Thrombospondin1 in tissue repair and fibrosis: TGF-β-dependent and independent mechanisms

Thrombospondin 1 (TSP1) plays major roles in both physiologic and pathologic tissue repair. TSP1 through its type 1 repeats is a known regulator of latent TGF-β activation and plays a role in wound healing and fibrosis. Binding of the TSP N-terminal domain to cell surface calreticulin in complex with LDL-receptor related protein 1 stimulates intermediate cell adhesion, cell migration, anoikis resistance, collagen expression and matrix deposition in an in vivo model of the foreign body response. There is also emerging evidence that TSP EGF-like repeats alter endothelial cell-cell interactions and stimulate epithelial migration through transactivation of EGF receptors. The mechanisms underlying these functions of TSP1 and the implications for physiologic and pathologic wound repair and fibrosis will be discussed.     

Demographic History,Adaptation, and NRAP Convergent Evolution at Amino Acid Residue 100 in the World Northernmost Cattle from Siberia

Native cattle breeds represent an important cultural heritage. They are a reservoir of genetic variation useful for properly responding to agriculture needs in the light of ongoing climate changes. Evolutionary processes that occur in response to extreme environmental conditions could also be better understood using adapted local populations. Herein, different evolutionary histories of the world northernmost native cattle breeds from Russia were investigated. They highlighted Kholmogory as a typical taurine cattle, whereas Yakut cattle separated from European taurines approximately 5,000 years ago and contain numerous ancestral and some novel genetic variants allowing their adaptation to harsh conditions of living above the Polar Circle. Scans for selection signatures pointed to several common gene pathways related to adaptation to harsh climates in both breeds. But genes affected by selection from these pathways were mostly different. A Yakut cattle breed-specific missense mutation in a highly conserved NRAP gene represents a unique example of a young amino acid residue convergent change shared with at least 16 species of hibernating/cold-adapted mammals from six distinct phylogenetic orders. This suggests a convergent evolution event along the mammalian phylogenetic tree and fast fixation in a single isolated cattle population exposed to a harsh climate.     

Mechanistic and structural analyses of the role of His67 in the yeast polyamine oxidase Fms1

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N(1)-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N(1)-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2-3 orders of magnitude, with the H67Q mutation having the smallest effect and H67N the largest. The k(cat)/K(O2) value changes very little upon mutation with N(1)-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The k(cat)/K(M)-pH profiles with N(1)-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pK(a) values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pK(a) as wild-type Fms1, about ～7.4; this pK(a) is assigned to the substrate N4. The k(cat)/K(O2)-pH profiles for wild-type Fms1 and the H67A enzyme both show a pK(a) of about ～6.9; this suggests His67 is not responsible for this pH behavior. With the H67Q, H67N, and H67A enzymes the k(cat) value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 ?. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure.     

Divergence and heterogeneity of the histone gene repeating units in the Drosophila melanogaster species subgroup

The repeating units of the histone gene cluster containing the H1, H2A, H2B and H4 genes were amplified by PCR from the Drosophila melanogaster species subgroup, i.e., D. yakuba, D. erecta, D. sechellia, D. mauritiana, D. teissieri and D. orena. The PCR products were cloned and their nucleotide sequences of about 4.6-4.8kbp were determined to elucidate the mechanism of molecular evolution of the histone gene family. The heterogeneity among the histone gene repeating units was 0.6% and 0.7% for D. yakuba and D. sechellia, respectively, indicating the same level of heterogeneity as in the H3 gene region of D. melanogaster. Divergence of the genes among species even in the most closely related ones was much greater than the heterogeneity among family members, indicating a concerted mode of evolution for the histone gene repeating units. Among the species in the D. melanogaster species subgroup, the histone gene regions as well as 3rd codon position of the coding region showed nearly the same GC contents. These results suggested that the previous conclusion on analysis of the H3 gene regions, the gene family evolution in a concerted fashion, holds true for the whole histone gene repeating unit.     

In vivo studies on the roles of Tic110, Tic40 and Hsp93 during chloroplast protein import

A multisubunit translocon of the inner envelope membrane, termed Tic, mediates the late stages of protein import into chloroplasts. Membrane proteins, Tic110 and Tic40, and a stromal chaperone, Hsp93, have been proposed to function together within the Tic complex. In Arabidopsis, single genes, atTIC110 and atTIC40, encode the Tic proteins, and two homologous genes, atHSP93-V and atHSP93-III, encode Hsp93. These four genes exhibited relatively uniform patterns of expression, suggesting important roles for plastid biogenesis throughout development and in all tissues. To investigate the roles played by these proteins in vivo, we conducted a comparative study of T-DNA knockout mutants for each Tic gene, and for the most abundantly expressed Hsp93 gene, atHSP93-V. In the homozygous state, the tic110 mutation caused embryo lethality, implying an essential role for atTic110 during plastid biogenesis. Homozygous tic110 embryos exhibited retarded growth, developmental arrest at the globular stage and a 'raspberry-like' embryo-proper phenotype. Heterozygous tic110 plants, and plants homozygous for the tic40 and hsp93-V mutations, exhibited chlorosis, aberrant chloroplast biogenesis, and inefficient chloroplast-import of both photosynthetic and non-photosynthetic preproteins. Non-additive interactions amongst the mutations occurred in double mutants, suggesting that the three components may cooperate during chloroplast protein import.     

Microsatellite analysis of Rosa damascena Mill. accessions reveals genetic similarity between genotypes used for rose oil production and old Damask rose varieties

Damask roses are grown in several European and Asiatic countries for rose oil production. Twenty-six oil-bearing Rosa damascena Mill. accessions and 13 garden Damask roses were assayed by molecular markers. Microsatellite genotyping demonstrated that R. damascena Mill. accessions from Bulgaria, Iran, and India and old European Damask rose varieties possess identical microsatellite profiles, suggesting a common origin. At the same time, the data indicated that modern industrial oil rose cultivation is based on a very narrow genepool and that oil rose collections contain many genetically identical accessions. The study of long-term vegetative propagation of the Damask roses also reveals high somatic stability for the microsatellite loci analyzed.