Translatable RNA Populations Associated with Maintenance of Primary Root Elongation and Inhibition of Mesocotyl Elongation by Abscisic Acid in Maize Seedlings at Low Water Potentials

Previous work indicated that accumulation of abscisic acid (ABA) acts differentially to maintain elongation of the primary root and inhibit elongation of the mesocotyl of maize (Zea mays L.) seedlings at low water potentials ([psi]w). Subsequent results indicated specific locations in the elongation zones where elongation is maintained, inhibited, or unaffected by endogenous ABA at low [psi]w. This information was utilized in this study to identify in vitro translation products of RNA associated with the maintenance or inhibition of elongation in the primary root and mesocotyl, respectively, by endogenous ABA at low [psi]w. The results distinguished products associated specifically with the elongation responses from those nonspecifically associated with ABA accumulation or low [psi]w, as well as normal cell development and maturation. In the primary root, the maintenance of elongation at low [psi]w by ABA was associated with the maintenance of expression of three products that were also expressed during elongation at high [psi]w, the expression of a novel product, and the suppression of two products. In the mesocotyl, the inhibition of elongation by ABA after transplanting to low [psi]w was associated with the induction of a novel translation product. However, the induction of this product, as well as accumulation of ABA and inhibition of elongation, occurred without a decline in tissue water content. The results demonstrate the necessity of examining the association of gene expression with elongation responses to low [psi]w with a high degree of spatial resolution.     

Comparison of Type 2 Inositol 1,4,5-Trisphosphate Receptor Distribution and Subcellular Ca2+ Release Sites that Support Ca2+ Waves in Cultured Astrocytes

Abstract: We have examined the mechanisms that underlie Ca²⁺ wave propagation in cultured cortical astrocytes. Norepinephrine evoked Ca²⁺ waves in astrocytes that began at discrete initiation loci and propagated throughout the cell by regenerative amplification at a number of cellular sites, as shown by very high Ca²⁺ release rates at these regions. We have hypothesized previously that domains displaying elevated Ca²⁺ release kinetics in astrocytes may correspond to sites of high inositol 1,4,5-trisphosphate receptor (InsP₃R) density. To examine this possibility, we compared the distribution pattern of endoplasmic reticulum (ER) and InsP₃Rs with Ca²⁺ release kinetics in subcellular regions during propagation of norepinephrine-evoked waves. 3,3'-Dihexyloxacarbocyanine iodide staining revealed that the ER in astrocytes exists as a meshwork of membranes extending throughout the cells, including fine processes. A specific antibody directed against type 2 InsP₃Rs (InsP₃R2) detected a 260-kDa band in western blotting of astrocyte membranes. Immunocytochemistry using this antibody stained the entire ER system in a punctate, variegated manner. When Ca²⁺ responses and InsP₃R2 immunofluorescence were compared in the same cell, domains of elevated Ca²⁺ response kinetics (high amplitude and rapid rate of rise) showed significant positive correlation with high local intensity of InsP₃R2 staining. It appears, therefore, that specializations in the ER responsible for discrete local Ca²⁺ release sites that support regenerative wave propagation include increased levels of InsP₃R2 expression.     

Biotechnology and agricultural improvement

A range of emerging technologies are expected to play a significant role in agricultural improvement in the next 20 years. Some are only now being explored, but others have already produced significant results. Recent progress in the tissue culture and genetic engineering of crop plants has opened the door to: (1) large scale and rapid propagation of genetically uniform plants from elite materials; (2) the selection of novel and improved varieties using somaclonal variation technology; (3) the development of new hybrids between different cultivars and species by means of protoplast fusion and (4) the use of recombinant DNA to introduce new genetic material into plant cells. It is expected that, by the year 2000, a wide range of crops will be affected by these advances in biotechnology.     

Calcium content and distribution as a function of growth and transformation in the mouse 3T3 cell

Total Ca content and that fraction of Ca sensitive to removal by the chelator ethylene glycol-bis(β-aminoethyl ether)N,N,N',N'-tetraacetate (EGTA) have been investigated in the mouse 3T3 cell as a function of growth stage, transformation with SV40 virus, and serum levels of the media. Cells were allowed to grow through several doublings in media containing (45)Ca. The cellular content of (45)Ca was used to access total cell Ca. That fraction of (45)Ca removed by EGTA was presumed to represent primarily surface-localized Ca. The data are expressed on a per cell volume basis to compensate for size differences as a function of growth stage and transformation. During exponential growth phase, the 3T3 cell contains 525pmol Ca/μl cell volume. Of this, approx. 457 pmol/μl is not removable by EGTA and, presumably, is cytoplasmically located. This value is in close agreement with previous studies on the HeLa cell (470 pmol Ca/μl cell water after the removal of the surface Ca). The low level of EGTA- removable Ca present in the 3T3 cell during early exponential growth (68 pmol Ca/μl cell volume) increases progressively with increasing cell density, and upon quiescence it is sevenfold greater. In contrast, SV40- transformed 3T3 cells growing exponentially possess total levels of Ca which are approximately two-thirds the levels of the normal 3T3 cell. However, their EGTA-sensitive Ca is not significantly different from that of exponentially growing, normal 3T3 cells. As the transformed cells continue to grow at high density, their total ca and their sensitivity to EGTA do not change, in contrast to the normal 3T3 cell. Thus, an increase in Ca associated with the cell surface appears to be correlated with growth inhibition. This has been investigated further by regulating growth of the normal and transformed cell with alterations in the serum level of the media. In 4 percent calf serum the normal cell is stopped from continued proliferation. Growth stoppage under these conditions is characterized by a nearly fourfold increase in EGTA-removable Ca, similar to the increase observed upon quiescence in depleted 10 percent serum. Similar treatment of the transformed cell does not reduce its growth rate, nor does it significantly alter Ca distribution. However, at 0.5 percent medium serum levels, the SV40 3T3 growth rate is substantially reduced and, under these conditions, EGTA-removable Ca increases twofold.     

Poliovirus aggregates and their survival in water.

Inactivation of aggregated poliovirus by bromine is characterized by a continuously decreasing reaction rate. Poliovirus released from infected cells in these experiments by alternate freezing and thawing in water without electrolytes has always been aggregated. The aggregates persist even on 7,000-fold dilution in ion-free water. Virus similarly released into phosphate-buffered saline solution may be well dispersed, but it aggregates when sedimented into a salt-free sucrose gradient or when it is diluted as little as 10-fold in water. Large one-step dilutions of dispersed virus in water remain dispersed. Aggregated virus was not dispersed by one-step dilution (7,000-fold) in distilled or untreated lake water but was dispersed if phosphate-buffered saline or clarified secondary sewage plant effluent was used as diluent. Dispersed virus aggregates at all dilutions in alum-treated, finished water from the city filter plant. This may be the result of complex formation with insoluble material rather than virion-virion aggregation. A simple procedure is described for rendering a very dilute suspension of mixed virion aggregates into a three-part spectrum of sizes.