A model of NaCl and water flow through paracellular pathways of renal proximal tubules.

To explain how hydrostatic pressure differences between tubule lumen and interstitium modulate isotonic reabsorption rates, we developed a model of NaCl and water flow through paracellular pathways of the proximal tubule. Structural elements of the model are a tight junction membrane, an intercellular channel whose walls transport NaCl actively at a constant rate, and a basement membrane. Equations of change were derived for the channel, boundary conditions were formulated from irreversible thermodynamics, and a pressure-area relationship typical of thin-walled tubing was assumed. The boundary value problem was solved numerically. The principal conclusions are: 1) channel NaCl concentration must remain within a few mOsm of isotonic values for reabsorption rates to be modulated by transtubular pressure differences known to affect this system: 2) basement membrane and channel wall parameters determine reabsorbate tonicity; tight junction parameters affect the sensitivity of reabsorption to transmural pressure; 3) channel NaCl concentration varies inversely with transmural pressure difference; this concentration variation controls NaCl diffusion through the tight junction; 4) modulation of NaCl diffusion through the tight junction controls the rate of isotonic reabsorption; modulation of water flow can increase sensitivity to transmural pressure; 5) no pressure-induced change in permeability of the tight junction or basement membrane is needed for pressure to modulate reabsorption; and 6) system performance is indifferent to the distribution of active transport sites, to the numerical value of the compliance function, and to the relationship between lumen and cell pressures.     

Deoxyribonucleic acid reassociation in the taxonomy of the genus Chlorella

DNA hybridization techniques showed Chlorella fusca var. vacuolata and C. kessleri to be homogeneous species with DNA homologies of 90–100% C. fusca var. fusca and var. rubescens, however, have only about 15% DNA homology with C. fusca var. vacuolata and should no longer be regarded as varieties. A good correlation was found so far between biochemical and physiological characters used in the taxonomy of Chlorella and DNA relatedness. Mutant strains of Chlorella were tested for DNA homologies to prove the reliability of the taxonomical interpretation.     

Die Beeinflussung der Aminosäuresequenz des serinhaltigen Mureins von Staphylococcus epidermidis Stamm 24 durch die Nährbodenzusammensetzung

Zusammenfassung Beim Wachstum von S. epidermidis, Stamm 24, in Hefe-Dextrose-Bouillon weist das Murein folgende Molverhältnisse auf (auf- bzw. abgerundete Zahlen): Mur-GlcNH₂:Ala:Glu:Lys:Gly:Ser=1:1:2,4:1:1:4,2:0,6. Die Glutaminsäure ist amidiert.Durch Isolierung und Identifizierung der Peptide des Partialhydrolysates des Mureins wurde die Aminosäuresequenz bestimmt. Die an die Muraminsäure gebundene Peptiduntereinheit (l-Ala-d-GluNH₂ -l-Lys-d-Ala) stimmt mit der von S. aureus, Copenhagen bzw. S. epidermidis, Stamm 66, überein. Bei knapp einem Drittel der Peptiduntereinheiten ist das C-terminale d-Alanin der Mureinvorstufe nicht abgespalten, so daß diese noch als Pentapeptide vorliegen. Dies konnte aus dem Verhältnis l-Ala/d-Ala (1:1,3), dem Ergebnis der Hydrazinolyse und der Isolierung der Muropeptide nach Spaltung der Zellwände mit Lysozym geschlossen werden.Bei etwa der Hälfte aller aus 5 Glycinresten aufgebauten Interpeptidketten ist ein Glycinrest durch l-Serin ersetzt. Die genaue Position des Serins konnte nicht bestimmt werden. Serin ist sicher nicht direkt an die -Aminogruppe des Lysins gebunden.In selteneren Fällen kann Lysin mit l-Alanin substituiert sein, das N-terminal vorliegt und nicht der Quervernetzung dient.Die Dinitrophenylierung des Mureins ergab, daß in etwa 3,5% der Fälle die Interpeptidketten fehlen und rund ein Drittel der Interpeptidketten nicht quervernetzt ist.Bei Wachstum in einem halbsynthetischen, glycinarmen Medium (Minimal, medium) nimmt der Glycinanteil des Mureins um rund 40% ab, während l-Alanin zunimmt. Es konnte gezeigt werden, daß rund 15% des Mureins ein an die -Amino-gruppe des Lysins gebundenes l-Alanin enthalten, das aber im Unterschied zu S. epidermidis, Stamm 66, nicht mit Glycin substituiert ist, sondern N-terminal bleibt und nicht zur Quervernetzung benützt werden kann. Weiterhin liegen hier rund 35% des Lysins unsubstituiert vor, und nur etwa 50% der Peptiduntereinheiten weisen eine Pentaglycyl-Interpeptidkette auf. Die Quervernetzung des Mureins ist bei den in Minimalmedium gewachsenen Zellen nur zu rund 30% durchgeführt. Bei Zusatz von Glycin zum Minimal-Nährboden wird der Glycingehalt im Murein erhöht, während der extra Alaninanteil praktisch verschwindet. Serinzusatz erhöht nicht nur den Serin-, sondern auch den Glycinanteil. Bei Alaninzusatz dagegen wird der Alaningehalt im Murein etwas erhöht und der Glycingehalt weiter erniedrigt.Die Ergebnisse dieser Untersuchungen wurden mit entsprechenden, vorläufigen Versuchen bei S. epidermidis, Stamm 66 und S. aureus, Stamm Copenhagen, verglichen. Es zeigt sich, daß trotz starker modifikativer Veränderungen der Mureinzusammensetzung eindeutige genetische Unterschiede zwischen diesen drei Stämmen vorliegen.

---

The effect of nutrition on the amino acid sequence of the serine containing murein of Staphylococcus epidermis strain 24

Summary The murein (peptidoglycan) of S. epidermidis strain 24 contains Mur GlcNH₂, Ala, Glu, Lys, Gly, Ser at a molar ratio of about 1:1:2.4:1:1:4.2:0.6 when grown in a yeast extract dextrose medium. Glutamic acid occurs as an amide.The amino acid sequence was determined by analysing the oligopeptides from partial acid hydrolysate. The tetrapeptide bound to the muramic acid (l-Ala-d-Glu-NH₂-l-Lys-d-Ala) is identical with those found in S. aureus and S. epidermidis strain 66. About 1/3 of the muropeptides is still present as pentapeptides, since the second d-alanine of the muramyl pentapeptide precursor is not split off. This fact is indicated by the ratio of l-Ala/d-Ala of 1:1.3, the isolation of muropentapeptides from the lysozyme lysates and by the result of the hydrazinolysis.About 50% of the pentaglycine interpeptide chains contain one mole of l-serine. The exact position of l-serine could not be determined. However, it could be shown, that serine is never bound to the -amino group of lysine. In very rare cases, the -amino group of lysine is substituted by l-alanine which remains N-terminal and can not be used for crosslinkages.As shown by dinitrophenylation, about 3.5% of the -amino groups of lysine is free and about 50% of the interpeptide chains are not cross-linked.If the organism is grown in a glycine deficient minimal medium, the glycine content of the murein drops by 40%, while l-alanine increases. Here, about 15% of the -amino groups of lysine is substituted by l-alanine, which again is not used for cross-linkages. Another 35% of the -amino groups of lysine remain free. From the existing interpeptide chains 30% are not cross-linked.The addition of glycine to the minimal medium causes an increase of the glycine content in the murein, however, the extra alanine protion nearly disappears. The addition of serine leads to an increase of not only the serine portion but also the glycine portion in the murein. However, when alanine is added the alanine portion of murein is slightly increased and the glycine portion further decreased.The results of these experiments were compared to corresponding preliminary experiments with S. epidermidis (strain 66) and S. aureus (strain Copenhagen). In spite of modificative changes in the murein composition, clear genetical differences between the 3 strains were obvious.

     

Functional analysis of a complex oncogene arrangement in biotype III Agrobacterium tumefaciens strains

The ubiquitous grapevine-associated octopine/cucumopine Ti plasmids of biotype III Agrobacterium tumefaciens strains carry two T regions, TA and TB, with a complex oncogene arrangement. Within the octopine/cucumopine group, two main strain types were identified: large TA strains with a TA region resembling the TL region of the biotype I octopine strain Ach5 and small TA strains with a similar T region organization as the large TA strains but with a large internal TA deletion. Structural and functional studies of the representative large TA strain Tm4 revealed six oncogenes. Each oncogene was inserted in a disarmed vector and tested for biological activity using the corresponding oncogenes of Ach5 as standards. Five Tm4 oncogenes, TA-iaaM, T-ipt, T-6b, TB-iaaH and TB-iaaM, were shown to be active, the IS-interrupted TA-iaaH gene was inactive. To study the role of each gene in the pTiTm4 context, several single and multiple pTiTm4 mutations were constructed. It was shown that whereas TA-iaaM and TB-iaaH are essential for tumour formation on grapevine, T-ipt, T-6b and TB-iaaM are not. The avirulence of the TA-iaaM ^- mutant was shown to be due to an inhibitory effect of the T-ipt gene, since a TA-iaaM ^- /T-ipt ^- double mutant was fully virulent. We conclude that the TA-iaaM gene of large TA strains is specifically required to counteract the tumour growth inhibiting activity of the T-ipt gene. Both TA-iaaM and T-ipt are absent from the small TA strains. A model on the roles and interactions of the different oncogenes in large TA and small TA strains is presented.     

Molecular evolution of pteridophytes and their relationship to seed plants: Evidence from complete 18S rRNA gene sequences

Complete 18S ribosomal RNA sequence data from representatives of all extant pteridophyte lineages together with RNA sequences from different seed plants were used to infer a molecular phylogeny of vascular plants that included all major land plant lineages. The molecular data indicate that lycopsids are monophyletic and are the earliest diverging group within the vascular land plants, whereasPsilotum nudum is more closely related to the seed plants than to other pteridophyte lineages. The phylogenetic trees based on maximum likelihood, parsimony and distance analyses show substantial agreement with the evolutionary relationships of land plants as interpreted from the fossil record.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.