Anti-GRWD1 antibody, rabbit polyclonal
Product code70-130Size

100 µgStorage-20℃

Concentration

1.0 mg/ml

Buffer

PBS- with 50% glycerol

PurityPurified IgG fraction with protein A from rabbit antiserum absorbed with GSTagarose column

ImmunogenPurified GST-GRWD1 (human, full-length) expressed in E. coli.

IsotypeRabbit IgG

Reactivity

Specific to GST and GST-tagged proteinsSpecial notes

Validated for Western Blotting by siRNA.

Application

1. Western blotting (1/1,000~1/3,000 dilution)
2. Immunoprecipitation (Assay dependent)
3. Chromatin Immuno-Precipitation (Assay dependent)
4. Immunofluorescence staining / Immunochemistry (1/100~1/1,000 dilution)
KEYWORDS: nucleolar stress response, oncogene, p53, PICT1, RPL11, tumor
suppressor

Background

Glutamate-rich WD repeat-containing protein 1 (GRWD1) consists of 446 aminoacids with molecular mass of 49.4 kDa. It has been found as a protein whichinteracts with METTL18 and CDT1 proteins. It has been implicated inregulation of DNA replication and/or in ribosome biogenesis.

Data Link

UniProtKB Q9BQ67 (GRWD1_HUMAN)

Q810D6 (GRWD1_MOUSE)

Please note: All products are FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC
PROCEDURES. NOT FOR MILITARY USE.

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Data Images: 70-130 Anti-GRWD1 antibody, rabbit polyclonal

Fig.1. Identication of GRWD1 proteins in whole cell lysates by western blotting with anti-GRWD1
antibody
Whole cell lysates of HeLa cells untreated (-) and treated (+) with DNA damaging agent, adriamycin(Ad), and NIH3T3 cells were analyzed by western blotting with anti-GRWD1 antibody at 1/1,000dilution. The samples were 10 μg. Second antibody was HRP-conjugated goat anti-rabbit IgG usedat 1/5,000 dilution. The revelation of multiple bands indicates post-translational modification such as
phosphorylation. The level of GRWD1 in the cell was not affected by DNA-damaging treatment. Theidentity of an additional band at ~85 kDa position other than the GRWD1 band in NIH-3T3 cell lysate
is not known. The GRWD1 proteins were identified at a position higher (~55 kDa) than expected fromthe molecular mass of GRWD1 indicated from cDNA sequence (49.4 kDa).

Fig.2. Immunofluorescence staining of GRWD1 protein in HeLa cells.
Hela cells were fixed in 4% paraformaldehyde overnight and permeabilized
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in 0.25% TritonX 100 in PBS for 10 min. Anti-GRWD1 antibody was used at 1/1,000 dilution. As
second antibody, goat anti-rabbit IgG conjugated with Alex488 was used at 1/5,000 dilution. As asignal enhancer, Can Get Signal Immunostain B (Toyobo, Osaka) was used according to the protocolof the supplier. Nuclei were stained with DAPI.
GRWD1 protein is localized in nuclei.

Fig.3. Nucleolar localization of GRWD1.
HCT116 cells were first extracted with Triton X-100 to remove nucleoplasmic proteins, doubleimmunostained with anti-GRWD1 (green) and anti-fibrillarin (red) antibodies as a marker for nucleoli,and counterstained with DAPI. The anti-GRWD antibody was used at 1/500 dilution and as the second
antibody, goat anti-rabbit IgG conjugated with Alex488 was used at 1/2,000 dilution.
Reference: This product has been described in Ref.1 and used in the following publication.
1.Sugimoto N. et al. Identification of novel human Cdt1-binding proteins by a proteomics approach:proteolytic regulation by APC/CCdh1. Mol Biol Cell. 2008 19(3):1007-21. WB (human)
2.Sugimoto N. et al. Cdt1-binding protein GRWD1 is a novel histone-binding protein that facilitates
MCM loading through its influence on chromatin architecture. Nucleic Acids Res. 2015 Jul
13;43(12):5898-911. PMID:25990725 WB, IP, ChIP, IC/IF (human)
3.Aizawa M. et al. Nucleosome assembly and disassembly activity of GRWD1, a novel Cdt1-binding
proteinthat promotes pre-replication

complexformation. Biochim Biophys

Acta. 2016

Nov;1863(11):2739-2748. PMID:27552915 WB (human)
4. Kayama K. et al. GRWD1 negatively regulates p53 via the RPL11-MDM2 pathway and promotes
tumorigenesis. EMBO Rep. 2017 Jan;18(1):123-137. PMID:27856536 WB,IF, IP (human)

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