Influence of Sulfur Nutrition on Developmental Patterns of Some Major Pea Seed Proteins and Their mRNAs

In addition to the marked reduction in legumin synthesis and legumin mRNA levels reported earlier (Chandler, Higgins, Randall, Spencer 1983 Plant Physiol 71: 47-54), pulse labeling of S-deficient Pisum sativum L. seeds showed that a high relative level of total vicilin (vicilin plus convicilin) synthesis was maintained throughout the entire phase of protein accumulation, whereas in nondeficient seeds vicilin synthesis is largely confined to the first half of this phase. Fractionation of pulse-labeled proteins on Na-dodecylsulfate-polyacrylamide gels showed that the synthesis of the M_r 50,000 family of vicilin polypeptides was increased and greatly extended in S-deficient seeds whereas that of convicilin was slightly reduced. Other changes apparent from pulse-labeling experiments include a depression, to different degrees, in the synthesis of three major albumin polypeptides.

The level of the mRNAs for seven major seed proteins was followed throughout development of control and sulfur-deficient seeds. In all cases, the changes in each mRNA closely reflected the pattern of synthesis of its corresponding polypeptide seen by pulse labeling. S-deficient seeds showed an elevated level of M_r 50,000 vicilin mRNA which remained high throughout seed formation, whereas legumin mRNA levels were greatly reduced at all stages of development.

When S-deficient plants were given an adequate supply of sulfate midway through seed development, there was a shift toward the protein synthesis profile characteristic of healthy plants. The synthesis of legumin and two albumins rapidly increased and the synthesis of M_r 50,000 vicilin declined more slowly. Similar responses were seen in detached, S-deficient seeds supplied directly with adequate sulfate.

     

Combined effect of growth medium,age of cells and phase of sporulation on heat resistance and recovery of Hansenula anomala

Effects of two growth media, age of cells and phase of sporulation on heat resistance of Hansenula anomala were determined. Cells were grown on two solid media, McClary's acetate and V8 juice agars, at 21 ° C for 16 days. Heat resistance of cells was determined in 0.06 M potassium phosphate buffer at 48 ° C. Heat-stressed cells were plated on four recovery media: yeast extract-malt extract-peptone-glucose (YMPG), pH 7.0; YMPG, pH 3.5; YMPG containing 6% NaCl, pH 7.0; and YMPG containing 20% sucrose, pH 7.0. The composition of sporulation medium influenced the extent of sporulation and the relative heat resistance of sporulating cells. One-day-old cells were the most sensitive to heat. The heat resistance of cells was generally increased as the incubation time was extended to 16 days. Heat treatment caused a greater increase in sensitivity to NaCl than to sucrose or acid pH in recovery media. Young cells were more sensitive to NaCl than were older cells.     

Nitrogenase — hydrogenase relationships in Rhizobium japonicum

The conditions necessary for coordinate derepression of nitrogenase and O₂-dependent hydrogenase activities in free-living cultures of Rhizobium japonicum were studied. Carbon sources were screened for their ability to support nitrogenase, and then hydrogenase activities. There was a positive correlation between the level of nitrogenase and corresponding hydrogenase activities among the various carbon substrates. The carbon substrate -ketoglutarate was able to support the highest levels of both nitrogenase and hydrogenase activities. When cells were incubated in -ketoglutarate-containing medium, without added H₂ but in the presence of acetylene (to block H₂ evolution from nitrogenase) significant hydrogenase activity was still observed. Complete inhibition of nitrogenase-dependent H₂ evolution by acetylene was verified by the use of a Hup^- mutant. Hydrogen is therefore not required to induce hydrogenase. The presence of 10% acetylene inhibited derepression of hydrogenase. Constitutive (Hup^c) mutants were isolated which contained up to 9 times the level of hydrogenase acitivity than the wild type in nitrogenase induction medium. These mutants did not have greater nitrogenase activities than the wild type.This is contribution number 1254 from the Department of Biology and the McCollum-Pratt Institute Abbreviations: -Ketoglutarate-containing medium (LOKG) and pre-adaptation medium (SRM) as described in Materials and methods     

Morphometric integration in the rat (Rattus sp.) scapula

Morphometric integration was analysed in 19 anatomical measures taken on the scapula and humerus in a population of 519 rats. As hypothesized, genetic integration was the highest, the average phenotypic genetic, and environmental correlations being 0·53, 0·67 and 0·42, and the index of integration 0·56, 0·69 and 0·48. Phenotypic and genetic correlation matrices were most similar (correlation =+0.79), genetic and environmental matrices least similar (correlation =+0.49). The first unrotated vector produced from principal components analysis explained a high percentage of the total variation (from 50% in the environmental to 70% in the genetic solution), and was highly heritable in all cases. Rotated vectors defined two length, one width, and one height grouping in the phenotypic solution, these being explained largely in terms of muscle assemblages. The four vectors produced in the genetic solution were similar to those from the phenotypic ones, but were more functionally interpretable. The five vectors produced from the environmental correlations paralleled those from the phenotypic correlations with regard to the length, but not the width measures. The general concordance among appropriate vectors from all three solutions was reasonably high. Twelve of the 13 vectors, as well as several hypothetical ones. exhibited moderate to high heritabilities.     

Effects of testosterone on messenger ribonucleic acid and protein synthesis in rat seminal vesicle

In a previous report [Higgins et al. (1976) Biochem. J. 158, 271–282] we described the effects of alterations in androgen status on the synthesis of two basic secretory proteins of the rat seminal vesicle. In the present paper we examine the effects of testosterone on the activity of mRNA in the seminal vesicle. Total cellular poly(A)-rich RNA was isolated and translated in a cell-free system prepared from wheat germ. Translation products were separated on denaturing polyacrylamide gels and the protein bands corresponding to the two basic secretory proteins were identified immunologically. Incorporation of radioactive methionine into these bands was taken as a measure of the individual mRNA activities. Total mRNA activity was estimated by radioactivity in total acid-precipitable material. The results show that 1 to 2 weeks after castration the activities of mRNA molecules for the basic secretory proteins were decreased 10–20-fold on a tissue basis. Testosterone given in vivo rapidly and substantially restores mRNA activity to normal. Since these changes correlate closely with variations in the rates of synthesis of the secretory proteins in whole cells it suggests that androgenic steroids control protein synthesis chiefly via mRNA availability. In this respect their action resembles those of other steroid hormones acting in other systems. However, these effects of testosterone on the mRNA molecules for the major secretory proteins could not be distinguished from those on total mRNA. Thus the proportion of the total mRNA population accounted for by the two specific mRNA molecules showed less than a 2-fold variation with androgen status. Similarly the two secretory proteins always accounted for 25–33% of general protein synthesis. This is in sharp contrast with the markedly differential effects of other steroid hormones controlling synthesis of major proteins in other well-studied systems. We interpret our results as indicating that testosterone regulates the mRNA population of the seminal vesicle as a whole.     

Coordinate regulation of protein synthesis and messenger RNA content during growth arrest of suspension Chinese hamster ovary cells

We have found Chinese Hamster Ovary cells, cultured in suspension, are subject to growth control by serum. When suspended in medium containing 0.5% serum the cells become reversibly arrested in the beginning of the G₁ phase of the cell cycle and can be maintained in this viable, nonproliferating state for several days. This system was used to examine the regulation of protein synthesis with growth rate. In particular, the experiments addressed the question whether mRNA content is the principal controlling factor determining the rate of protein synthesis. The rate of leucine incorporation in resting cells in low serum is 2- to 2.5-fold lower than that of cells growing in 10% serum. The steady-state number of cytoplasmic poly A (+) RNA molecules shows a proportional decrease, consistent with it being a determining factor controlling the rate of protein synthesis. Furthermore, the rate of production of poly A (+) and poly A (?) RNA appears to be regulated coordinately. Regulation of the rate of initiation of translation would result in fewer ribosomes bound per active message and/or a lower proportion of total mRNA's being active. Our measurements indicate that the fraction of cytoplasmic poly A (+) mRNA in polyribosomes and the relative degree of loading of each active poly A(+) mRNA with ribosomes is the same in resting and growing cells. Thus these cells resemble 3T6 and translational control does not appear to be an important part of the change in protein synthetic rate with the state of growth.     

Comparison of various ultraviolet sources for fluorescent detection of ethidium bromide-DNA complexes in polyacrylamide gels

Ultraviolet sources with output wavelengths of 254, 300, and 366 nm were compared for detection of ethidium bromide-DNA complexes in acrylamide gels. The 254- and 300-nm sources were both much more sensitive than the 366-nm source. The 254-nm source produced a great deal of photodamage, photonicking and photodimerization, and photobleaching, while the longer wavelength sources cause little damage or bleaching. The 300-nm source is clearly the most suitable source, providing high sensitivity and a relatively low amount of photodamage and photobleaching.     

Temporal patterning of feeding by hummingbirds

For five species of hummingbirds in the laboratory, time between meals was related to energy intake on the first meal and rate of energy expenditure between meals. Field observations gave similar results. Average meal sizes were similar at one intake rate independent of food caloric density; females averaged longer bouts than males. When rate of intake was approximately halved, meal duration approximately doubled and volume intake remained similar. We postulate that feeding is initiated when crop contents reach a lower threshold and that feeding is terminated after ingestion of an optimal volume determined by the added weight of the meal.     

Ascorbic acid uptake in guinea pig intestinal mucosa

Cellular accumulation of ascorbic acid was investigated in vitro in distal intestinal mucosa of guinea pig. With ¹⁴C-ascorbic acid present at 8 μM/L in the bathing media, tissue/media (T/M) concentration ratios of at least 5 were routinely achieved. Recently absorbed ascorbic acid appeared to be free in solution in the cellular fluid in that it diffused from tissue exposed to poisons with a disappearance half-time of approximately 10 minutes. Ascorbic acid uptake was highly dependent on the presence of sodium in the bathing media; total Tris substitution resulted in a 97% decrease in uptake. Also, metabolically depleted tissue did not accumulate ascorbic acid against a concentration gradient. Uptake of ¹⁴C-ascorbic acid from a bathing solution concentration of 8 μM/L was reduced 67% in the presence of 0.8 mM/L nonlabeled ascorbic acid. Recently absorbed ¹⁴C-ascorbic acid moved more rapidly back into the lumen when the luminal solution contained nonlabeled ascorbic acid (5 mM) than when it contained mannitol (5mM). This demonstration of counter transport substantiates a carrier mechanism in the brush border.