Evolution of disintegrin cysteine-rich and mammalian matrix-degrading metalloproteinases: Gene duplication and divergence of a common ancestor rather than convergent evolution

The evolution of the Metalloproteinase Disintegrin Cysteine-rich (MDC) gene family and that of the mammalian Matrix-degrading Metalloproteinases (MMPs) are compared. The alignment of snake venom and mammalian MDC and MMP precursor sequences generated a phylogenetic tree that grouped these proteins mainly according to their function. Based on this observation, a common ancestry is suggested for mammalian and snake venom MDCs; it is also possible that gene duplication of the already-assembled domain structure, followed by divergence of the copies, may have significantly contributed to the evolution of the functionally diverse MDC proteins. The data also suggest that the structural resemblance of the zinc-binding motif of venom MDCs and MMPs may best be explained by common ancestry and conservation of the proteolytic motifs during the divergence of the proteins rather than through convergent evolution. Correspondence to: J.M. Crampton     

FIXATION PROBABILITIES OF SELFING RATE MODIFIERS IN SIMULATIONS WITH SEVERAL DELETERIOUS ALLELES WITH LINKAGE

The fixation rates of selfing rate modifiers were found by stochastic simulation in an infinite site model, including effects of several deleterious alleles with variable effects, which were randomly distributed in the genome without assuming any pollen discounting. Previous results on the evolution of selfing obtained by more precise methods were in this study further validated, and it was concluded that the effect of genetic associations on the evolution of mating systems is small except in the case of full pollen discounting. Furthermore, attention was given to the uneven distribution of the genetic load in the population, and the accompanying large among-genome variation in fixation rates. This among-genome variation will be of significance for the evolution of mating systems.     

Gluconate production by spores of Aspergillus niger

Spores of Aspergillus niger obtained by solid state fermentation on buckwheat seeds produced gluconic acid from glucose with a high yield, near 1.06 g gluconic acid/g glucose, close to the stoichiometric value. The reaction itself could be carried out either with purified biocatalyst or with the whole buckwheat medium resulting from spore production process. 200 g gluconic acid/L were obtained in 200 h with sequential feedings of glucose up to 190 g/L.     

Shape of the pelvis and posture of the hindlimbs inPlateosaurus

The pelvis ofPlateosaurus is examined from a biomechanical point of view. The shape of the acetabulum in particular is analysed in order to determine the range of possible directions of the forces exchanged between femur and pelvis. These forces must have been more or less confined to a sagittal plane. From a quasi-static analysis under consideration of the major hip muscles ofPlateosaurus, a nearly but not fully extended posture of the hindlimbs can be deduced. The hip joints ofPlateosaurus and probably of some other dinosaurs with a narrow biacetabular width were balanced rather by adducting than by abducting muscles.     

Detoxification of xenobiotics by plant cells: characterisation of vacuolar amphiphilic organic anion transporters

The transport activities of two primary ATP-dependent organic-anion transporters in the tonoplast of isolated barley (Hordeum vulgare L. cv. Klaxon) vacuoles have been characterised with N-ethylmaleimide glutathione (NEM-SG) and taurocholate as substrates. The transporters showed different sensitivities to organic anions and a variety of transport inhibitors and drugs. The vacuolar uptake of NEM-SG was inhibited by carbonylcyanide 4-trifluoromethoxyphenylhydrazone, 4,4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), S-(2,4-dinitrophenyl)glutathione, alkyl-S-glutathione derivatives and taurocholate but stimulated by probenecid. The uptake of taurocholate was inhibited by vinblastine, DIDS and probenecid. Both transporters were unaffected by verapamil. The kinetic properties of the transporters indicate a general preference for amphiphilic anions with some substrate overlap. These characteristics of the transporters are similar to those displayed by the multidrug resistance protein of mammalian drug-resistant cells. We suggest that these vacuolar transporters be described as plant multispecific organic anion transporters (pMOATs).Abbreviations Bm-S bimane S-glutathione - DIDS 4,4-diisothiocyanatostilbene-2,2-disulfonic acid - DNP-SG S-(2,4-dinitrophenyl)glutathione - FCCP carbonylcyanide 4-trifluoromethoxyphenylhydrazone - LTC₄ cysteinyl leukotriene - MDR multidrug transporter - MRP multidrug resistance protein - NEM-SG N-ethylmaleimide glutathione We thank Prof E. Martinoia for technical advice on the uptake experiments and Prof J. Palmer for helpful discussions and suggestions. M.B.-K. was partially sponsored by a grant from Stichting VSB Fonds, The Netherlands. IACR receives grant-aided support from the Biotechnology and Biological Science Research Council of the United Kingdom     

Identification and Characterization of a cDNA and the Structural Gene Encoding the Mouse Epithelial Membrane Protein-1

The PMP22/EMP/MP20 gene family includes four closely related proteins, peripheral myelin protein-22 (PMP22), epithelial membrane protein-1 (EMP-1), epithelial membrane protein-2 (EMP-2), and epithelial membrane protein-3 (EMP-3), which share amino acid identities ranging from 33 to 43%. In addition, the lens-specific membrane protein MP20 represents a more distant relative. Functionally, this family of proteins is likely to play important roles in the control of cell proliferation, cell differentiation, and cell death. In particular, mutations affecting thePMP22gene are responsible for various hereditary peripheral neuropathies in humans and mice. We report the isolation and characterization of a mouse EMP-1 cDNA and the correspondingemp-1gene. Mouse EMP-1 displays 93% amino acid identity to rat EMP-1 and 39% identity to mouse PMP22. The cDNA-predicted EMP-1 protein contains four putative membrane-associated domains and can beN-linked glycosylatedin vitro.EMP-1 is encoded by a single-copy gene with the positions of introns exactly conserved betweenemp-1andPMP22,corroborating the hypothesis that both genes belong to the same family. Computer-predicted structural domains of EMP-1 are partially mirrored by the exon/intron structure ofemp-1.Most interestingly, exon 4, which covers the potential second transmembrane domain, a small intracellular loop, and half of the third transmembrane domain, encodes the most highly conserved regions between the EMP-1 and PMP22 proteins and is also remarkably conserved in the MP20 gene, indicating some shared functional significance for this module in the PMP22/EMP/MP20 family.     

Homogenisation and oxygen transfer rates in large agitated and sparged animal cell bioreactors: Some implications for growth and production

Because of concern for cell damage, very low agitation energy inputs have been used in industrial animal cell bioreactors, typical values being two orders of magnitude less than those found in bacterial fermentations. Aeration rates are also very small. As a result, such bioreactors might be both poorly mixed and also unable to provide the higher oxygen up-take rates demanded by more intensive operation. This paper reports experimental studies both of K _L a and of mixing (via pH measurements) in bioreactors up to 8 m³ at Wellcome and of scaled down models of such reactors at Birmingham. Alongside these physical measurements, sensitivity of certain cell lines to continuously controlled dO₂ has been studied and the oxygen up-take rates measured in representative growth conditions. An analysis of characteristic times and mixing theory, together with other recent work showing that more vigorous agitation and aeration can be used especially in the presence of Pluronic F-68, indicates ways of improving their performance. pH gradients offer a special challenge.     

Localization and organization of phenol degradation genes ofPseudomonas putida strain H

The genetic organization of the DNA region encoding the phenol degradation pathway ofPseudomonas putida H has been investigated. This strain can utilize phenol or some of its methylated derivatives as its sole source of carbon and energy. The first step in this process is the conversion of phenol into catechol. Catechol is then further metabolized via themeta-cleavage pathway into TCA cycle intermediates. Genes encoding these enzymes are clustered on the plasmid pPGH1. A region of contiguous DNA spanning about 16 kb contains all of the genetic information necessary for inducible phenol degradation. The analysis of mutants generated by insertion of transposons and cassettes indicates that all of the catabolic genes are contained in a single operon. This codes for a multicomponent phenol hydroxylase andmeta-cleavage pathway enzymes. Catabolic genes are subject to positive control by the gene product(s) of a second locus.     

Responses to recent climatewarming of Pinus sylvestris and Pinus cembra within their montane transition zone in the Swiss Alps

Abstract. Altitudinal and latitudinal distribution limits of trees are mainly controlled by temperature. Therefore climate warming is expected to induce upslope or poleward migrations. In the Swiss Central Alps, summers in the period 1982-1991 were on average 0.8 °C warmer than those of the period 30 yr before. We investigated whether populations of conifers at the montane Pinus sylvestris-Pinus cembra ecocline exhibit demographic trends in response to that warming. We found no evidence for this. Young seedlings of Pinus sylvestris, the species which is expected to expand its range upward in a warmer climate, were virtually absent from all sites, whereas large fractions of Pinus cembra populations were observed in the seedling and juvenile categories even below the present lower distribution limit of adult trees. This suggests that there are no major altitudinal shifts in response to the recent sequence of warmer summers. Germination and seedling survival trials with Pinus sylvestris suggest that temperature per se would not exclude this species even from establishing at the current treeline in the Swiss Central Alps. Similar results were found at the polar treeline. Phytotron tests of seedling survival showed much less drought resistance in Pinus sylvestris than in Pinus cembra which is in contrast to their phytogeographic distributions. Thus, the montane pine ecocline in the Swiss Central Alps seems to be stabilized by species interactions and may not be directly responsive to moderate climatic change, which needs to be taken into account in predictive attempts.