Occurrence of rye (Secale cereale) 350-family DNA sequences inAgropyron and otherTriticeae

Evidence is presented that in the R and P genomes (Secale cereale andAgropyron cristatum, respectively) of theTriticeae there exist closely related 350-family DNA sequences in the terminal heterochromatin. This observation is compared to the relationships between these two genomes derived from a comparison of theNor and5 S DNA loci as well as the available data on morphological characters, chromosome pairing, and isozyme studies. It is concluded that the R and P genomes are not closely related and that the common presence of very similar 350-family DNA sequences reflects the parallel amplification of this family of DNA sequences.     

Frequent duplication and deletion events in the 5S RNA genes and the associated spacer regions of theTriticeae

The 5 S DNA units from 15 grasses in theTriticeae were analysed at the DNA sequence level. Four units carried duplications near the 3-end of the 5 S RNA gene with 3 of the duplications centred on the same base pairs as a duplication previously reported byGerlach & Dyer. The fourth duplication was located 3 downstream from the gene, in the spacer region. Apparent deletions were very frequent when units of the different grasses were compared and it was clear that these deletions did not extend into a 75 bp spacer region upstream from the 5 S RNA gene. This 75 bp region also tended to be more conserved between the grasses as compared to the high level of sequence change in the rest of the spacer region. — Phenetic relationships were established between the grasses using the sequence data. The relationships were generally consistent with the data from other parameters and, in addition, showed that two Australian grasses were closely related to the other Northern hemisphere genera examined. The data concerning the Australian grasses is discussed in relation to the isolated nature of Australia.     

Proteolytic specificity of chicken cathepsin L on bovineβ-casein

The proteolytic specificity of chicken cathepsin L was studied using bovine -casein as substrate. The peptide mixtures obtained after various times of hydrolysis were separated by RP-HPLC and ten peptides were identified. Chicken cathepsin L accepts proline residues in all positions except P ₁ . Looking at the amino acid residues on the amino side of the scissile bond we found three times the Tyr-Pro pair at P ₁ –P ₂ positions and that the S ₁ subsite can interact with modified amino acids such as phosphoserine.Abbreviations RP-HPLC reverse phase high performance liquid chromatography - NMec N-methyl coumarylamide - TEA triethylamine - TFA trifluoroacetic acid     

Immobilization of enzymes on alumina by means of pyridoxal 5′-phosphate

A number of proteases have been immobilized on alumina in a two-step procedure: the first step converted them into semisynthetic phosphoproteins which, in the second step, spontaneously bonded to alumina through their phosphate function. The immobilized enzymes thus obtained showed the physical properties typical of the inorganic carrier and a high activity on low molecular weight substrates.     

Effects of auxin and gibberellin on conidial germination and elongation of young hyphae in a cyclic 3′:5′ adenosine monophosphate-dependent protein kinase mutant of Neurospora crassa

The possibility that plant growth regulators may relate to a cyclic 3:5 adenosine monophosphate (cAMP)-dependent protein kinase through the control of cAMP level in the conidial germination process of Neurospora crassa was examined using a cAPM-dependent protein kinase mutant (cpk mutant) which is thought to be cAMP-independent because of defect in the regulatory subunit of cAMP-dependent protein kinase. IAA, 2,4-D and GA₃ promoted conidial germination and elongation of young hyphae in the mutant as well as in the wild-type. The result suggests that the effects of auxin and gibberellin on germination and hyphal elongation are not mediated by cAMP.     

[3H]Imipramine Binding of Protein Nature in Human Platelets: Inhibition by 5-Hydroxytryptamine and 5-Hydroxytryptamine Uptake Inhibitors

Abstract: The nature of [³H]imipramine binding to human platelets was investigated. Desipramine and 5-hydroxytryptamine (5-HT) displaced the same amount of binding and the binding was sensitive to protease treatment. The nature of pharmacological inhibition of [³H]imipramine binding was investigated in saturation experiments. Increases in K d without changes in B _max were noted with the addition of 5-HT, desipramine, norzimeldine, or 5-methoxytryptoline. Reductions in B _max without alterations in K _D were obtained when citalopram or clomipramine was added. It is concluded that the [³H]imipramine binding site in human platelets is of protein nature and that this binding site contains the substrate recognition site for 5-HT uptake. In addition, [³H]imipramine and other 5-HT uptake inhibitors have bonds to other parts of the 5-HT uptake carrier or to the surrounding lipid membrane. This additional binding outside the substrate recognition site is not one single site but most likely represents sites that are specific for the chemical structure of each uptake inhibitor, respectively.     

Stereoselective Nicotine-Induced Release of Dopamine from Striatal Synaptosomes: Concentration Dependence and Repetitive Stimulation

Using a sensitive perfusion system we have studied the nicotine-induced release of [3H]dopamine ([( 3H]DA) from striatal synaptosomes. Nicotine-evoked release was concentration dependent with an EC50 of 3.8 microM. The response to 1 microM nicotine was comparable to that to 16 mM K+; 10 microM veratridine evoked a larger response. All three stimuli were Ca2+ dependent but only the response to veratridine was blocked by tetrodotoxin. Repetitive stimulations by 1 microM (-)-nicotine (100 microliters) at 30-min intervals resulted in similar levels of [3H]DA release; higher concentrations of (-)-nicotine resulted in an attenuation of the response particularly following the third stimulation. This may reflect desensitisation or tachyphylaxis of the presynaptic nicotinic receptor. The action of nicotine was markedly stereoselective: a 100-fold higher concentration of (+)-nicotine was necessary to evoke the same level of response as 1 microM (-)-nicotine. It is proposed that these presynaptic nicotinic receptors on striatal terminals are equivalent to high-affinity nicotine binding sites described in mammalian brain.     

Inhibition by Cyclic AMP of Phorbol Ester-Potentiated Norepinephrine Release from Guinea Pig Brain Cortical Synaptosomes

The involvement of Ca2+/phospholipid-dependent protein kinase (protein kinase C, PKC) and cyclic AMP-dependent protein kinase in the K+-evoked release of norepinephrine (NE) was studied using guinea pig brain cortical synaptosomes preloaded with [3H]NE. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a potent activator of PKC, enhanced the K+-evoked release of [3H]NE, in a concentration-dependent manner, but with no effect on the spontaneous outflow and uptake of [3H]NE in the synaptosomes. The apparent affinity of the evoked release for added calcium but not the maximally evoked release was increased by TPA (10(-7) M). Inhibitors of PKC, polymyxin B, and a more potent inhibitor, staurosporine, counteracted the TPA-induced potentiation of the evoked release. Both forskolin and dibutyryl cyclic AMP (DBcAMP) enhanced the evoked release, but reduced the TPA-potentiated NE release. A novel inhibitor of cyclic AMP-dependent protein kinase, KT5720, blocked both the forskolin-induced increase in the evoked release and its inhibition of TPA-induced potentiation in the evoked release, thereby suggesting that forskolin or DBcAMP counteracts the Ca2+-dependent release of NE by activating cyclic AMP-dependent protein kinase. These results suggest that the activation of PKC potentiates the evoked release of NE and that the activation of cyclic AMP-dependent protein kinase acts negatively on the PKC-activated exocytotic neurotransmitter release process in brain synaptosomes of the guinea pig.