The relationship between turgor pressure and titratable acidity in mesophyll cells of intact leaves of a Crassulacean-acid-metabolism plant,Kalanchoe daigremontiana Hamet et Perr

Day/night changes in turgor pressure (P) and titratable acidity content were investigated in the (Crassulacean-acid-metabolism (CAM) plant Kalanchoe daigremontiana. Measurements of P were made on individual mesophyll cells of intact attached leaves using the pressure-probe technique. Under conditions of high relative humidity, when transpiration rates were minimal, changes in P correlated well with changes in the level of titratable acidity. During the standard 12 h light/12 h dark cycle, maximum turgor pressure (0.15 MPa) occurred at the end of the dark period when the level of titratable acidity was highest (about 300 eq H⁺·g^-1 fresh weight). A close relationship between P and titratable acidity was also seen in leaves exposed to perturbations of the standard light/dark cycle. (The dark period was either prolonged, or else only CO₂-free air was supplied in this period). In plants deprived of irrigation for five weeks, diurnal changes in titratable acidity of the leaves were reduced (H=160 eq H⁺·g^-1 fresh weight) and P increased from essentially zero at the end of the light period to 0.02 MPa at the end of the dark period. Following more severe water stress (experiments were made on leaves which had been detached for five weeks), P was zero throughout day and night, yet small diurnal changes in titratable acidity were still measured. These findings are discussed in relation to a hypothesis by Lüttge et al. 1975 (Plant Physiol. 56,613-616) for the role of P in the regulation of acidification/de-acidification cycles of plants exhibiting CAM.Abbreviations CAM crassulacean acid metabolism - FW fresh weight - P turgor pressure     

Posttranslational modification and microtubule stability

We have probed the relationship between tubulin posttranslational modification and microtubule stability, using a variation of the antibody-blocking technique. In human retinoblastoma cells we find that acetylated and detyrosinated microtubules represent congruent subsets of the cells' total microtubules. We also find that stable microtubules defined as those that had not undergone polymerization within 1 h after injection of biotin-tubulin were all posttranslationally modified; furthermore dynamic microtubules were all unmodified. We therefore conclude that in these cells the stable, acetylated, and detyrosinated microtubules represent the same subset of the cells' total network. Posttranslational modification, however, is not a prerequisite for microtubule stability and vice versa. Potorous tridactylis kidney cells have no detectable acetylated microtubules but do have a sizable subset of stable ones, and chick embryo fibroblast cells are extensively modified but have few stable microtubules. We conclude that different cell types can create specific microtubule subsets by modulating the relative rates of posttranslational modification and microtubule turnover.     

Experimental evidence on the affinities ofPapaver somniferum (Papaveraceae)

Results obtained from crossing experiments betweenP. somniferum subsp.somniferum (2n = 22) and subsp.setigerum (2n = 44),P. glaucum (2n = 14) andP. gracile (2n = 14) and from the observation of meiotic chromosome pairing in the various hybrids obtained do not provide straightforward evidence for the hypothesis thatP. somniferum originated as a triploid hybrid between taxa similar toP. glaucum andP. gracile (Kadereit 1986a, b).—On the one hand, the pattern of crossability found reflects the closer similarity of subsp.somniferum toP. glaucum and of subsp.setigerum toP. gracile, which was interpreted as segregation of parental characters, and the high frequency of 2n = 28 chromosomes among F₂-progeny from the hybrid subsp.somniferum × subsp.setigerum (2n = 33) might reveal n = 7 as the base number also ofP. somniferum. On the other hand, however, the general difficulty of obtaining hybrids, and the low incidence of bivalent formation in their meiosis, probably indicating a lack of chromosome homology between the different species, do not fit the above hypothesis.—These results are in marked contrast to the morphological similarity between the three species involved.     

Incorporation of highly purified melittin into phosphatidylcholine bilayer vesicles

Melittin free of phospholipase A2 was prepared. In the absence of salt this highly pure protein starts to aggregate in solution at a protein concentration of Cp greater than 10(-3) M. In high salt solution (2 M) aggregation starts at Cp greater than 10(-6) M. This was determined from the blue shift of the intrinsic fluorescence of the protein. Reinvestigation of the quenching behaviour clearly shows that self-aggregation cannot be deduced from quenching experiments using nitrate or 2,2,6,6-tetramethylpiperidine-1-oxyl as quencher. The incorporation of melittin into phosphatidylcholine bilayer vesicles was studied by fluorescence quenching and by energy-transfer experiments using 2- and 6-anthroyloxypalmitic acid as acceptor and peptide tryptophan as donor. Incorporation of melittin into small unilamellar vesicles was found to be reduced below the lipid phase transition temperature, Tt, whereas it incorporates and distributes more randomly above Tt. Cooling the temperature below Tt after incubation at T greater than Tt leads to a deeper incorporation of the peptide into the lipid bilayer due to electrostatic interaction between the lipid phosphate groups and the positively charged amino acids. This stabilizing effect is lost above Tt and melittin is extruded to the polar phase. Quenching experiments support this finding. EPR measurements clearly demonstrate that even in the presence of high amounts of melittin up to 10 mol% with respect to the lipid broadening of the phase transition curves was only observed with fatty acid spin labels, where the doxyl group is localized near the bilayer surface. The order degree of the inner part of the bilayer remains almost unchanged even in the presence of high melittin content.     

Regression of blood vessels precedes cartilage differentiation during chick limb development

We have previously investigated distinct areas of vascular regression in the developing vascular system of the chick limb bud. Avascular areas appear in a characteristic spatial and temporal pattern, and are correlated with the position of developing cartilage. In the present study, we examined limb-bud sections which had been double labeled for endothelial cells and developing cartilage in order to determine the relationship between the appearance of cartilage and the disappearance of capillaries. Endothelial cells, which specifically take up acetylated low-density lipoprotein (acLDL), were labeled by intravenously injecting fluorescent acLDL (DiIacLDL) into chick embryos at Hamburger and Hamilton stages 26-30. Avascular zones, which correspond to the developing digits, were clearly visible within the fluorescently labeled distal vasculature. The same sections were labeled with monoclonal antibodies specific for cartilage. We found that progressing avascularity in the digital regions was followed by increased staining for cartilage antigens in the same areas. Zones of avascularity always developed earlier than morphologically and immunologically detectable cartilage in all planes of section and were always larger than the areas of cartilage. These results demonstrate that blood vessels disappear in predictable areas prior to the overt differentiation of cartilage.     

Mapping of antibody specificities to V H gene families

V _H gene segments represent the products of the repeated duplication and subsequent diversification of a primordial V gene element. It is widely assumed that natural selection, operating via pathogens, has played the dominant role in this process. Here, we screen some 3.7 × 10⁴ C ⁺ colonies of mitogen-activated B cells for the production of antibodies specific for phosphorylcholine or hen egg lysozyme and expression of the V _H X-24, S107, Q52, or J558 gene families. These gene families were expressed at frequencies proportional to their genomic complexity among both unselected and antigen-specific C ⁺ colonies. Thus, the capacity to encode equivalent antibody-combining sites is dispersed uniformly among V _Hfamilies. This result suggests that individual V _H genes have not evolved to address specific antigens.     

The distribution of the Hb Constant Spring gene in Southeast Asian populations

Summary The distribution of the hemoglobin Constant Spring (Hb CS) gene in eight populations in Southeast Asia (including Assam) was determined using oligonucleotide hybridization. Hb CS was absent in two Assamese populations with a high prevalence of Hb E. The Hb CS gene frequency was 0.033 in northern Thailand and near 0.01 in central Thailand and Cambodia. High frequencies, between 0.05 and 0.06, were observed in northeastern Thailand. The present data and a similar study in Laotians suggest that the Lao-speaking populations of the Mekong River basin in northeastern Thailand and Laos have the highest frequencies of the Hb CS gene in Southeast Asia.     

Purification of acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyltransferase from Catharanthus roseus leaves

The enzyme acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyltransferase which terminates vindoline biosynthesis has been isolated from Catharanthus roseus leaves, further characterized and purified to homogeneity by three step column chromatography and subsequent preparative isoelectric focusing. Kinetic properties concerning the enzyme reaction are discussed. Five multiple forms of the acetyl-transferase could be observed, each consisting of two subunits. This enzyme is now the best characterized of the enzymes involved in vindoline biosynthesis.Abbreviations DTE dithiothreitol - EDTA ethylenediamine-tetraacetic acid - HEPES N-(2-hydroxyethyl)-piperazine-N-2-ethanesulfonic acid - IEF isoelectric focusing - KP_i potassium phosphate - M_r rel.molecular mass - PEG polyethylene glycol - SDS-PAGE sodium dodecylsulfate polyacrylamide gel electrophoresis - Tris 2-amino-2-(hydroxymethyl)-1,3-propandiol     

Maternal-embryonic relationships in the goodeid teleost,Xenoophorus captivus

In goodeid teleosts, prolonged embryonic development takes place within the ovarian cavity. Apposed maternal and embryonic epithelia interface via a nutritive liquid (embryotrophe) and facilitate aplacental matrotrophy. The role of the internal ovarian epithelium (IOE) in providing proteins for the embryotrophe has been studied using transmission electron-microscopic examinations of both the resting and the active ovarian lining, and isoelectric focusing of embryotrophe and maternal blood serum. The simple IOE is apparently composed of only one, filament-containing cell-type. In the non-gravid ovary these cells are cuboidal to columnar in shape, and are either compact and electron-dense or oedematous and light. During gestation, swelling of the ovarian connective tissue gives rise to dovetailing of the IOE with the subjacent capillary plexus. Part of the IOE overlying the capillaries becomes stretched, resulting in a thin endothelium-like demarcation. The nuclei and the bulk of the cytoplasm are usually recessed between the meshes of the protruding capillary network. The blood-embryotrophe pathway is thus reduced in places, to less than one m. The active form of the IOE contains a well-developed vacuolar apparatus composed of small vesicles, vacuoles, multivesicular bodies, and a few lysosomes. Elements of the RER are sparsely distributed throughout the cytoplasm. Endocytotic activity is observable at the apical and basolateral plasma membrane. Isoelectric focusing of both serum and embryotrophe produces numerous bands each between pI 4–8, which reveal many homologies. The intensity of corresponding bands varies considerably. It is concluded that the cells of the IOE provide a transport pathway for serum-derived macromolecular substances rather than produce proteinaceous secretions.