Auxin regulation of a proton translocating ATPase in pea root plasma membrane vesicles

Pea root microsomal vesicles have been fractionated on a Dextran step gradient to give three fractions, each of which carries out ATP-dependent proton accumulation as measured by fluorescence quenching of quinacrine. The fraction at the 4/6% Dextran interface is enriched in plasma membrane, as determined by UDPG sterol glucosyltransferase and vanadate-inhibited ATPase. The vanadate-sensitive phosphohydrolase is not specific for ATP, has a K_m of about 0.23 millimolar for MgATP, is only slightly affected by K⁺ or Cl⁻ and is insensitive to auxin. Proton transport, on the other hand, is more specific for ATP, enhanced by anions (NO₃⁻ > Cl⁻) and has a K_m of about 0.7 millimolar. Auxins decrease the K_m to about 0.35 millimolar, with no significant effect on the V_max, while antiauxins or weak acids have no such effect. It appears that auxin has the ability to alter the efficiency of the ATP-driven proton transport.     

Dilute solution properties of proteoglycan fractions from bovine nasal cartilage

Fresh proteogycans (adult bovine nasal cartilage) isolated from the densest portion of a dissociative density gradient had a weight-average molecular weight of ca. 10⁶ in 4M guanidine hydrochloride (GdnHCI) by light scattering. Fractions of such material obtained by elution with 4M GdnHCI from 2% agarose gel, both normal and cross-linkd, has proteoglycan subunit molecular weights ranging from 0.8 to 2.6 × 10⁶ and root-mean-square radii ranging from 35 to 52 nm in the same solvent. The protein molecular weight per proteoglycan subunit was about 1.2 × 10⁵ and that of keratan sulfate about 1.8 × 10⁵, both independent of total molecular weight. A random-flight “graft copolymer” model having uniform side chains of chondroitin sulfate (40 disaccharides) and keratan sulfate (15 disaccharides) and a random-coil polypeptide back bone was used to estimate the unperturbed radius, whihc was about 19 nm for a mol wt of 1.5 × 10⁶. Experimental light-scattering data for fractions were fitted very well by theoretical curves for the particale scattering factor for both linear and appropriate branched polymers. Examination of coil expansion on the basis of perturbation calculations for branched polymer models suggested that expansion did not account for the experimentally observed radii in terms of unperturbed radii calculated from the model. A possible explanation is that substantial local stiffening of the polypeptide chain due to substitution of side-chain clusters increases the unperturbed radii. The intrinsic viscosity [η] is 4M GdnHCI ranged from 120 to 180 ml/g, and could be interpreted in terms of th eequivalent sphere model; the Flory number has approximately its normal value for flexible linear polymers. The treatment of the sedimentation coefficient by this is less successful, since the Man delkern-Flory parameter β apparently increases with increasing molecular weight; average value are similar to those for flexible polymers, but the variation in β makes this method useful only for rough estimation of molecular weight of proteoglycans. Molecular weights of purified proteoglycans are the same in 0.2M NaCI as in 4M GdnHCI, while crude preparations gave higher molecular weights in 0.2M NaCI, probably because of association due to incomplete removel of “linking” proteins.     

Comparison of the flowering behavior of the long-day plant Lemna gibba G3 from different laboratories

In an effort to determine the cause for the wide discrepanciesin the level of flowering response reported for the long-dayplant Lemna gibba L., strain G3, cultures of L. gibba G3 wereobtained from the laboratories of W. S. Hillman (G3-H), R. Kandeler(G3-K), Y. Oota (G3-O) and and A. Pieterse (G3-P) and comparedto the L. gibba G3 (G3-C) from this laboratory. Under continuouslight all cultures gave FL% values of 77 or above, and on a9L:15D short-day treatment, all cultures were completely vegetative.However, on daylengths of 10 to 12 hr, small but statisticallysignificant differences were obtained for the different cultures.The critical daylength curves for G3-G, which showed the shortestcritical daylength, and G3-K, which showed the longest criticaldaylength, differed by approximately one hour. Salicylic acidtreatment caused flower promotion in each culture, but statisticallysignificant differences were obtained between some of the culturesin their response to salicylic acid. It is concluded that the large discrepancies in the floweringresponses of L. gibba G3 that have been reported are due primarilyto differences in culturing methods and counting proceduresin the different laboratories. However, the results also indicatethat there may be distinct cultures of L. gibba G3 that exhibitsmall physiological and/or genetic differences that would makeprecise quantitative comparison between different laboratoriesvery difficult. (Received January 23, 1979; )     

Determination of the screw sense specificity of bovine liver fructokinase

Fructokinase from beef liver showed a clear reversal in specificity when the two isomers of ATP beta S were used as substrates with Mg2+ and Cd2+, with the Sp isomer having the higher V/K value with Mg2+ and the Rp isomer the higher value with Cd2+. The delta isomer of MgATP is thus the active form of the substrate. The substitution of sulfur for oxygen in the noncoordinated position of the beta-phosphate caused a 102-fold decrease in V/K over the value seen with MgATP, while substitution in the coordinated position gave a 21-fold decrease over the V/K value seen with CdATP. The Km values were little affected by sulfur substitution, showing that the wrong screw sense isomers were nonproductively bound almost as well as the correct ones. When ADP alpha S was used as a substrate in the reverse reaction, the Sp isomer showed the highest V/K value with both Mg2+ and Cd2+, suggesting that the metal ion is not coordinated to the alpha-phosphate during transphosphorylation. The failure of CrATP to act as a substrate for fructokinase suggests that the enzyme inserts one of its side chains into the inner coordination sphere of the metal ion during the reaction.     

A biological system for studies on mixing in bioreactors

A method for studying bioreactor inhomogeneity is suggested. Oxygen depletion in an E. coli cultivation leads to mixed acid fermentation with hydrogen gas production. Using a palladium metal-oxide semiconductor (Pd-MOS) hydrogen sensor on line in the effluent gas, it is shown that in a batch culture of 1 m³ hydrogen gas was evolved even before the dissolved oxygen tension (DOT) as measured by the probe reached zero. This suggests an appreciable degree of inhomogeneity with respect to DOT. Using measurements performed off line, it was observed that the lag time for hydrogen to appear became greater if the cells had been subjected to oxygen depletion. Lack of oxygen also resulted in a lowered hydrogen evolution capacity of the cells.     

Epistemological issues in the study of microbial life: alternative terran biospheres?

The assumption that all life on Earth today shares the same basic molecular architecture and biochemistry is part of the paradigm of modern biology. This paper argues that there is little theoretical or empirical support for this widely held assumption. Scientists know that life could have been at least modestly different at the molecular level and it is clear that alternative molecular building blocks for life were available on the early Earth. If the emergence of life is, like other natural phenomena, highly probable given the right chemical and physical conditions then it seems likely that the early Earth hosted multiple origins of life, some of which produced chemical variations on life as we know it. While these points are often conceded, it is nevertheless maintained that any primitive alternatives to familiar life would have been eliminated long ago, either amalgamated into a single form of life through lateral gene transfer (LGT) or alternatively out-competed by our putatively more evolutionarily robust form of life. Besides, the argument continues, if such life forms still existed, we surely would have encountered telling signs of them by now. These arguments do not hold up well under close scrutiny. They reflect a host of assumptions that are grounded in our experience with large multicellular organisms and, most importantly, do not apply to microbial forms of life, which cannot be easily studied without the aid of sophisticated technologies. Significantly, the most powerful molecular biology techniques available-polymerase chain reaction (PCR) amplification of rRNA genes augmented by metagenomic analysis-could not detect such microbes if they existed. Given the profound philosophical and scientific importance that such a discovery would represent, a dedicated search for 'shadow microbes' (heretofore unrecognized 'alien' forms of terran microbial life) seems in order. The best place to start such a search is with puzzling (anomalous) phenomena, such as desert varnish, that resist classification as 'biological' or 'nonbiological'.     

Efficacy and mechanisms of action of vitamin D in experimental polyarthritis

Vitamin D (vit D) status has been linked to the occurrence and severity of auto-immune and inflammatory diseases. This study evaluates the effects of vit D status on adoptive transfer of adjuvant-induced arthritis (ATA). Rats maintained on diets replete or deficient in vit D3 received arthritogenic thoracic duct cells and were monitored for severity of arthritis. CD45(+) cells obtained by collagenase digestion of hind-paw synovium-rich tissues (SRTs) were analysed to observe the effects of dietary vit D3 on the inflammatory process. Arthritis was more severe in vitamin D-deficient (vit-D(-)) rats compared with vitamin D-replete (vit-D(+)) rats. Resolution was delayed in vit-D(-) rats compared with vit-D(+) rats, or rats fed standard chow. During the acute phase of ATA, numbers of CD45(+) cells were significantly increased in the SRTs of vit-D(-) rats compared with vit-D(+) rats. This increase involved T-cells, polymorphonuclear leukocytes, macrophages, dendritic cells (DCs) and MHC II(hi) cells that resemble activated monocytes. A major difference between the dietary groups was that most DCs at the peak of inflammation in vit-D(-) rats were CD4(-), whereas in convalescent vit-D(+) rats most expressed CD4. Multiple categories of genes expressed by DCs differed between deficient and replete rats, with deficiency being associated with relative upregulation of certain pro-inflammatory genes and replete status being associated with upregulation of genes associated with resolution of inflammation. The findings indicate that ATA is more severe and prolonged in vit-D deficiency, that vit-D deficiency promotes accumulation of CD4(-) DCs in synovium during ATA and that a gene-expression profile is likely to contribute to the observed increased severity and duration of arthritis.