Regulation of sperm flagellar movement by protein phosphorylation and dephosphorylation

Flagellar motility of Triton models of sea urchin spermatozoa was reactivated by cyclic AMP-dependent protein kinase and a protein factor, termed motility activator, both of which were prepared from the detergent-extract of sea urchin spermatozoa. It was shown that phosphorylation of the motility activator by the protein kinase is necessary for the reactivation of flagellar motility [Ishiguro et al, J. Cell Biol. 92:777-782, 1982; Murofushi et al, in "Biological Functions of Microtubules and Related Structures," Academic Press, 1982]. Reactivating factor was also detected in a KCl-extract of the axoneme fraction devoid of the detergent-extractable materials. The activity of this factor was also cyclic AMP- and protein kinase-dependent. Furthermore, when freshly prepared Triton models were treated with phosphoprotein phosphatase prepared from bovine cardiac muscle, the flagellar motility was drastically suppressed. This inhibition of the motility was partially recovered by the addition of cyclic AMP and protein kinase to the phosphatase-treated models.     

Modulatory Action of Taurine on the Release of GABA in Cerebellar Slices of the Guinea Pig

Abstract: For the purpose of demonstrating the action of taurine as a neuromodulator in addition to its suggested neurotransmitter function, the effects of taurine and muscimol on the depolarization-induced Ca-dependent release of [³H]γ-aminobutyric acid (pH]GABA) and l -[³H]glutamate in cerebellar slices from guinea pigs were investigated. The release of [³H]GABA was found to be greatly decreased by a GABA agonist, muscimol, and by taurine, but not by glycine. The release of l -[³H]glutamate was little affected by taurine. The release of [³H]GABA was enhanced by bicuculline and strychnine, but not by picrotoxin, and the suppressive action of muscimol on the GABA release was antagonized by bicuculline, picrotoxin, and strychnine, suggesting the possible existence of presynaptic autoreceptors for GABA in the cerebellum. The suppressive action of taurine on the release of [³H]GABA, on the other hand, was blocked only by bicuculline. These results suggest that taurine reduced the release of [³H]GABA from cerebellar slices by acting on the GABA autoreceptors or, more likely, on other types of receptors that are sensitive to bicuculline. As a possible mechanism for this modulatory action of taurine, the blockade by this amino acid of the influx of Ca²⁺ into cerebellar tissues was tentatively suggested.     

In vivo Studies on the Occurrence of Stored mRNA in Embryonic Axes of Vigna unguiculata Seeds

-Amanitin and cordycepin at various concentrations were testedfor their inhibitory effect on the fresh weight increase ofVigna unguiculata embryonic axes after the onset of imbibitionand on the incorporation rate of ³H-labeled leucine into proteinin axes of the 36–38 h stage. -Amanitin at 0.5–5µ/Kg/ml clearly exerted an inhibitory effect on both thefresh weight increase and the protein synthesis. This drug at1 µg/ml, however, showed no significant effect on theprotein synthesis at an early stage of imbibition (4 h), whereascycloheximide was a very potent inhibitor. By experiments inwhich ‘dry’ axes were allowed to imbibe 3H-lebeledadenosine solution for 4 and 12 h in the presence of -amanitin,it was found that poly A⁺RNA was newly synthesized to some extentin axes as early as 4 h after the onset of imbibition and thatthe drug effectively inhibited the poly A⁺RNA synthesis. Theresults may indicate the occurrence of stored mRNA in embryonicaxes of V. unguiculata seeds. (Received June 11, 1983; Accepted August 16, 1983)     

Differential staining of insect neurons with nickel and cobalt

Pairs of neurons were differentially stained by intracellular injection of the divalent cations, nickel and cobalt. Pairs of neuronal tracts could be similarly labelled following immersion of their axon bundles in the markers described. Use of nickel and a 4 : 6 mixture of cobalt and nickel according to the protocols presented here revealed the morphology of cricket neuron processes as thin as 1 μm in diameter and yielded a favourable contrast between the cells marked, as well as with respect to the background of unstained tissue.     

Replication of Deoxyribonucleic Acid in Escherichia coli C Mutants Temperature Sensitive in the Initiation of Chromosome Replication

An Escherichia coli HF4704S mutant temperature sensitive in deoxyribonucleic acid (DNA) synthesis and different from any previously characterized mutant was isolated. The mutated gene in this strain was designated dnaH. The mutant could grow normally at 27 C but not at 43 C, and DNA synthesis continued for an hour at a decreasing rate and then ceased. After temperature shift-up, the increased amount of DNA was 40 to 50%. When the culture was incubated at 43 C for 70 min and then transferred to 27 C, DNA synthesis resumed after about 50 min, initiating synchronously at a fixed region on the bacterial chromosome. The initiation step in DNA replication sensitive to 30 mug of chloramphenicol per ml occurs synchronously before the resumption of DNA replication after the temperature shift-down, being completed about 30 min before the start of DNA replication. When the cells incubated at 27 C in the presence of 30 mug of chloramphenicol per ml after the temperature shift-down to 27 C were transferred to 43 C with simultaneous removal of the antibiotic, no resumption of DNA replication was observed. When the culture was returned to 43 C after being released from high-temperature inhibition at 30 min before the start of DNA replication, no recovery replication was observed; whereas at 20 min, the recovery of replication was observed. These results indicated that HF4704S was temperature sensitive in the initiation of DNA replication. Analysis of HF4704S, by an interrupted conjugation experiment, indicated that gene dnaH was located at about 64 min on the E. coli C linkage map. In E. coli S1814 (a K-12 derivative), which was a dnaH(ts) transductant from HF4704S (C strain) with phage P1, the mutated gene (dnaH) was demonstrated to be closely linked to the thyA marker by conjugation and P1 transduction experiments and to be distinct from genes dnaA through dnaG.     

Rapid isolation of nucleoli from detergent purified nuclei of various tumor and tissue culture cells

A rapid isolation procedure of nucleoli from detergent purified nuclei of some tumor and tissue culture cells is described. The procedure makes use of a non-ionic detergent, Nonidet P40 and sodium deoxycholate to purify nuclei followed by the addition of Ca²⁺ or Mg²⁺ to strengthen the nucleoli against sonication. Enzymatically active (with respect to nucleolar RNA polymerase) nucleoli containing undegraded nucleolar RNAs may be isolated from a mouse hepatoma MH134, Ehrlich ascites tumor, HeLa cells, L cells and C3H2K cells with this procedure.