Protein phosphorylation in nervous tissue: possible involvement in nervous tissue function and relationship to cyclic nucleotide metabolism

Williams, Michael; Rodnight, Richard

doi:10.1016/0301-0082(77)90006-5

Cited by 92 publications

(18 citation statements)

References 344 publications

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“…This involved quantitatively extracting Proteins la and III from each of the subcellular fractions, phosphorylating them, subjecting the extracts to SDS-PAGE, and quantitating the amounts of Proteins la and Ib as described previously (36) . 45 Itg for control and 90 jig for proteolytic digestion) and crude synaptic vesicle (SV, 30 hg for control and 60 1~g for proteolytic digestion) fractions were phosphorylated and subjected to SDS-gel electrophoresis as described in Materials and Methods under Assay of Protein I . The gel was stained, destained, and dried, and subjected to autoradiography to identify the positions of Protein la and Protein Ib in the gel.…”

Section: Phosphorylation Of Proteins Ia and Ib In Subcellular Fractionsmentioning

confidence: 99%

Subcellular distribution in cerebral cortex of two proteins phosphorylated by a cAMP-dependent protein kinase.

Ueda

Greengard

Berzins

et al. 1979

The Journal of Cell Biology

198

104

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The subcellular distribution of Proteins la and lb, two proteins which serve as specific substrates for protein kinases present in mammalian brain, was studied in the dog cerebral cortex. Proteins Ia and Ib were found to be most highly enriched in synaptic vesicle fractions ; they were also present in postsynaptic density and synaptic membrane fractions in significant amounts, Proteins la and Ib present in the synaptic vesicle fraction appear to be similar, if not identical, to those present in the postsynaptic density fraction as judged by several criteria : (a) the ability to serve as substrate for cAMP-dependent protein kinase, (b) electrophoretic mobility in the presence of sodium dodecyl sulfate, (c) extractability with NH4 C1 or EGTA, and (d) fragmentation to electrophoretically similar peptides by a purified Staphylococcus aureus protease . In addition, the postsynaptic density fraction has been found to contain cAMP-dependent Protein Ia and Protein Ib kinase activity . The subcellular localization of Proteins la and Ib suggests a role for these proteins in the physiology of the synapse .

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Section: Phosphorylation Of Proteins Ia and Ib In Subcellular Fractionsmentioning

confidence: 99%

Subcellular distribution in cerebral cortex of two proteins phosphorylated by a cAMP-dependent protein kinase.

Ueda

Greengard

Berzins

et al. 1979

The Journal of Cell Biology

198

104

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“…6). Hence it is plausible that AcCho liberation, triggered in vivo and in vitro by an increased level of free Ca2+ in the nerve ending's cytoplasm (7)(8)(9), is accompanied by the phosphorylation of specific proteins.…”

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confidence: 99%

Mechanism of acetylcholine release: possible involvement of presynaptic muscarinic receptors in regulation of acetylcholine release and protein phosphorylation.

Michaelson¹,

Avissar²,

Kloog³

et al. 1979

Proc. Natl. Acad. Sci. U.S.A.

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Acetylcholine (AcCho) release from purely cholinergic Torpedo synaptosomes was evoked by K+ depolarization in the presence of Ca2+. Activation of muscarinic receptors, present in the synaptosomal fraction, by the agonist oxotremorine resulted in the inhibition of AcCho liberation. This inhibition was abolished by the muscarinic antagonist atropine, which by itself has no effect. These findings suggest that the muscarinic receptor, present in the electric organ of Torpedo, is presynaptic and that its physiological function is to regulate AcCho release by negative feedback. The mechanism of presynaptic muscarinic inhibition was investigated by examining the effect of muscarinic ligands on synaptosomal 45Ca2+ uptake and on the level of phosphorylation of specific synaptosomal proteins. Ca2+-dependent K+ depolarization-induced synaptosomal AcCho release was accompanied by 45Ca2+ uptake and by a marked increase in the phosphorylation of a specific synaptosomal protein (band a) of approximately 100,000 daltons. Activation of the muscarinic receptor by the agonist oxotremorine had no detectable effect on synaptosomal 45Ca2+ uptake but resulted in the concomitant inhibition of AcCho release and of phosphorylation of band a. The muscarinic antagonist atropine abolished the inhibitory effect of oxotremorine both on AcCho liberation and on phosphorylation of band a. These findings suggest that phosphorylation of band a may be involved in regulation of the presynaptic processes that underly AcCho release and that activation of the muscarinic receptor' by agonists may inhibit AcCho release by blocking the phosphorylation of band a.It has been suggested that some cholinergic synapses contain presynaptic muscarinic receptors which regulate the extent of acetylcholine (AcCho) release by feedback inhibition (1-5). The molecular mechanisms underlying AcCho liberation, and in particular its modulation by muscarinic receptors, are not known.Protein phosphorylation and dephosphorylation is now recognized as a ubiquitous regulatory mechanism of enzymatic and cellular activities (for review, see ref. 6). Hence it is plausible that AcCho liberation, triggered in vivo and in vitro by an increased level of free Ca2+ in the nerve ending's cytoplasm (7-9), is accompanied by the phosphorylation of specific proteins. Krueger et al. (10) have shown that agents which increase Ca2+ transport into intact rat brain synaptosomes stimulate the incorporation of 32P, into synaptosomal proteins of apparent subunit molecular weights of 80,000 and 86,000. DeLorenzo (11) and Hershkowitz (12) used a preparation of rat brain synaptosomes and [y-32P]ATP and found Ca2+-dependent phosphorylation of specific proteins in the range of 60,000, 50,000, and 42,000 daltons. These findings show that Ca2+ can modulate protein phosphorylation of rat brain synaptosomes. Brain synaptosomes contain a mixture of nerve endings, and these endings use various neurotransmitters. Consequently, the phosphorylation observed in such preparations represents that of an ave...

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“…In postdoctoral work, M.W. (one of the authors of this appendix) was researching changes in the phosphorylation of synaptic proteins as a molecular substrate of memory (Williams and Rodnight, ). After 10 months of intensive research, the initial increases in phosphorylation of the P2 synaptosomal fraction of rat brain isolated via a standard ultracentrifugation technique using molar sucrose concentrations could not be replicated.…”

Section: Hard Copy Paper Notebooksmentioning

confidence: 99%

Laboratory Notebooks and Data Storage

Williams¹,

Bozyczko-Coyne²,

Dorsey³

et al. 2008

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The laboratory notebook, whether in hardcopy or electronic (ELN) format, represents a true, chronological record of a scientist's bench work, incorporating the primary source of all materials and information related to the design, execution, and outcomes (data) for a specific experiment. The notebook content thus represents a precise, legible record of what was done, why, and by whom. It also describes the outcome of the experiment and how this is related to the initial hypothesis on which performing the experiment was based. As an archive, a laboratory notebook allows others previously unassociated with the work to understand what was done and, if necessary, independently recreate the experiments. A laboratory notebook and its associated records (e.g., computer printouts or HPLC traces) also represent the vital record of the conception date of an invention and its reduction to practice, providing a timely and necessary legal record to support and defend patent applications.

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Protein phosphorylation in nervous tissue: possible involvement in nervous tissue function and relationship to cyclic nucleotide metabolism

Cited by 92 publications

References 344 publications

Subcellular distribution in cerebral cortex of two proteins phosphorylated by a cAMP-dependent protein kinase.

Subcellular distribution in cerebral cortex of two proteins phosphorylated by a cAMP-dependent protein kinase.

Mechanism of acetylcholine release: possible involvement of presynaptic muscarinic receptors in regulation of acetylcholine release and protein phosphorylation.

Laboratory Notebooks and Data Storage

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