Clonal variation of Populus tremuloides responses to diurnal drought stress.

We have developed an automated microprocessor controlled system for subjecting hydroponically grown plants to drought. Pumps and valves were used to move nutrient solutions into and out of a system of culture vessels in a growth chamber to provide periods of drought. Drought conditions were obtained by exposing the roots of hydroponically grown clones of aspen, Populus tremuloides Michx., to air in culture vessels temporarily emptied of nutrient medium. Over a 3-week period, the daily duration of drought was increased from 0 to 6 h. During this period, the plants became increasingly tolerant to drought, as shown by a decreasing propensity to wilt. All three clones sustained diurnal drought periods of 6 h for up to 5 weeks without detectable deterioration of health. Typical drought stress symptoms were observed including inhibition of growth, increased tissue amino acid content, and decreased water, solute, and turgor potentials in young leaves. In all clones, control plants had leaf water potentials between -1.0 and -1.6 MPa, whereas leaf water potentials of drought-treated plants were significantly lower, ranging from -1.7 to -3.0 MPa. Only one of the clones showed a significant decrease in leaf solute potential in response to drought. The decrease in leaf solute potential paralleled the decrease in water potential resulting in no significant difference in turgor potential. The other two clones had nonsignificant decreases to more negative leaf solute potentials under drought conditions resulting in significantly lowered turgor potentials. Leaf water potentials, solute potentials, and turgor potentials of the drought-treated plants returned to control values within two hours after rewatering. The growth inhibitions observed could not have been the consequence of loss of turgor. These results demonstrate genetic differences among aspen clones in water relations responses to drought.

Abscisic Aldehyde Is an Intermediate in the Enzymatic Conversion of Xanthoxin to Abscisic Acid in Phaseolus vulgaris L. Leaves.

The enzymatic conversion of xanthoxin to abscisic acid by cell-free extracts of Phaseolus vulgaris L. leaves has been found to be a two-step reaction catalyzed by two different enzymes. Xanthoxin was first converted to abscisic aldehyde followed by conversion of the latter to abscisic acid. The enzyme activity catalyzing the synthesis of abscisic aldehyde from xanthoxin (xanthoxin oxidase) was present in cell-free leaf extracts from both wild type and the abscisic acid-deficient molybdopterin cofactor mutant, Az34 (nar2a) of Hordeum vulgare L. However, the enzyme activity catalyzing the synthesis of abscisic acid from abscisic aldehyde (abscisic aldehyde oxidase) was present only in extracts of the wild type and no activity could be detected in either turgid or water stressed leaf extracts of the Az34 mutant. Furthermore, the wilty tomato mutants, sitiens and flacca, which do not accumulate abscisic acid in response to water stress, have been shown to lack abscisic aldehyde oxidase activity. When this enzyme fraction was isolated from leaf extracts of P. vulgaris L. and added to extracts prepared from sitiens and flacca, xanthoxin was converted to abscisic acid. Abscisic aldehyde oxidase has been purified about 145-fold from P. vulgaris L. leaves. It exhibited optimum catalytic activity at pH 7.25 in potassium phosphate buffer.

Hearing loss.

Defective hearing is the most prevalent chronic health problem in the United States, making it a primary concern of the physician. Failure to diagnose, misdiagnose, and delay of diagnosis of hearing loss are common errors that have serious implications. However, they can be avoided. If children exhibiting the symptoms of delayed language, articulation defects, and academic problems (especially with reading or a medical history of certain diseases) are referred for audiological testing, most cases of hearing impairment will be promptly detected. With adults the implications of hearing loss are different, both medically and psychosocially. After 30 years of age the prevalence of hearing defects increases rapidly. Some major causes are noise, otosclerosis, otitis media, and presbycusis. Surgery and amplification are the principle treatments. Management of nonmedical aspects should involve programs such as the Division of Vocational Rehabilitation, state schools for the deaf, and sign language classes.

Resistance of selected saprobic and zoopathogenic fungi to cycloheximide.

Spore germination was used as an assay to measure the sensitivities of selected fungi (Achlya bisexualis, Cladosporium sp., Trichophyton mentagrophytes and Microsporum gypseum) to cyclohexamide and to determine their abilities to adapt to the drug. Two patterns of response were noted. The saprobes, A. bisexualis and Cladosporium sp., demonstrated acquired resistance. Spores from hyphae previously exposed to cycloheximide either germinated in the presence of concentrations of the drug that completely inhibited spores from unexposed hyphae (Achlya), or germinates with a shorter lag and to a greater extent in the presence of the antibiotic than did spores from unexposed hyphae (Cladosporium). Hyphae of Achlya adapted at concentrations of cycloheximide in which spores did not germinate and hyphae of Cladosporium adapted more rapidly than spores. Achlya adapted to only 12 muM-cycloheximide whereas Cladosporium acquired resistance to 18 mM-cycloheximide. These fungi lost this acquired resistance after a single transfer to media lacking cycloheximide. The zoopathogens, T. mentagrophytes and M. gypseum, had a contrasting response, indicating constitutive resistance. Conidia from unexposed hyphae showed 90 to 100% germination on media containing up to 18 mM-cycloheximide; prior exposure to the drug did not affect their response.

Inhibition of DNA synthesis by cycloheximide and blasticidin-S is independent of their effect on protein synthesis.

The effects of cycloheximide and related glutarimide antibiotics on DNA synthesis in Achlya bisexualis Coker and A. Couch were compared to those of other protein synthesis inhibitors, puromycin, p-fluorophenylalanine and blasticidin-S. The inhibitors had no significant effects on intrahyphal pool sizes of dTTP, dCTP, ATP, UTP and CTP, nor on the specific activity of the dTTP pool labelled by [3H] thymidine. DNA was the sole acid-insoluble product of [3H]-thymidine incorporation. Cycloheximide, isocycloheximide, streptimidone and blasticidin-S inhibited DNA synthesis rapidly and completely and anhydrocycloheximide was less effective. Cycloheximide acetate, puromycin and p-fluorophenylalanine did not inhibit DNA synthesis. It is concluded that the effects of the several glutarimide antibiotics and of blasticidin-S on DNA synthesis were independent of their effects on protein synthesis.

Differential effects of analogs of cycloheximide on protein and RNA synthesis in Achlya.

Analogs of the glutarimide antibiotic cycloheximide were tested for their effect on growth and incorporation of proline and uridine into acid-insoluble material in Achlya bisexualis. Each of the compounds tested had reduced antibiotic activity as compared to cycloheximide. The effects of the antibiotics on protein and RNA synthesis were varied. While cycloheximide inhibited both protein and RNA synthesis immediately, two of the analogs inhibited proline incorporation without effect on uridine incorporation, while three, each representing a modification of the hydroxyl of cycloheximide, stimulated uridine incorporation and either had no effect on or inhibited protein synthesis. These results indicate that the control of RNA synthesis by protein synthesis in Achlya can be released by glutarimide antibiotics.

Protein synthesis during the differentiation of sporangia in the water mold Achlya.

During the synchronous differentiation of sporangia in the absence of added nutrients, the water mold Achlya bisexualis (Coker and Couch) actively synthesized protein. Inhibition of protein synthesis at any time during the sporulation process completely inhibited further differentiation. Large changes in the rate of radioactive amino acid uptake resulted in changes in the specific activity of the cellular amino acid pool. The rate of protein synthesis was calculated from the amino acid pool specific activity and the incorporation of isotope into protein. During the 1st h after induction of the sporulation process, the rate of protein synthesis increased to two times the initial value. The amino acid precursors for this synthesis were supplied by the degradation of preexisting protein. Proteolytic enzyme activity assayed in vitro increased in proportion to the in vivo rates of protein synthesis and degradation. Differentiation was accompanied by a slight decline in dry weight of the mycelium as well as by a decrease in the protein content, whereas the relative size of the amino acid pools remained constant.

Specific inhibitors of the three RNA polymerases from the aquatic fungus Blastocladiella emersonii.

Specific inhibitors of each of the three RNA polymerases of Blastocladiella emersonii have been found. Cycloheximide specifically inhibited the in vitro activity of the DEAE-fraction I enzyme, alpha-amanitin specifically inhibited the DEAE-fraction II enzyme, and rifampicin specifically inhibited the fraction III enzyme. DNA stimulation and dependency on the four riboside triphosphates were shown to be characteristic of each of the three fractions. Optimum concentrations of magnesium ions required were shown to differ among the three fractions and to be somewhat higher than optimum concentrations of manganese ions. The effect of pH on activity was essentially identical for each of the three fractions. Kinetic experiments and nuclease assays indicated the presence of some interfering substances in the partially purified RNA polymerase fractions.

Cytochrome Oxidase Activity in Blastocladiella emersonii.

Studies of cytochrome oxidase in isolated mitochondria of Blastocladiella emersonii Cant. and Hyatt show that the enzyme was present in zoospores and throughout the development of ordinary colorless sporangia and of resistant sporangia. The enzyme activity was present in KCl, NaCl, NH(4)Cl, and KHCO(3) induced resistant sporangia, and was shown to be as active or more active than the enzyme found in ordinary colorless sporangia and zoospores. Interfering substances causing difficulties in the measurement of cytochrome oxidase activity were found in whole cell homogenates of KHCO(3) grown resistant sporangia, but not in KCl, NaCl, or NH(4)Cl grown thalli. These substances could be removed by dialysis or by sedimentation of the mitochondria.

Ribonucleic acid synthesis during the differentiation of sporangia in the water mold Achlya.

The role of ribonucleic acid (RNA) synthesis in the development of sporangia in the saprolegniaceous mold Achlya was studied. Methods were developed for growing and treating large populations of mycelia so that the hyphal tips would differentiate into sporangia with considerable synchrony. Under the starvation conditions imposed for the differentiation of sporangia, net RNA, deoxyribonucleic acid (DNA), and protein synthesis ceased. However, incorporation of radioactive precursors into RNA continued at a high rate throughout the period of differentiation, showing that the enzymatic mechanism for RNA synthesis was still in an active state. Actinomycin D inhibited the differentiation of sporangia and the incorporation of labeled precursors into RNA. The level of actinomycin used did not inhibit the normal outgrowth and branching of the mycelia that occurred during differentiation. Thus, DNA-dependent RNA synthesis was required for the differentiation of sporangia. Sucrose gradient analysis of newly synthesized RNA showed that only the ribosomal and soluble fractions of RNA were labeled during vegetative growth. During the differentiation of sporangia, ribosomal and soluble RNA fractions were also labeled, and, in addition, a heterodisperse fraction of labeled RNA which was heavier than ribosomal RNA appeared; this fraction was not evident in the newly synthesized RNA from vegetative mycelia.

Effect of Electrolytes on Differentiation in Achlya sp.

Some factors controlling the differentiation of sporangia and gemmae in an isolate of Achlya sp. were investigated by replacement culture technique. Factors in the replacement medium which affect differentiation were as follows: A) Calcium is required for the differentiation of both sporangia and gemmae. B) Calcium cannot be replaced by magnesium, potassium or sodium. C) Gemmae are induced when the replacement medium contains salt at 40 to 80 mm. D) The salt induction of gemmae is nonspecific, sodium, potassium, magnesium and calcium chlorides are equally effective.