Localization of Two Functions of the Phosphoribosyl Anthranilate Transferase of Escherichia coli to Distinct Regions of the Polypeptide Chain

The trpD gene specifies a polypeptide which has both glutamine amidotransferase and phosphoribosyl anthranilate (PRA) transferase activities. Deletions fusing segments of trpD to the gene preceding it in the operon, trpE, were selected in strains carrying various trpD point mutations. The selection procedure required both that a deletion enter trpE and that it restore the PRA transferase activity which the parent trpD point mutant lacked. Deletion mutants were found which had PRA transferase activity although the first third of trpD was deleted. The existence of the mutants proves that a terminal segment of trpD is sufficient to specify a polypeptide having PRA transferase activity. The location of the deletion end points on the genetic map of trpD defines the extent of the trpD segment required for PRA transferase activity. This segment did not overlap the initial region of trpD required to specify the glutamine amidotransferase function of the trpD polypeptide. These results support the hypothesis (M. Grieshaber and R. Bauerle, 1972; H. Zalkin and L. H. Hwang, 1971) that the bifunctional trpD polypeptide might have evolved by fusion of a gene specifying a glutamine amidotransferase with a gene directing PRA transferase synthesis.     

HISTOLOGICAL CHANGES DURING THE DEVELOPMENT OF THE BOVINE NUCHAL LIGAMENT

The histological changes occurring during the development of the bovine nuchal ligament have been observed in sections of formalin-fixed material from 21 animals ranging in age from 110 days of gestation to 10 yr. The elastic fibers which constitute the bulk of the adult ligament were initially few in number. During fetal development, the fibers showed a rapid increase both in number and in their stainability with the usual elastic stains. The average diameter of these elastic fibers increased only slowly until the last uterine month, at which time it began to increase very rapidly. This rapid rate of increase continued through the first 6 postnatal months, after which the rate of increase slowed markedly. However, the fiber diameter continued to rise steadily throughout the period of the study. During the fetal stage of development, the fibroblastic cells of the ligament exhibited unusual nuclear appearances which distinguish them from other fibroblasts. These consisted of marked clumping of the chromatin and an associated nuclear vacuolation or vesiculation. While these changes seem likely to be artefacts of fixation, their temporal correlation with elastin deposition and their demonstration in other tissue cells engaged in elastin production suggest that the factors responsible for these appearances may be related to elastin synthesis.     

Studies onAzotobacter species in soil

Summary Methods for countingAzotobacter species in soil have been examined. The highest counts were obtained from soil suspensions shaken in sterile distilled water containing 10-g glass beads and plated on to glucose agar. Mannitol has been rejected as a suitable substrate in agar media because it gives lower counts of Azotobacter than glucose, an effect which is further enhanced by drying the agar plates. A clear medium free from precipitated phosphate and CaCO₃ is recommended for the agar-plate method; the Azotobacter count is affected by the phosphate concentration.The agar-plate and dilution-tube methods were compared; the latter is less accurate but more convenient when many soil samples have to be examined.     

Regulation of Mitosis in Living Cells. I. Mitosis under Controlled Conditions

A quantitative method has been devised to study mitosis in vitro by phase contrast and polarization microscopy. Mitosis in cell-wall-free endosperm cells of Haemanthus kathrinar Baker (the African blood lily) has been divided into 18 arbitrary stages or events. The time course for the various stapes, as well as the percentage of cells that proceed from one stage to another during a four hour observation period, are presented. Cells that were in prophase when selected for study proceeded from nuclear membrane breakdown to melaphase in 60 minutes and remained in melaphase for 30 minutes. Only 13 minutes was required to proceed from onset of anaphase to mid-anaphasc. Mid-anaphase provides a clear and precise baseline for determining the time required for succeeding stages to appear. The cell plate made its appearance 40 minutes after mid-anaphase and was completely formed 20 minutes later. The nuclear membranes also became evident at this latter time and nucleoli were visible 30 minutes later. Thus, the average time for a cell observed initially in prophase to proceed from nuclear membrane breakdown to formation of two daughter cells was just over three hours. A high percentage of cells that were in late prophase or later stages of mitosis at the time of initial observation completed mitosis during the observation period. The effect of the length of time a cell is subjected to experimental conditions upon its subsequent behaviour is assessed. These results form the basis for future studies of the effects of chemicals, particularly herbicides, upon cells in mitosis as observed in vitro by phase contrast and polarization microscopy.