Phosphorylation of Chromosomal Proteins in Resting and Wounded Potato Tuber Tissues

Wounding of quiescent white potato tuber tissue enhances chromatin-boundprotein phosphokinase activity, which exhibits two distinctphases during wound-healing. A moderate activation of the enzymesup to 20 hr after injury is followed by a dramatic increasein activity with a peak at 50 hr. This time-course resemblesthat of chromatinbound DNA-dependent RNA polymerase with a peakin activity at about 48 hr after wounding. The kinases phosphorylateendogenous proteins as well as added histones, phosvitin andcasein. The incorporated phosphate is stable under standardassay conditions, indicating the absence of protein phosphatases.Sensitivity of the incorporated phosphate toward trypsin andalkali, but not DNase, RNase, hydroxylamine or succinic acidpoints to seryl- and threonyl-bonds and proteins as acceptormolecules. Kinases from resting tissues are only weakly stimulatedeven by 100 mM MgCl₂, those from wounded tissues exhibit pronouncedMg^$$-optima at 5–10 mM with endogenous proteins, phosvitinand casein and 50 mM MgCl₂ with histones. Wounding also increasesthe sensitivity of the kinases toward p-hydroxymercuribenzoate. Chromatin preparations from both resting and wounded tissuescontain about 40 protein bands after polyacrylamide disc gelelectrophoresis. In vitro phosphorylation of these proteinsin chromatin from quiescent tissues is comparably low and uniform.Wounding induces changes in the protein and phosphorylationpattern with a general enhancement of phosphorylative capacityand preferential phosphorylation of low molecular weight proteins. (Received August 10, 1981; Accepted November 18, 1981)     

Site-directed mutagenesis in the uracil-repair system. Preparation of mutated forms of human alpha 2 interferon

The mutant forms of human IFN-alpha 2 gene are obtained by oligonucleotide-directed mutagenesis with the use of uracil-repair system. To intensify the process the procedure of the uracil-containing DNA template preparation is modified. It was determined that when mutagenesis is performed in the uracil-repair system the yield of the process depends on the mutant DNA-strand in vitro synthesis efficiency. It is shown that the stability of the 5'-end primer-template complex and the level of the endogenic primers elongation are the basis factors, that determine induction mutations.     

Identification of Cell Cycle Dependent Interaction Partners of the Septins by Quantitative Mass Spectrometry

The septins are a conserved family of GTP-binding proteins that, in the baker''s yeast, assemble into a highly ordered array of filaments at the mother bud neck. These filaments undergo significant structural rearrangements during the cell cycle. We aimed at identifying key components that are involved in or regulate the transitions of the septins. By combining cell synchronization and quantitative affinity-purification mass-spectrometry, we performed a screen for specific interaction partners of the septins at three distinct stages of the cell cycle. A total of 83 interaction partners of the septins were assigned. Surprisingly, we detected DNA-interacting/nuclear proteins and proteins involved in ribosome biogenesis and protein synthesis predominantly present in alpha-factor arrested that do not display an assembled septin structure. Furthermore, two distinct sets of regulatory proteins that are specific for cells at S-phase with a stable septin collar or at mitosis with split septin rings were identified.Complementary methods like SPLIFF and immunoprecipitation allowed us to more exactly define the spatial and temporal characteristics of selected hits of the AP-MS screen.     

Chemoreceptor function and metabolism of the carotid body in the rat

Using fluorimetric and polarographic determination of cell respiration, metabolic reactions of the carotid body of rats to adequate stimulation have been revealed. Functional relationships of the dose--effect type, which were found for these reactions, may be approximated by equations which are similar to those describing the unconditioned reflexes from the carotid chemoreceptors. However, reactions of the metabolics systems of the carotid body account for the perception of not all chemical stimuli, but only of acids and some alkaloids. These data reveal the heteregeneity of the carotid chemosensory system not only in the amphibians (which was known earlier), but in mammals as well.     

Histochemical study of reverse reactions in experimental leprosy in armadillos

Morphological and histochemical characteristics of reverse reactions in M. leprae-infected armadillos are reviewed. The reverse reactions develop in generalized lepromatous disease and are characterized by a dramatic decrease in mycobacterial macrophage load and the appearance of lymphocytes, epithelioid and giant multinuclear cells with peripheral nucleus distribution. Histochemical investigation has shown a decrease in the activity of redox enzymes and alpha-naphthyl acetate esterase and an increase in beta-glucuronidase activity. The reverse reactions in the liver are accompanied by hepatic granuloma and hepatocyte destruction.