not only for erythropoiesis, but also for cellular bioenergetics. The impact of myocardial iron deficiency (MID) on mitochondrial function, measured directly in the failing human heart, is unknown. Methods and results Left ventricular samples were obtained from 91 consecutive HF patients undergoing transplantation and 38 HF‐free organ donors (controls). Total myocardial iron content, mitochondrial respiration, citric acid cycle and respiratory chain enzyme activities, respiratory chain components (complex I–V), and protein content of reactive oxygen species (ROS)‐protective enzymes were measured in tissue homogenates to quantify mitochondrial function. Myocardial iron content was lower in HF compared with controls (156 ± 41 vs. 200 ± 38 µg·g⁻¹ dry weight, P < 0.001), independently of anaemia. MID (the lowest iron tercile in HF) was associated with more extensive coronary disease and less beta‐blocker usage compared with non‐MID HF patients. Compared with controls, HF patients displayed reduced myocardial oxygen₂ respiration and reduced activity of all examined mitochondrial enzymes (all P < 0.001). MID in HF was associated with preserved activity of respiratory chain enzymes but reduced activity of aconitase and citrate synthase (by –26% and –15%, P < 0.05) and reduced expression of catalase, glutathione peroxidase, and superoxide dismutase 2. Conclusion Myocardial iron content is decreased and mitochondrial functions are impaired in advanced HF. MID in HF is associated with diminished citric acid cycle enzyme activities and decreased ROS‐protecting enzymes. MID may contribute to altered myocardial substrate use and to worsening of mitochondrial dysfunction that exists in HF.

Diamond‐Blackfan anemia (DBA) is a congenital erythroid aplasia characterized as a normochromic macrocytic anemia with a selective deficiency in red blood cell precursors in otherwise normocellular bone marrow. In 40% of DBA patients, various physical anomalies are also present. Currently two genes are associated with the DBA phenotype—the ribosomal protein (RP) S19 mutated in 25% of DBA patients and RPS24 mutated in ∼1.4% of DBA patients. Here we report the identification of a mutation in yet another ribosomal protein, RPS17. The mutation affects the translation initiation start codon, changing T to G (c.2T>G), thus eliminating the natural start of RPS17 protein biosynthesis. RNA analysis revealed that the mutated allele was expressed, and the next downstream start codon located at position +158 should give rise to a short peptide of only four amino acids (Met‐Ser‐Arg‐Ile). The mutation arose de novo, since all healthy family members carry the wild‐type alleles. The identification of a mutation in the third RP of the small ribosomal subunit in DBA patients further supports the theory that impaired translation may be the main cause of DBA pathogenesis. Hum Mutat 28(12),1178–1182, 2007.

Hepcidin is a major regulator of iron metabolism. Hepcidin-based therapeutics/diagnostics could play roles in hematology in the future, and thus, hepcidin transport is crucial to understand. In this study, we identify α₂-macroglobulin (α₂-M) as the specific hepcidin-binding molecule in blood. Interaction of ¹²⁵I-hepcidin with α₂-M was identified using fractionation of plasma proteins followed by native gradient polyacrylamide gel electrophoresis and mass spectrometry. Hepcidin binding to nonactivated α₂-M displays high affinity (K_d 177 ± 27 nM), whereas hepcidin binding to albumin was nonspecific and displayed nonsaturable kinetics. Surprisingly, the interaction of hepcidin with activated α₂-M exhibited a classical sigmoidal binding curve demonstrating cooperative binding of 4 high-affinity (K_d 0.3 μM) hepcidin-binding sites. This property probably enables efficient sequestration of hepcidin and its subsequent release or inactivation that may be important for its effector functions. Because α₂-M rapidly targets ligands to cells via receptor-mediated endocytosis, the binding of hepcidin to α₂-M may influence its functions. In fact, the α₂-M–hepcidin complex decreased ferroportin expression in J774 cells more effectively than hepcidin alone. The demonstration that α₂-M is the hepcidin transporter could lead to better understanding of hepcidin physiology, methods for its sensitive measurement and the development of novel drugs for the treatment of iron-related diseases.

Diamond‐Blackfan anemia (DBA) is a congenital red blood cell aplasia that is usually diagnosed during early infancy. Apart from defects in red blood cell maturation, the disorder is also associated with various physical anomalies in 40% of patients. Mutations in the ribosomal protein (RP) S19 are found in 25% of patients, while mutations in other proteins of the small ribosomal subunit—RPS17 and RPS24—have been found in a fraction of patients. Recently, mutations in RPL5, RPL11, and RPL35a of the large ribosomal subunit have also been reported in several DBA patients. Here, we present the identification of mutations in the RPL5 and RPL11 genes in patients from the Czech DBA Registry. Mutations in RPL5 were identified in eight patients from 6 out of 28 families (21.4%), and mutations in RPL11 in two patients from 2 out of 28 families (7.1%). Interestingly, all 10 patients with either an RPL5 or RPL11 mutation exhibited one or more physical anomalies; specifically, thumb anomalies (flat thenar) were always present, while no such anomaly was observed in seven patients with an RPS19 mutation. Moreover, 9 out of 10 patients with either an RPL5 or RPL11 mutation were born small for gestational age (SGA) compared to 3 out of 7 patients from the RPS19‐mutated group. These observations may suggest that mutations, at least in RPL5, seem to generally have more profound impact on fetal development than mutations in RPS19. Since RPL5 and RPL11, together with RPL23, are also involved in the MDM2‐mediated p53 pathway regulation, we also screened the RPL23 gene for mutations; however, no mutations were identified. Hum Mutat 0, 1–7, 2009.