BBA - Molecular Basis of Disease (v.1792, #2)

A generally accepted view posits that insulin resistant condition in type 2 diabetes is caused by defects at one or several levels of the insulin-signaling cascade in skeletal muscles, adipose tissue and liver, that quantitatively constitute the bulk of the insulin-responsive tissues. Hence, the gradual uncovering of the biochemical events defining the intracellular signaling of insulin has been quickly followed by clinical studies on humans attempting to define the molecular defect(s) responsible for the establishment of the insulin resistant state. While the existence of molecular defects within the insulin signal transduction machinery is undisputed, contrasting data exist on what is the principal molecular alteration leading to insulin resistance. Such discrepancies in the literature may depend on: 1) different subject characteristics, 2) methodological differences 3) small cohorts of subjects, and – not least – 4) intrinsic limitations in studying every detail of the insulin signaling cascade. Here, we review the studies on humans exploring the defects of the insulin signaling cascade generated by insulin resistance and type 2 diabetes, focusing on muscle and adipose tissue – which account for most of the glucose disposal capacity of the body – with focus on the unresolved discrepancies present in the literature.
Keywords: Insulin signaling; Insulin resistance; Type 2 diabetes; Adipose tissue; Skeletal muscle; Myotube;

Magnetic susceptibility of iron in malaria-infected red blood cells by S. Hackett; J. Hamzah; T.M.E. Davis; T.G. St Pierre (93-99).
During intra-erythrocytic maturation, malaria parasites catabolize up to 80% of cellular haemoglobin. Haem is liberated inside the parasite and converted to haemozoin, preventing haem iron from participating in cell-damaging reactions. Several experimental techniques exploit the relatively large paramagnetic susceptibility of malaria-infected cells as a means of sorting cells or investigating haemoglobin degradation, but the source of the dramatic increase in cellular magnetic susceptibility during parasite growth has not been unequivocally determined. Plasmodium falciparum cultures were enriched using high-gradient magnetic fractionation columns and the magnetic susceptibility of cell contents was directly measured. The forms of haem iron in the erythrocytes were quantified spectroscopically. In the 3D7 laboratory strain, the parasites converted approximately 60% of host cell haemoglobin to haemozoin and this product was the primary source of the increase in cell magnetic susceptibility. Haemozoin iron was found to have a magnetic susceptibility of (11.0 ± 0.9) × 10− 3 mL mol− 1. The calculated volumetric magnetic susceptibility (SI units) of the magnetically enriched cells was (1.88 ± 0.60) × 10− 6 relative to water while that of uninfected cells was not significantly different from water. Magnetic enrichment of parasitised cells can therefore be considered dependent primarily on the magnetic susceptibility of the parasitised cells.
Keywords: Haemozoin; Haemoglobin; Magnetic susceptibility; Malaria;

Notch signaling is an evolutionarily conserved mechanism that determines cell fate in a variety of contexts during development. This is achieved through different modes of action that are context dependent. One mode involves boundary formation between two groups of cells. With this mode of action, Notch signaling is central to vertebrate evolution as it drives the segmentation of paraxial mesoderm in the formation of somites, which are the precursors of the vertebra. In this case, boundary formation facilitates a mesenchymal to epithelial transition, leading to the creation of a somite. In addition, the boundary establishes a signaling center that patterns the somite, a feature that directly impacts on vertebral column formation. Studies in Xenopus, zebrafish, chicken and mouse have established the importance of Notch signaling in somitogenesis, and indeed in mouse how perturbations in somitogenesis affect vertebral column formation. Spondylocostal dysostosis is a congenital disorder characterized by formation of abnormal vertebrae. Here, mutation in Notch pathway genes demonstrates that Notch signaling is also required for normal somite formation and vertebral column development in humans; of particular interest here is mutation of the LUNATIC FRINGE (LFNG) gene, which causes SCD type 3. LUNATIC FRINGE encodes for a fucose-specific β1,3-N-acetylglucosaminyltransferase, which modifies Notch receptors and alters Notch signaling activity. This review will focus on Notch glycolsylation, and the role of LUNATIC FRINGE in somite formation and vertebral column development in mice and humans.
Keywords: Notch; Lunatic fringe; N-acetylglucosaminyltransferase; Somite; Vertebra; Spondylocostal dysostosis;

Characterization of the first FGFRL1 mutation identified in a craniosynostosis patient by Thorsten Rieckmann; Lei Zhuang; Christa E. Flück; Beat Trueb (112-121).
Fibroblast growth factor receptor-like 1 (FGFRL1) is a recently discovered transmembrane protein whose functions remain unclear. Since mutations in the related receptors FGFR1-3 cause skeletal malformations, DNA samples from 55 patients suffering from congenital skeletal malformations and 109 controls were searched for mutations in FGFRL1. One patient was identified harboring a frameshift mutation in the intracellular domain of this novel receptor. The patient showed craniosynostosis, radio-ulnar synostosis and genital abnormalities and had previously been diagnosed with Antley–Bixler syndrome. The effect of the FGFRL1 mutation was studied in vitro. In a reporter gene assay, the wild-type as well as the mutant receptor inhibited FGF signaling. However, the mutant protein differed from the wild-type protein in its subcellular localization. Mutant FGFRL1 was mainly found at the plasma membrane where it interacted with FGF ligands, while the wild-type protein was preferentially located in vesicular structures and the Golgi complex. Two motifs from the intracellular domain of FGFRL1 appeared to be responsible for this differential distribution, a tandem tyrosine based motif and a histidine-rich sequence. Deletion of either one led to the preferential redistribution of FGFRL1 to the plasma membrane. It is therefore likely that mutant FGFRL1 contributes to the skeletal malformations of the patient.
Keywords: Fibroblast growth factor (FGF); Fibroblast growth factor receptor (FGFR); FGFRL1; Craniosynostosis; Antley-Bixler syndrome ABS; Skeletal malformation;

Ectopic expression of Ligand-of-Numb protein X promoted TGF-β induced epithelial to mesenchymal transition of proximal tubular epithelial cells by Jing Nie; Qiaoyuan Wu; Wei Liu; Fengxin Zhu; Fanghua Qiu; Qin Zhou; Jinjin Fan; Xiuqing Dong; Xueqing Yu (122-131).
Ligand-of-Numb protein X (LNX) was initially characterized as a RING finger type E3 ubiquitin ligase that targeted the intrinsic cell fate determinant Numb for ubiquitination dependent degradation. However, the physiological function of LNX remains largely unknown. In the present study, we demonstrate that ectopic expression of LNX in human proximal tubular epithelial cells (HK-2 cells) significantly enhanced TGF-β1 induced epithelial to mesenchymal transition (EMT). The EMT-promoting effect of LNX manifested as strong inhibition of E-cadherin expression, enhanced expression of vimentin, fibronectin or PAI-1, and increased cell migration. This function of LNX was shown to be independent of its ligase activity because ectopic expression of a mutant form of LNX (C48ALNX) that lacks E3 ligase activity had the similar effect as the wild-type LNX. Overexpression of E-cadherin could inhibit LNX augmented EMT. This study suggests a potential role for LNX in promoting EMT in human proximal tubular epithelial cells.
Keywords: LNX; TGF-β; Epithelial to mesenchymal transition; Ubiquitin ligase; Proximal tubular epithelial cell;

Structure–function defects of the twinkle amino-terminal region in progressive external ophthalmoplegia by Teresa Holmlund; Géraldine Farge; Vineet Pande; Jenny Korhonen; Lennart Nilsson; Maria Falkenberg (132-139).
TWINKLE is a DNA helicase needed for mitochondrial DNA replication. In lower eukaryotes the protein also harbors a primase activity, which is lost from TWINKLE encoded by mammalian cells. Mutations in TWINKLE underlie autosomal dominant progressive external ophthalmoplegia (adPEO), a disorder associated with multiple deletions in the mtDNA. Four different adPEO-causing mutations (W315L, K319T, R334Q, and P335L) are located in the N-terminal domain of TWINKLE. The mutations cause a dramatic decrease in ATPase activity, which is partially overcome in the presence of single-stranded DNA. The mutated proteins have defects in DNA helicase activity and cannot support normal levels of DNA replication. To explain the phenotypes, we use a molecular model of TWINKLE based on sequence similarities with the phage T7 gene 4 protein. The four adPEO-causing mutations are located in a region required to bind single-stranded DNA. These mutations may therefore impair an essential element of the catalytic cycle in hexameric helicases, i.e. the interplay between single-stranded DNA binding and ATP hydrolysis.
Keywords: DNA replication; Mitochondrion; DNA helicase; TWINKLE;

Activating Fgfr3 Y367C mutation causes hearing loss and inner ear defect in a mouse model of chondrodysplasia by Stéphanie Pannier; Vincent Couloigner; Nadia Messaddeq; Monique Elmaleh-Bergès; Arnold Munnich; Raymond Romand; Laurence Legeai-Mallet (140-147).
Fibroblast growth factor receptor 3 (FGFR3) is a key regulator of skeletal development and activating mutations in FGFR3 cause skeletal dysplasias, including hypochondroplasia, achondroplasia and thanatophoric dysplasia. The introduction of the Y367C mutation corresponding to the human Y373C thanatophoric dysplasia type I (TDI) mutation into the mouse genome, resulted in dwarfism with a skeletal phenotype remarkably similar to that of human chondrodysplasia. To investigate the role of the activating Fgfr3 Y367C mutation in auditory function, the middle and inner ear of the heterozygous mutant Fgfr3 Y367C/+ mice were examined. The mutant Fgfr3 Y367C/+ mice exhibit fully penetrant deafness with a significantly elevated auditory brainstem response threshold for all frequencies tested. The inner ear defect is mainly associated with an increased number of pillar cells or modified supporting cells in the organ of Corti. Hearing loss in the Fgfr3 Y367C/+ mouse model demonstrates the crucial role of Fgfr3 in the development of the inner ear and provides novel insight on the biological consequences of FGFR3 mutations in chondrodysplasia.
Keywords: Chondrodysplasia; Fgfr3; Hearing loss; Cochlea; Pillar cell;

The role of redox status on chemokine expression in acute pancreatitis by S. Yubero; L. Ramudo; M.A. Manso; I. De Dios (148-154).
This study focused on the involvement of oxidative stress in the mechanisms mediating chemokine production in different cell sources during mild and severe acute pancreatitis (AP) induced by bile-pancreatic duct obstruction (BPDO) and 3.5% NaTc, respectively. N-Acetylcysteine (NAC) was used as antioxidant treatment. Pancreatic glutathione depletion, acinar overexpression of monocyte chemoattractant protein-1 (MCP-1) and cytokine-induced neutrophil chemoattractant (CINC), and activation of p38MAPK, NF-κB and STAT3 were found in both AP models. NAC reduced the depletion of glutathione in BPDO- but not in NaTc-induced AP, in which oxidative stress overwhelmed the antioxidant capability of NAC. As a result, inhibition of the acinar chemokine expression and signalling pathways occurs in mild, but not in severe AP. However, MCP-1 and CINC expressions in whole pancreas and plasma chemokine levels were not reduced by NAC, even in BPDO-induced AP, suggesting that in addition to acini, other pancreatic cells produced chemokines by antioxidant resistant mechanisms. The high Il-6 plasma levels found during AP, both in NAC-treated and non-treated rats, pointed out cytokines as activating factors of chemokine expression in non-acinar cells. In conclusion, from early AP oxidant-mediated MAPK, NF-κB and STAT3 activation triggers the chemokine expression in acini but not in non-acinar cells.
Keywords: Acinar cell; Acute pancreatitis; Chemokine; Oxidative stress; Signalling pathway;