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Mouse psd95 coimmunoprecipitation from excitatory or inhibitory neurons and mass spectrometry

ABSTRACT: We used a psd95-GFP cre dependent KI mouse to label PSD95 with GFP specifically in excitatory (CaMK) or inhibitory (vGAT) neurons. PSD95 was pulled down with a GFP nanobody and the resulting complex was used for massspectrometry analysis.

INSTRUMENT(S): LTQ

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Brain

SUBMITTER: Alexei Bygrave

LAB HEAD: Richard Huganir

PROVIDER: PXD041635 | Pride | 2023-06-03

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	TB-QC-190717_HuganirR_AB_Area_PhosST_Pairwise_All_peptides-MGF.mgf	Mgf
	TB-QC-190717_HuganirR_AB_Area_PhosST_Pairwise_All_peptides.msf	Msf
	TB-QC-190717_HuganirR_AB_Area_PhosST_Pairwise_All_peptides.mzData	Other
	TB-QC-190717_HuganirR_AB_Area_PhosST_Pairwise_All_peptides.pep.xml	Pepxml
	TB-QC-190717_HuganirR_AB_Area_PhosST_Pairwise_All_peptides.prot.xml	Xml

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1 - 5 of 8

Publications

Btbd11 supports cell-type-specific synaptic function.

Bygrave Alexei M AM Sengupta Ayesha A Jackert Ella P EP Ahmed Mehroz M Adenuga Beloved B Nelson Erik E Goldschmidt Hana L HL Johnson Richard C RC Zhong Haining H Yeh Felix L FL Sheng Morgan M Huganir Richard L RL

Cell reports 20230531 6

Synapses in the brain exhibit cell-type-specific differences in basal synaptic transmission and plasticity. Here, we evaluated cell-type-specific specializations in the composition of glutamatergic synapses, identifying Btbd11 as an inhibitory interneuron-specific, synapse-enriched protein. Btbd11 is highly conserved across species and binds to core postsynaptic proteins, including Psd-95. Intriguingly, we show that Btbd11 can undergo liquid-liquid phase separation when expressed with Psd-95, su ...[more]

PMID: 37261953

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fiber transformation without CaMK pathway in electric-stimulation

Project description:Comparing the gene expression profiles of day 3 of in vitro cultured FDB fibers with or without slow-fiber-type pattern of electrical stimulation, with inhibition of CaMK signaling pathways(KN93 or KN62), with amplified RNA (cRNA probes). ----CaMK inhibition (KN93): GSM154001, GSM154002, GSM154003, GSM154004, GSM154025, and GSM154026. ----CaMK inhibition (KN62)GSM154027, GSM154028, GSM154029, GSM154030, GSM154031, and GSM154032. ----Negative control (KN92): GSM154033, GSM154034, GSM154035, GSM154036, GSM154037, and GSM154038. Keywords: cell treatment

2007-01-18 | GSE6660 | GEO

Ptchd1 deficiency induces excitatory synaptic and cognitive dysfunctions in mouse.

Project description:Synapse development and neuronal activity represent fundamental processes for the establishment of cognitive function. Structural organization as well as signalling pathways from receptor stimulation to gene expression regulation are mediated by synaptic activity and misregulated in neurodevelopmental disorders such as autism spectrum disorder (ASD) and intellectual disability (ID). Deleterious mutations in the PTCHD1 (Patched domain containing 1) gene have been described in male patients with X-linked ID and/or ASD. The structure of PTCHD1 protein is similar to the Patched (PTCH1) receptor; however, the cellular mechanisms and pathways associated with PTCHD1 in the developing brain are poorly determined. Here we show that PTCHD1 displays a C-terminal PDZ-binding motif that binds to the postsynaptic proteins PSD95 and SAP102. We also report that PTCHD1 is unable to rescue the canonical sonic hedgehog (SHH) pathway in cells depleted of PTCH1, suggesting that both proteins are involved in distinct cellular signalling pathways. We find that Ptchd1 deficiency in male mice (Ptchd1-/y) induces global changes in synaptic gene expression, affects the expression of the immediate-early expression genes Egr1 and Npas4 and finally impairs excitatory synaptic structure and neuronal excitatory activity in the hippocampus, leading to cognitive dysfunction, motor disabilities and hyperactivity. Thus our results support that PTCHD1 deficiency induces a neurodevelopmental disorder causing excitatory synaptic dysfunction.

2017-04-14 | GSE80312 | GEO

Comparing Control and Schizophrenic hiPSC-derived Neurons

Project description:Schizophrenia is a debilitating neurological disorder for which no cure exists. Few defining characteristics of schizophrenic neurons have been identified and the molecular mechanisms responsible for schizophrenia are not well understood, in part due to the lack of patient material for study. Human induced pluripotent stem cells (hiPSCs) offer a new strategy for studying schizophrenia. We have created the first cell-based human model of a complex genetic psychiatric disease by generating hiPSCs from schizophrenic patients and subsequently differentiating these cells to hiPSC-derived neurons in vitro. Schizophrenic hiPSC-derived neurons showed diminished neuronal connectivity in conjunction with decreased neurite number, PSD95-protein levels and glutamate receptor expression. Gene expression profiles of schizophrenic hiPSC-derived neurons identified altered expression of many components of the cAMP and WNT signaling pathways. Key cellular and molecular elements of the schizophrenic phenotype were ameliorated following treatment of schizophrenic hiPSC-derived neurons with the antipsychotic loxapine. 3 independent differentiations (biological replicates) for each of four control and four schizophrenic patients were analyzed.

2011-05-01 | E-GEOD-25673 | biostudies-arrayexpress

coimmunoprecipitation of BdRFS identifies putative itneracting proteins

Project description:In this work we sought to identify putative interacting proteins of the Brachypodium distachyon Regulator of Stress and Flowering protein (BdRFS) from healthy leaf tissue. A truncated form of BdRFS lacking the N-terminal 45 residues was necessary for solubilization. THe protein is tagged N-terminally with His-SUMO tag.

2023-03-11 | PXD033835 | Pride

Two-Step Coimmunoprecipitation (TIP) Enables Efficient and Highly Selective Isolation of Native Protein Complexes.

Project description:Coimmunoprecipitation (co-IP) is one of the most frequently used techniques to study protein-protein (PPIs) or protein-nucleic acid interactions (PNIs). However, the presence of coprecipitated contaminants is a well-recognized issue associated with single-step co-IPs. To overcome this limitation, we developed the two-step co-IP (TIP) strategy that enables sequential coimmunoprecipitations of endogenous protein complexes. TIP can be performed with a broad range of mono- and polyclonal antibodies targeting a single protein or different components of a given complex. TIP results in a highly selective enrichment of protein complexes and thus outperforms single-step co-IPs for downstream applications such as mass spectrometry for the identification of PPIs and quantitative PCR for the analysis of PNIs. We benchmarked TIP for the identification of CD95/FAS-interacting proteins in primary human CD4+ T cells, which recapitulated all major known interactors, but also enabled the proteomics discovery of PPM1G and IPO7 as new interaction partners. For its feasibility and high performance, we propose TIP as an advanced tool for the isolation of highly purified protein-protein and protein-nucleic acid complexes under native expression conditions.

| S-EPMC5930409 | biostudies-literature

Endogenous brain ARC complexes

Project description:Arc is an activity regulated neuronal protein yet little is known about its protein interactions, assembly into multiprotein complexes, role in human disease and cognition. We applied an integrated proteomic and genetic strategy using targeted tagging of a Tandem Affinity Purification (TAP) tag and Venus fluorescent protein into the endogenous Arc gene in mice, biochemical and proteomic characterization of native complexes in wild type and knockout mice, and human genetic analyses of disease and intelligence. TAP tagging enabled efficient purification of complexes and identification of many novel Arcinteracting proteins, of which PSD95 was the most abundant. PSD95 was essential for Arc assembly into 1.5 MDa complexes and activity-dependent recruitment to excitatory synapses. Integrating human genetic data with proteomic data showed postsynaptic Arc- PSD95 complexes are enriched in schizophrenia, intellectual disability, autism and epilepsy mutations and normal variants in intelligence. Arc-PSD95 postsynaptic complexes are a molecular substrate for the convergence of normal and pathological genetic variants impacting on human cognitive function.

2017-10-19 | PXD007283 | Pride

Project description:A survey of pairwise epistasis in PSD95 PDZ3

| PRJNA642160 | ENA

Mus musculus

Project description:fiber transformation without CaMK pathway in electric-stimulation

| PRJNA104157 | ENA

Transcriptome analysis of wt Pilus-1 expressing Streptococcus pneumoniae strains

Project description:The Streptococcus pneumoniae Pilus-1 Displays a Biphasic Expression Pattern

2011-07-01 | E-TABM-1154 | biostudies-arrayexpress

Coimmunoprecipitation and following mass spectrometry analysis of EhRacJ

Project description:To find out the binding partners of EhRacJ, crosslinking and coimmunoprecipitation were performed. Then, eluted samples were submitted to Mass Spectrometry and Proteomics Core Facility, Johns Hopkins University School of Medicine and analyzed by mass spctrometry (shotgun proteomics). The coimmunoprecipitation and following analysis were conducted only a single time. You can find data of HA-EhRacJ from "2023_3_13_28_AM_FileSize_1261863517_Byte_F012933.mzid.gz", "2023_3_13_28_AM_FileSize_1261863517_Byte_F012933.mzid_2023_3_13_28_AM_File_Size__1261863517__Byte__F012933.MGF", "JS-E480-CS_230524_NozakiT_MS_DDA_S1_10pct.msf", "JS-E480-CS_230524_NozakiT_MS_DDA_S1_10pct.mzML", and "JS-E480-CS_230524_NozakiT_MS_DDA_S1_10pct.RAW". You can find data of pEhEx-HA (mock) from "2023_6_54_27_PM_FileSize_1168419053_Byte_F012936.mzid.gz", "2023_6_54_27_PM_File Size_1168419053_Byte_F012936.mzid_2023_6_54_27_PM_File_Size__1168419053__Byte__F012936.MGF", "JS-E480-CS_230524_NozakiT_MS_DDA_S4_10pct.msf", "JS-E480-CS_230524_NozakiT_MS_DDA_S4_10pct.mzML", and "JS-E480-CS_230524_NozakiT_MS_DDA_S4_10pct.RAW".

2024-06-20 | PXD051913 | Pride

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