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Molecular Brain Sep 2021We recently showed that synaptophysin (Syph) and synapsin (Syn) can induce liquid-liquid phase separation (LLPS) to cluster small synaptic-like microvesicles in living...
We recently showed that synaptophysin (Syph) and synapsin (Syn) can induce liquid-liquid phase separation (LLPS) to cluster small synaptic-like microvesicles in living cells which are highly reminiscent of SV cluster. However, as there is no physical interaction between them, the underlying mechanism for their coacervation remains unknown. Here, we showed that the coacervation between Syph and Syn is primarily governed by multivalent pi-cation electrostatic interactions among tyrosine residues of Syph C-terminal (Ct) and positively charged Syn. We found that Syph Ct is intrinsically disordered and it alone can form liquid droplets by interactions among themselves at high concentration in a crowding environment in vitro or when assisted by additional interactions by tagging with light-sensitive CRY2PHR or subunits of a multimeric protein in living cells. Syph Ct contains 10 repeated sequences, 9 of them start with tyrosine, and mutating 9 tyrosine to serine (9YS) completely abolished the phase separating property of Syph Ct, indicating tyrosine-mediated pi-interactions are critical. We further found that 9YS mutation failed to coacervate with Syn, and since 9YS retains Syph's negative charge, the results indicate that pi-cation interactions rather than simple charge interactions are responsible for their coacervation. In addition to revealing the underlying mechanism of Syph and Syn coacervation, our results also raise the possibility that physiological regulation of pi-cation interactions between Syph and Syn during synaptic activity may contribute to the dynamics of synaptic vesicle clustering.
Topics: Amino Acid Substitution; Animals; Buffers; COS Cells; Chlorocebus aethiops; Fluorescence Recovery After Photobleaching; Genes, Reporter; Glycols; Humans; Hydrophobic and Hydrophilic Interactions; Ionic Liquids; Luminescent Proteins; Mice; Mutation, Missense; Osmolar Concentration; Phase Transition; Photochemistry; Point Mutation; Recombinant Fusion Proteins; Secretory Vesicles; Static Electricity; Synapsins; Synaptophysin; Time-Lapse Imaging; Tyrosine; Red Fluorescent Protein
PubMed: 34496937
DOI: 10.1186/s13041-021-00846-y -
Cell and Tissue Research Dec 1995Synaptophysin is one of the major integral membrane proteins of the small (30-50nm diameter) electron-translucent transmitter-containing vesicles in neurons and of...
Synaptophysin is one of the major integral membrane proteins of the small (30-50nm diameter) electron-translucent transmitter-containing vesicles in neurons and of similar vesicles in neuroendocrine cells. Since its expression is tightly linked to the occurrence of these vesicle types, we mutated the X-chromosomally located synaptophysin gene in embryonic stem cells for the generation of synaptophysin-deficient mice in order to study the consequence of synaptophysin ablation for the formation and function of such vesicles in vivo. The behavior and appearance of mice lacking synaptophysin was indistinguishable from that of their litter mates and reproductive capacity was comparable to normal mice. Furthermore, no drastic compensatory changes were noted in the expression of several other neuronal polypeptides or in the mRNA levels of synaptophysin isoforms, the closely related neuronal synaptoporin/synaptophysinII, and the ubiquitous pantophysin. Immunofluorescence microscopy of several neuronal and neuroendocrine tissues showed that overall tissue architecture was maintained in the absence of synaptophysin, and that the distribution of other synaptic vesicle components was not visibly affected. In electron-microscopic preparations, large numbers of vesicles with a diameter of 39.9nm and an electron-translucent interior were seen in synaptic regions of synaptophysin-deficient mice; these vesicles could be labeled by antibodies against synaptic vesicle proteins, such as synaptobrevin 2.
Topics: Adrenal Medulla; Animals; Brain; Cell Line; Cloning, Molecular; Female; Fertility; Mice; Mice, Inbred C57BL; Mice, Transgenic; Synapses; Synaptic Vesicles; Synaptophysin
PubMed: 8581936
DOI: 10.1007/BF00318874 -
Proceedings of the National Academy of... May 1996Synaptophysin (syp I) is a synaptic vesicle membrane protein that constitutes approximately 7% of the total vesicle protein. Multiple lines of evidence implicate syp I...
Synaptophysin (syp I) is a synaptic vesicle membrane protein that constitutes approximately 7% of the total vesicle protein. Multiple lines of evidence implicate syp I in a number of nerve terminal functions. To test these, we have disrupted the murine Syp I gene. Mutant mice lacking syp I were viable and fertile. No changes in the structure and protein composition of the mutant brains were observed except for a decrease in synaptobrevin/VAMP II. Synaptic transmission was normal with no detectable changes in synaptic plasticity or the probability of release. Our data demonstrate that one of the major synaptic vesicle membrane proteins is not essential for synaptic transmission, suggesting that its function is either redundant or that it has a more subtle function not apparent in the assays used.
Topics: Animals; Base Sequence; Calcium; Chromosome Mapping; DNA Primers; Electrophysiology; Humans; Mice; Mice, Knockout; Molecular Sequence Data; Neuronal Plasticity; Neurotransmitter Agents; Phenotype; Synaptic Transmission; Synaptic Vesicles; Synaptophysin
PubMed: 8643476
DOI: 10.1073/pnas.93.10.4760 -
Neurobiology of Disease Dec 2019The effects of prolonged physical training on memory performance and underlying presynaptic mechanisms were investigated in old C57BL/6 mice. Training via voluntary...
The effects of prolonged physical training on memory performance and underlying presynaptic mechanisms were investigated in old C57BL/6 mice. Training via voluntary running wheels was initiated at 16 months of age and continued for 5 months (1 h per day, 5 days per week), followed by testing of learning and memory functions and counting of presynaptic puncta and cholinergic inputs in the hippocampus. Trained old mice were compared to their age-matched sedentary controls and adult controls. This training strategy improved hippocampal-dependent spatial memory function tested via a novel location task, and enhanced memory was accompanied by restored presynaptic puncta and cholinergic fibers in area CA1 and DG of the hippocampus in old mice. Particularly, the training selectively affected presynaptic vesicle protein synaptophysin but not growth associated protein GAP-43, and the increased number of synaptophysin puncta positively correlates with improved memory performance. To better understand the neurochemical mechanisms by which prolonged physical training protects against aging-related memory deficits, the cholinergic inputs to the hippocampus were compared among the three groups of mice and correlated with memory performance. While the running prevented age-related loss of cholinergic inputs, it has limited impact on the projection source cells in the medial septum-diagonal band (MS-DB). Importantly, cholinergic fibers in area CA1 and DG positively correlated with spatial memory function. These data suggest that the preservation of presynaptic inputs, particularly those involved in the integrity of memory performance, contributes critically to the beneficial effects of physical running initiated at an older age.
Topics: Aging; Animals; Cholinergic Neurons; Hippocampus; Mice; Mice, Inbred C57BL; Physical Conditioning, Animal; Presynaptic Terminals; Spatial Memory; Synaptophysin
PubMed: 31470103
DOI: 10.1016/j.nbd.2019.104586 -
Hearing Research Nov 2003Light microscopy and immunohistochemical analyses of a freshly prepared human cochlea, removed at meningioma skull base surgery, were performed with particular emphasis...
Light microscopy and immunohistochemical analyses of a freshly prepared human cochlea, removed at meningioma skull base surgery, were performed with particular emphasis on synaptophysin (SY) reactivity. Synaptophysin, a 38-kDa glycoprotein, is one of the most abundant integral membrane proteins of small presynaptic vesicles and is a useful marker for sites of synaptic transmission of the efferent olivocochlear system in the cochlea. Following fixation and decalcification, cryosections of 30 microm were prepared. To introduce immunostaining, free-floating sections were exposed to monoclonal SY antibody. Positive SY immunostaining was solely restricted to the neural and sensory structures and did not include supporting cells of the organ of Corti. Dense reaction products were noted around the hair cells, especially at the basal portion of the inner and outer hair cells and their neural poles, as well as around the inner spiral bundle, tunnel spiral bundle, outer spiral bundle and upper tunnel crossing fibers. The majority of spiral ganglion cells stained positively. An intermingling network of thin unmyelinated nerve fibers stained densely, especially at the basal portions of the cochlea. The spiral limbus, inner and outer sulcus cells, basilar membrane, myelinated nerve fibers, spiral ligament and the stria vascularis were unstained. Human cochlea obtained during surgery offers excellent conditions for immunohistochemical analysis. In the basal cochlea in the organ of Corti, outer hair cell area, there may be alterations due to noise trauma from the drilling procedure.
Topics: Adult; Cochlea; Female; Hair Cells, Auditory; Humans; Immunohistochemistry; Nerve Fibers; Organ of Corti; Spiral Ganglion; Synaptophysin; Tissue Distribution
PubMed: 14599690
DOI: 10.1016/s0378-5955(03)00228-4 -
The British Journal of Psychiatry : the... Mar 2000Decreased expression of proteins such as synaptophysin in the hippocampus and prefrontal cortex in schizophrenia is suggestive of synaptic pathology. However, the...
BACKGROUND
Decreased expression of proteins such as synaptophysin in the hippocampus and prefrontal cortex in schizophrenia is suggestive of synaptic pathology. However, the overall profile of changes is unclear.
AIMS
To investigate synaptophysin gene expression in the cerebral cortex in schizophrenia.
METHOD
The dorsolateral prefrontal (Brodmann area [BA] 9/46), anterior cingulate (BA 24), superior temporal (BA 22) and occipital (BA 17) cortex were studied in two series of brains, totalling 19 cases and 19 controls. Synaptophysin was measured by immunoautoradiography and immunoblotting. Synaptophysin messenger RNA (mRNA) was measured using in situ hybridisation.
RESULTS
Synaptophysin was unchanged in schizophrenia, except for a reduction in BA 17 of one brain series. Synaptophysin mRNA was decreased in BA 17, and in BA 22 in the women with schizophrenia. No alterations were seen in BA 9/46.
CONCLUSIONS
Synaptophysin expression is decreased in some cortical areas in schizophrenia. The alterations affect the mRNA more than the protein, and have an unexpected regional distribution. The characteristics of the implied synaptic pathology remain to be determined.
Topics: Adult; Aged; Aged, 80 and over; Autoradiography; Blotting, Western; Case-Control Studies; Cerebral Cortex; Female; Gene Expression; Humans; In Situ Hybridization; Male; Middle Aged; RNA, Messenger; Regression Analysis; Schizophrenia; Synaptophysin
PubMed: 10755070
DOI: 10.1192/bjp.176.3.236 -
Synapse (New York, N.Y.) May 2007In the present study, unilateral cochlear ablations were performed in adult ferrets to evaluate possible time-dependent modifications of synaptophysin and insulin-like...
Synaptophysin and insulin-like growth factor-1 immunostaining in the central nucleus of the inferior colliculus in adult ferrets following unilateral cochlear removal: a densitometric analysis.
In the present study, unilateral cochlear ablations were performed in adult ferrets to evaluate possible time-dependent modifications of synaptophysin and insulin-like growth factor-1 (IGF-1) in the central nucleus of the inferior colliculus (CNIC). Using densitometric analysis, synaptophysin and IGF-1 immunostaining were assessed at 1 (PA1) and 90 (PA90) days after cochlear ablation. The results demonstrated that 1 day after the lesion there was an increase in the levels of synaptophysin immunostaining bilaterally in the CNIC compared to control animals. That increase was no longer present at 90 days after the ablation. Overall levels of IGF-1 immunostaining at PA1 were increased significantly within neurons and neuropil. However, at PA90, only IGF-1 immunostaining contralateral to the lesion was elevated compared to control animals, although elevation was less than that observed at PA1. These results suggest that cochlear ablation appears to affect synaptophysin and IGF-1 protein levels bilaterally in the CNIC.
Topics: Animals; Auditory Pathways; Cochlea; Densitometry; Functional Laterality; Immunohistochemistry; Inferior Colliculi; Insulin-Like Growth Factor I; Neuronal Plasticity; Synaptophysin; Time Factors
PubMed: 17318882
DOI: 10.1002/syn.20373 -
Journal of Neurochemistry Jul 2004Synaptophysin is one of the most abundant membrane proteins of small synaptic vesicles. In mature nerve terminals it forms a complex with the vesicular membrane protein...
Synaptophysin is one of the most abundant membrane proteins of small synaptic vesicles. In mature nerve terminals it forms a complex with the vesicular membrane protein synaptobrevin, which appears to modulate synaptobrevin's interaction with the plasma membrane-associated proteins syntaxin and SNAP25 to form the SNARE complex as a prerequisite for membrane fusion. Here we show that synaptobrevin is preferentially cleaved by tetanus toxin while bound to synaptophysin or when existing as a homodimer. The synaptophysin/synaptobrevin complex is, however, not affected when neuronal secretion is blocked by botulinum A toxin which cleaves SNAP25. Excessive stimulation with alpha-latrotoxin or Ca(2+)-ionophores dissociates the synaptophysin/synaptobrevin complex and increases the interaction of the other SNARE proteins. The stimulation-induced dissociation of the synaptophysin/synaptobrevin complex is not inhibited by pre-incubating neurones with botulinum A toxin, but depends on extracellular calcium. However, the synaptophysin/synaptobrevin complex cannot be directly dissociated by calcium alone or in combination with magnesium. The dissociation of synaptobrevin from synaptophysin appears to precede its interaction with the other SNARE proteins and does not depend on the final fusion event. This finding further supports the modulatory role the synaptophysin/synaptobrevin complex may play in mature neurones.
Topics: Animals; Botulinum Toxins, Type A; Brain Chemistry; Calcium; Cells, Cultured; Dimerization; Exocytosis; Hippocampus; Ionophores; Macromolecular Substances; Magnesium; Membrane Proteins; Mice; Nerve Tissue Proteins; Neurons; Protein Binding; R-SNARE Proteins; Rats; Spider Venoms; Stimulation, Chemical; Synaptophysin; Synaptosomal-Associated Protein 25; Synaptosomes; Tetanus Toxin
PubMed: 15198661
DOI: 10.1111/j.1471-4159.2004.02472.x -
Hippocampus 2006In the human neocortex, progressive synaptogenesis in early postnatal life is followed by a decline in synaptic density, then stability from adolescence until middle...
In the human neocortex, progressive synaptogenesis in early postnatal life is followed by a decline in synaptic density, then stability from adolescence until middle age. No comparable data are available in the hippocampus. In this study, the integral synaptic vesicle protein synaptophysin, measured immunoautoradiographically, was used as an index of synaptic terminal abundance in the hippocampal formation of 37 subjects from 5 weeks to 86 yr old, divided into 4 age groups (10 infants, 15 adolescents/young adults, 6 adults, and 6 elderly). In all hippocampal subfields, synaptophysin was lowest in infancy, but did not differ significantly between the older age groups, except in dentate gyrus (DG) where the rise was delayed until adulthood. A similar developmental profile was found in the rat hippocampus. We also measured synaptophysin mRNA in the human subjects and found no age-related changes, except in parahippocampal gyrus wherein the mRNA declined from infancy to adolescence, and again in old age. The synaptophysin protein data demonstrate a significant presynaptic component to human postnatal hippocampal development. In so far as synaptophysin abundance reflects synaptic density, the findings support an increase in hippocampal and parahippocampal synapse formation during early childhood, but provide no evidence for adolescent synaptic pruning. The mRNA data indicate that the maturational increases in synaptophysin protein are either translational rather than transcriptional in origin, or else are secondary to mRNA increases in neurons, the cell bodies of which lie outside the hippocampal formation.
Topics: Adolescent; Adult; Age Factors; Aged; Aged, 80 and over; Aging; Animals; Child; Child, Preschool; Hippocampus; Humans; Infant; Middle Aged; RNA, Messenger; Rats; Statistics as Topic; Synaptophysin
PubMed: 16807900
DOI: 10.1002/hipo.20194 -
Biophysical Journal Dec 2002The synaptic vesicle protein synaptophysin was solubilized from rat brain synaptosomes with a relatively low concentration of Triton X-100 (0.2%) and was highly purified...
The synaptic vesicle protein synaptophysin was solubilized from rat brain synaptosomes with a relatively low concentration of Triton X-100 (0.2%) and was highly purified (above 95%) using a rapid single chromatography step on hydroxyapatite/celite resin. Purified synaptophysin was reconstituted into a planar lipid bilayer and the channel activity of synaptophysin was characterized. In asymmetric KCl solutions (cis 300 mM/trans 100 mM), synaptophysin formed a fast-fluctuating channel with a conductance of 414 +/- 13 pS at +60 mV. The open probability of synaptophysin channels was decreased upon depolarization, and channels were found to be cation-selective. Synaptophysin channels showed higher selectivity for K(+) over Cl(-) (P(K(+))/P(Cl(-)) > 8) and preferred K(+) over Li(+), Na(+), Rb(+), Cs(+), or choline(+). The synaptophysin channel is impermeable to Ca(2+), which has no effect on its channel activity. This study is the second demonstration of purified synaptophysin channel activity, but the first biophysical characterization of its channel properties. The availability of large amounts of purified synaptophysin and of its characteristic channel properties might help to establish the role of synaptophysin in synaptic transmission.
Topics: Animals; Brain; Cesium; Chlorides; Choline; Comet Assay; Gluconates; Ion Channel Gating; Ion Channels; Lipid Bilayers; Membrane Potentials; Membranes, Artificial; Potassium Chloride; Rats; Rubidium; Sodium Chloride; Synaptic Vesicles; Synaptophysin; Synaptosomes
PubMed: 12496091
DOI: 10.1016/S0006-3495(02)75324-1