Loss of Omi mitochondrial protease activity causes the neuromuscular disorder of mnd2 mutant mice

The mouse mutant mnd2 (motor neuron degeneration 2) exhibits muscle wasting, neurodegeneration, involution of the spleen and thymus, and death by 40 days of age. Degeneration of striatal neurons, with astrogliosis and microglia activation, begins at around 3 weeks of age, and other neurons are affected at later stages. Here we have identified the mnd2 mutation as the missense mutation Ser276Cys in the protease domain of the nuclear-encoded mitochondrial serine protease Omi (also known as HtrA2 or Prss25). Protease activity of Omi is greatly reduced in tissues of mnd2 mice but is restored in mice rescued by a bacterial artificial chromosome transgene containing the wild-type Omi gene. Deletion of the PDZ domain partially restores protease activity to the inactive recombinant Omi protein carrying the Ser276Cys mutation, suggesting that the mutation impairs substrate access or binding to the active site pocket. Loss of Omi protease activity increases the susceptibility of mitochondria to induction of the permeability transition, and increases the sensitivity of mouse embryonic fibroblasts to stress-induced cell death. The neurodegeneration and juvenile lethality in mnd2 mice result from this defect in mitochondrial Omi protease.     

Nanoindentation. Simulation of defect nucleation in a crystal

Nanoindentation is the penetration of a surface to nanometre depths using an indenting device. It can be simulated using the Bragg bubble-raft model, in which a close-packed array of soap bubbles corresponds to the equilibrium positions of atoms in a crystalline solid. Here we show that homogeneous defect nucleation occurs within a crystal when its surface roughness is comparable to the radius of the indenter tip, and that the depth of the nucleation site below the surface is proportional to the half-width of the contact. Our results may explain the unusually high local stress required for defect nucleation in nano-indented face-centred cubic crystals.     

GSK-3α/β kinases and amyloid production in vivo

Arising from C. J. Phiel, C. A. Wilson, V. M.-Y. Lee & P. S. Klein 423, 435-439 (2003)A major unresolved issue in Alzheimer's disease is identifying the mechanisms that regulate proteolytic processing of amyloid precursor protein (APP)-glycogen synthase kinase-3 (GSK-3) isozymes are thought to be important in this regulation. Phiel et al. proposed that GSK-3α, but not GSK-3β, controls production of amyloid. We analysed the proteolytic processing of mouse and human APP in mouse brain in vivo in five different genetic and viral models. Our data do not yield evidence for either GSK-3α-mediated or GSK-3β-mediated control of APP processing in brain in vivo.     

X-ray structures of general anaesthetics bound to a pentameric ligand-gated ion channel

General anaesthetics have enjoyed long and widespread use but their molecular mechanism of action remains poorly understood. There is good evidence that their principal targets are pentameric ligand-gated ion channels (pLGICs) such as inhibitory GABA(A) (γ-aminobutyric acid) receptors and excitatory nicotinic acetylcholine receptors, which are respectively potentiated and inhibited by general anaesthetics. The bacterial homologue from Gloeobacter violaceus (GLIC), whose X-ray structure was recently solved, is also sensitive to clinical concentrations of general anaesthetics. Here we describe the crystal structures of the complexes propofol/GLIC and desflurane/GLIC. These reveal a common general-anaesthetic binding site, which pre-exists in the apo-structure in the upper part of the transmembrane domain of each protomer. Both molecules establish van der Waals interactions with the protein; propofol binds at the entrance of the cavity whereas the smaller, more flexible, desflurane binds deeper inside. Mutations of some amino acids lining the binding site profoundly alter the ionic response of GLIC to protons, and affect its general-anaesthetic pharmacology. Molecular dynamics simulations, performed on the wild type (WT) and two GLIC mutants, highlight differences in mobility of propofol in its binding site and help to explain these effects. These data provide a novel structural framework for the design of general anaesthetics and of allosteric modulators of brain pLGICs.     

Activation-independent binding of human memory T cells to adhesion molecule ELAM-1.

The induction of an ensemble of adhesion molecules on endothelial cells by inflammatory cytokines is likely to be crucial to the differential migration of T-lymphocyte subsets into inflammatory sites. Two molecular pathways involving the VLA-4 and LFA-1 integrins are known to mediate T-cell adhesion to activated endothelium. Here we show that a third pathway involving the rapidly inducible endothelial cell-surface adhesion molecule ELAM-1 contributes to the binding of resting CD4+ T cells to IL-1-induced human endothelial cells. All three pathways contribute to the greater adhesion to endothelium of memory T cells than naive T cells. There are two unique features of T-cell adhesion to purified ELAM-1: first, ELAM-1 exclusively mediates adhesion of memory T cells; second, memory T-cell binding to ELAM-1 is independent of acute activation events that regulate integrin-mediated adhesion. Thus, ELAM-1 may be of primary importance in the initial attachment of memory T cells to inflamed endothelium in vivo and to the preferential migration of memory T cells into tissue and inflammatory sites.     

Apolipoprotein L-I is the trypanosome lytic factor of human serum

Human sleeping sickness in east Africa is caused by the parasite Trypanosoma brucei rhodesiense. The basis of this pathology is the resistance of these parasites to lysis by normal human serum (NHS). Resistance to NHS is conferred by a gene that encodes a truncated form of the variant surface glycoprotein termed serum resistance associated protein (SRA). We show that SRA is a lysosomal protein, and that the amino-terminal alpha-helix of SRA is responsible for resistance to NHS. This domain interacts strongly with a carboxy-terminal alpha-helix of the human-specific serum protein apolipoprotein L-I (apoL-I). Depleting NHS of apoL-I, by incubation with SRA or anti-apoL-I, led to the complete loss of trypanolytic activity. Addition of native or recombinant apoL-I either to apoL-I-depleted NHS or to fetal calf serum induced lysis of NHS-sensitive, but not NHS-resistant, trypanosomes. Confocal microscopy demonstrated that apoL-I is taken up through the endocytic pathway into the lysosome. We propose that apoL-I is the trypanosome lytic factor of NHS, and that SRA confers resistance to lysis by interaction with apoL-I in the lysosome.     

Craniometric evidence for Palaeoamerican survival in Baja California

A current issue on the settlement of the Americas refers to the lack of morphological affinities between early Holocene human remains (Palaeoamericans) and modern Amerindian groups, as well as the degree of contribution of the former to the gene pool of the latter. A different origin for Palaeoamericans and Amerindians is invoked to explain such a phenomenon. Under this hypothesis, the origin of Palaeoamericans must be traced back to a common ancestor for Palaeoamericans and Australians, which departed from somewhere in southern Asia and arrived in the Australian continent and the Americas around 40,000 and 12,000 years before present, respectively. Most modern Amerindians are believed to be part of a second, morphologically differentiated migration. Here we present evidence of a modern Amerindian group from the Baja California Peninsula in Mexico, showing clearer affinities with Palaeoamerican remains than with modern Amerindians. Climatic changes during the Middle Holocene probably generated the conditions for isolation from the continent, restricting the gene flow of the original group with northern populations, which resulted in the temporal continuity of the Palaeoamerican morphological pattern to the present.     

The Polycomb group protein EZH2 directly controls DNA methylation

The establishment and maintenance of epigenetic gene silencing is fundamental to cell determination and function. The essential epigenetic systems involved in heritable repression of gene activity are the Polycomb group (PcG) proteins and the DNA methylation systems. Here we show that the corresponding silencing pathways are mechanistically linked. We find that the PcG protein EZH2 (Enhancer of Zeste homolog 2) interacts-within the context of the Polycomb repressive complexes 2 and 3 (PRC2/3)-with DNA methyltransferases (DNMTs) and associates with DNMT activity in vivo. Chromatin immunoprecipitations indicate that binding of DNMTs to several EZH2-repressed genes depends on the presence of EZH2. Furthermore, we show by bisulphite genomic sequencing that EZH2 is required for DNA methylation of EZH2-target promoters. Our results suggest that EZH2 serves as a recruitment platform for DNA methyltransferases, thus highlighting a previously unrecognized direct connection between two key epigenetic repression systems.