The human PPP1R9A gene encoding neurabin I is adjacent to SGCE, PEG10, and ASB4. Neurabin I (neural tissue specific F-actin-binding protein I) was originally purified from rat brain and has been characterised as a key candidate molecule in synaptic formation and function. It has also been shown that neurabin I is highly concentrated in the synapse of developed neurons and is involved in neurite formation. The deduced 1098 amino acid sequence for rat neurabin I contains an F-actin-binding domain, a PDZ domain, and coiled-coil domains. As the name indicates, the expression of the gene and protein was shown to be brain specific in adult rat tissues. The gene symbol is based on ‘‘protein phosphatase 1, regulatory (inhibitor) subunit 9A’’ because neurabin I has been shown to bind protein phosphatase I (PP1) and inhibit its activity. The PP1/ neurabin I complex has been demonstrated to regulate actin cytoskeleton dynamics. Furthermore, molecules implicated in the control of neuronal morphology, such as p70S6K and kalirin-7 (a GTP-exchange factor), have been identified to bind the PDZ domain of neurabin I. Lastly, the C-terminus of neurabin I binds the trans-Golgi protein TGN38, suggesting it is a multi-functional molecule that regulates membrane and cytoskeletal functions.
Background References
1. Qiao M. et. al. Polymorphisms in PEG10 and PPP1R9A genes are associated with porcine carcass and meat quality traits. Anim Genet. 2016 Apr
2. Konopaske GT. et. al. Altered prefrontal cortical MARCKS and PPP1R9A mRNA expression in schizophrenia and bipolar disorder. Schizophr Res. 2015 May
3. K Nakabayashi. et al. Genomic imprinting of PPP1R9A encoding neurabin I in skeletal muscle and extra-embryonic tissues.J Med Genet. 2004 Aug
Western blot analysis of Neurabin 1 on rat brain lysate with Rabbit anti-Neurabin 1 antibody (HA500169) at 1/1,000 dilution.
Lysates/proteins at 15 µg/Lane. Exposure time: 30 seconds; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature Primary antibody: HA500169, 1/1,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃ Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 123.3 kDa Observed band size: 123 kDa
ICC staining of Neurabin 1 in SH-SY5Y cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (HA500169, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
Immunohistochemical analysis of paraffin-embedded rat brain tissue using anti-Neurabin 1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500169, 1/400) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded human liver carcinoma tissue using anti-Neurabin 1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500169, 1/400) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded mouse brain tissue using anti-Neurabin 1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500169, 1/400) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Flow cytometric analysis of Neurabin 1 was done on SHSY5Y cells. The cells were fixed, permeabilized and stained with the primary antibody (HA500169, 1/50) (red). After incubation of the primary antibody at room temperature for an hour, the cells were stained with a Alexa Fluor 488-conjugated Goat anti-Rabbit IgG Secondary antibody at 1/1000 dilution for 30 minutes.Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
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