Shank3 is a PSD scaffold protein that received increased attention in recent years, following observations that its mutations correlate with autism spectrum disorders. When Shank3 antibodies became suitable for immuno electron microscopy,we examined the association of this protein with the PSD. Labeling of dissociated hippocampal cultures with two different Shank3 antibodies showed a similar labeling pattern in the PSD complex, with a broad laminar distribution 30-100 nm from the postsynaptic membrane. Upon depolarization with high K+ for 2 min, or application of NMDA (50 M, 2 min), both the labeling intensity at PSD and the median distance of label from the postsynaptic membrane increased significantly, indicating that Shank3 molecules are preferentially recruited to the distal regions of the PSD complex under these excitatory conditions. Activity-induced accumulation of Shank at the PSD was reversed within 30 min upon return to basal medium. However, presence of Zn2+ in the incubation medium prevented loss of NMDA-induced Shank3 increase at the PSD. These data suggest that activity promotes reversible association of Shank3 with the PSD, which can be stabilized in the presence of Zn2+. These observations may help explain the roles of Zn2+ deficiency and Shank3 mutations in promoting autism spectrum disorders. Another project focused on the dynamics of AIDA-1 at the PSD. AIDA-1 is highly enriched PSD fractions and is considered to be a major component of the PSD complex. In cultured rat hippocampal neurons under basal conditions, immunogold label for AIDA-1 is mostly located within the dense core of the PSD, with a median distance of 30 nm from the postsynaptic membrane. Under excitatory conditions, such as depolarization with high K+ (90 mM, 2 min) or application of NMDA (50 M, 2 min), AIDA-1 label density at the PSD core was reduced to 40 % of controls and the median distance of label from the postsynaptic membrane increased to 55 nm. The movement of AIDA-1 was blocked by the application of a specific inhibitor of CaMKII. The effect of excitatory conditions on the postsynaptic distribution of AIDA-1 was reversed within 30 minutes after returning to control conditions. The reversible removal of AIDA-1 from the PSD core under excitatory conditions is similar to the redistribution of another abundant PSD protein, SynGAP. Both SynGAP-alpha1 and AIDA-1 are known to bind PSD-95. We speculate that activity-induced, CaMKII-mediated, transient translocation of these abundant proteins from the PSD core could promote structural flexibility, vacate sites on PSD-95 for the insertion of other components and thus may create a window for synaptic modification. An inhibitor of CaMKII demonstrated that accumulation of Shank under excitatory conditions is mediated by CaMKII. Under excitatory conditions, CaMKII mediates accumulation and activation of another PSD component, the deubiquitinase CYLD, in the pallium. Under basal conditions, IKK, another kinase present at the PSD, phosphorylates CYLD to promote CYLD activity, although at a lower level compared than by CaMKII. Altogether these data indicate that translocation of CYLD to the PSD results in its activation both under basal and excitatory conditions, and thus translocation to the pallium may have a regulatory role in synaptic function. In sum, this work reveals that the pallium, a second distinct layer lying deep to the core layer of the PSD and attached to it by GKAP-Shank interactions, is involved in regulation of proteins during synaptic activity.
| Dosemeci, Ayse; Burch, Amelia; Loo, Hannah et al. (2017) IRSp53 accumulates at the postsynaptic density under excitatory conditions. PLoS One 12:e0190250 |
| Burch, Amelia; Tao-Cheng, Jung-Hwa; Dosemeci, Ayse (2017) A novel synaptic junction preparation for the identification and characterization of cleft proteins. PLoS One 12:e0174895 |
| Dosemeci, Ayse; Toy, Dana; Burch, Amelia et al. (2016) CaMKII-mediated displacement of AIDA-1 out of the postsynaptic density core. FEBS Lett 590:2934-9 |
| Tao-Cheng, Jung-Hwa; Toy, Dana; Winters, Christine A et al. (2016) Zinc Stabilizes Shank3 at the Postsynaptic Density of Hippocampal Synapses. PLoS One 11:e0153979 |
| Dosemeci, Ayse; Weinberg, Richard J; Reese, Thomas S et al. (2016) The Postsynaptic Density: There Is More than Meets the Eye. Front Synaptic Neurosci 8:23 |
| Tao-Cheng, Jung-Hwa; Yang, Yijung; Reese, Thomas S et al. (2015) Differential distribution of Shank and GKAP at the postsynaptic density. PLoS One 10:e0118750 |
| Dosemeci, Ayse; Toy, Dana; Reese, Thomas S et al. (2015) AIDA-1 Moves out of the Postsynaptic Density Core under Excitatory Conditions. PLoS One 10:e0137216 |
| Thein, Soe; Pham, Anna; Bayer, K Ulrich et al. (2014) IKK regulates the deubiquitinase CYLD at the postsynaptic density. Biochem Biophys Res Commun 450:550-4 |
| Tao-Cheng, J-H; Thein, S; Yang, Y et al. (2014) Homer is concentrated at the postsynaptic density and does not redistribute after acute synaptic stimulation. Neuroscience 266:80-90 |
| Thein, Soe; Tao-Cheng, Jung-Hwa; Li, Yan et al. (2014) CaMKII mediates recruitment and activation of the deubiquitinase CYLD at the postsynaptic density. PLoS One 9:e91312 |
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