Root Growth Maintenance at Low Water Potentials (Increased Activity of Xyloglucan Endotransglycosylase and Its Possible Regulation by Abscisic Acid)

Previous work suggested that an increase in cell wall-loosening contributes to the maintenance of maize (Zea mays L.) primary root elongation at low water potentials ([psi]w). It was also shown that root elongation at low [psi]w requires increased levels of abscisic acid (ABA). In this study we investigated the effects of low [psi]w and ABA status on xyloglucan endotransglycosylase (XET) activity in the root elongation zone. XET is believed to contribute to wall-loosening by reversibly cleaving xyloglucan molecules that tether cellulose microfibrils. The activity of XET per unit fresh weight in the apical 10 mm (encompassing the elongation zone) was constant at high [psi]w but increased by more than 2-fold at a [psi]w of -1.6 MPa. Treatment with fluridone to decrease ABA accumulation greatly delayed the increase in activity at low [psi]w. This effect was largely overcome when internal ABA levels were restored by exogenous application. Spatial distribution studies showed that XET activity was increased in the apical 6 mm at low [psi]w whether expressed per unit fresh weight, total soluble protein, or cell wall dry weight, corresponding to the region of continued elongation. Treatment with fluridone progressively inhibited the increase in activity with distance from the apex, correlating with the pattern of inhibition of elongation. Added ABA partly restored activity at all positions. The increase in XET activity at low [psi]w was due to maintenance of the rate of deposition of activity despite decreased deposition of wall material. The loss of activity associated with decreased ABA was due to inhibition of the deposition of activity. The results demonstrate that increased XET activity is associated with maintenance of root elongation at low [psi]w and that this response requires increased ABA.     

Growth of Arabidopsis thaliana seedlings under water deficit studied by control of water potential in nutrient-agar media

We have characterized the growth responses of Arabidopsis thaliana seedlings to water deficit. To manipulate the water potential, we developed a method whereby the nutrient-agar medium could be supplemented with polyethylene glycol (PEG 8000); PEG was introduced into gelled media by diffusion, which produced media with water potential as low as -1.6 MPa. For dark-grown plants, hypocotyl growth had a hyperbolic dependence on water potential, and was virtually stopped by -1 MPa. In contrast, primary root elongation was stimulated by moderate deficit and even at -1.6 MPa was not significantly less than the control. That these results did not depend on a direct effect of PEG was attested by obtaining indistinguishable results when a dialysis membrane impermeable to PEG was placed between the medium and the seedlings. For light-grown seedlings, moderate deficit also stimulated primary root elongation and severe deficit reduced elongation only partially. These changes in elongation were paralleled by changes in root system dry weight. At moderate deficit, lateral root elongation and initiation were unaffected and at higher stress levels both were inhibited. Primary root diameter increased steadily with time in well-watered controls and under water deficit increased transiently before stabilizing at a diameter that was inversely proportional to the deficit. Along with stimulated primary root elongation, moderate water deficit also stimulated the rate of cell production. Thus, A. thaliana responds to water deficit vigorously, which enhances its use as a model to uncover mechanisms underlying plant responses to water deficit.     

Effects of Vendor and Genetic Background on the Composition of the Fecal Microbiota of Inbred Mice

The commensal gut microbiota has been implicated as a determinant in several human diseases and conditions. There is mounting evidence that the gut microbiota of laboratory mice (Mus musculus) similarly modulates the phenotype of mouse models used to study human disease and development. While differing model phenotypes have been reported using mice purchased from different vendors, the composition and uniformity of the fecal microbiota in mice of various genetic backgrounds from different vendors is unclear. Using culture-independent methods and robust statistical analysis, we demonstrate significant differences in the richness and diversity of fecal microbial populations in mice purchased from two large commercial vendors. Moreover, the abundance of many operational taxonomic units, often identified to the species level, as well as several higher taxa, differed in vendor- and strain-dependent manners. Such differences were evident in the fecal microbiota of weanling mice and persisted throughout the study, to twenty-four weeks of age. These data provide the first in-depth analysis of the developmental trajectory of the fecal microbiota in mice from different vendors, and a starting point from which researchers may be able to refine animal models affected by differences in the gut microbiota and thus possibly reduce the number of animals required to perform studies with sufficient statistical power.     

Relationship between force and intracellular [Ca2+] in tetanized mammalian heart muscle

To determine features of the steady state [Ca2+]-tension relationship in intact heart, we measured steady force and intracellular [Ca2+] ([Ca2+]i) in tetanized ferret papillary muscles. [Ca2+]i was estimated from the luminescence emitted by muscles that had been microinjected with aequorin, a Ca2+-sensitive, bioluminescent protein. We found that by raising extracellular [Ca2+] and/or by exposing muscles to the Ca2+ channel agonist Bay K 8644, tension development could be varied from rest to an apparently saturating level, at which increases in [Ca2+]i produced no further rise in force. 95% of maximal Ca2+-activated force was reached at a [Ca2+]i of 0.85 +/- 0.06 microM (mean +/- SEM; n = 7), which suggests that the sensitivity of the myofilaments to [Ca2+]i is far greater than anticipated from studies of skinned heart preparations (or from previous studies using Ca2+-sensitive microelectrodes in intact heart). Our finding that maximal force was reached by approximately 1 microM also allowed us to calculate that the steady state [Ca2+]i-tension relationship, as it might be observed in intact muscle, should be steep (Hill coefficient of greater than 4), which is consistent with the Hill coefficient estimated from the entire [Ca2+]i-tension relationship derived from families of variably activated tetani (6.08 +/- 0.68; n = 7). Finally, with regard to whether steady state measurements can be applied directly toward understanding physiological contractions, we found that the relation between steady force and [Ca2+]i obtained during tetani was steeper than that between peak force and peak [Ca2+]i observed during physiological twitches.     

Maintenance of shoot growth by endogenous ABA: genetic assessment of the involvement of ethylene suppression

Previous work demonstrated that normal levels of endogenous abscisic acid (ABA) are required to maintain shoot growth in well-watered tomato plants independently of effects of hormone status on plant water balance. The results suggested that the impairment of shoot growth in ABA-deficient mutants is at least partly attributable to increased ethylene production. To assess the extent to which ABA maintains shoot growth by ethylene suppression, the growth of ABA-deficient (aba2-1) and ethylene-insensitive (etr1-1) single- and double-mutants of Arabidopsis was examined. To ensure that the results were independent of effects of hormone status on plant water balance, differential relative humidity regimes were used to achieve similar leaf water potentials in all genotypes and treatments. In aba2-1, shoot growth was substantially inhibited and ethylene evolution was doubled compared with the wild type, consistent with the results for tomato. In the aba2-1 etr1-1 double mutant, in which ABA was equally as deficient as in aba2-1 and shoot growth was shown to be insensitive to ethylene, shoot growth was substantially, although incompletely, restored relative to etr1-1. Treatment with ABA resulted in the complete recovery of shoot growth in aba2-1 relative to the wild type, and also significantly increased the growth of aba2-1 etr1-1 such that total leaf area and shoot fresh weight were not significantly lower than in etr1-1. In addition, ABA treatment of aba2-1 etr1-1 restored the wider leaf morphology phenotype exhibited by etr1-1. The results demonstrate that normal levels of endogenous ABA maintain shoot development, particularly leaf expansion, in well-watered Arabidopsis plants, partly by suppressing ethylene synthesis and partly by another mechanism that is independent of ethylene.