Rabbit Polyclonal eIF4E phospho S209 antibody. Suitable for WB, ICC/IF and reacts with Rat, Human samples. Cited in 1 publication. Immunogen corresponding to Synthetic Peptide within Human EIF4E phospho S209.
pH: 7.3
Preservative: 0.05% Sodium azide
Constituents: PBS, 50% Glycerol (glycerin, glycerine), 1% BSA
WB | ICC/IF | |
---|---|---|
Human | Tested | Tested |
Mouse | Predicted | Predicted |
Rat | Tested | Expected |
Rabbit | Predicted | Predicted |
Species | Dilution info | Notes |
---|---|---|
Species Rat | Dilution info 1/1000 | Notes - |
Species Human | Dilution info 1/1000 | Notes - |
Species | Dilution info | Notes |
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Species Mouse, Rabbit | Dilution info - | Notes - |
Species | Dilution info | Notes |
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Species Human | Dilution info 1/250 | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Rat | Dilution info Use at an assay dependent concentration. | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rabbit | Dilution info - | Notes - |
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Acts in the cytoplasm to initiate and regulate protein synthesis and is required in the nucleus for export of a subset of mRNAs from the nucleus to the cytoplasm which promotes processes such as RNA capping, processing and splicing (PubMed:11606200, PubMed:22578813, PubMed:22684010, PubMed:24335285, PubMed:29987188). Component of the protein complex eIF4F, which is involved in the recognition of the mRNA cap, ATP-dependent unwinding of 5'-terminal secondary structure and recruitment of mRNA to the ribosome (By similarity). This protein recognizes and binds the 7-methylguanosine (m7G)-containing mRNA cap during an early step in the initiation of protein synthesis and facilitates ribosome binding by inducing the unwinding of the mRNAs secondary structures (PubMed:16271312, PubMed:22578813). Together with EIF4G1, antagonizes the scanning promoted by EIF1-EIF4G1 and is required for TISU translation, a process where the TISU element recognition makes scanning unnecessary (PubMed:29987188). In addition to its role in translation initiation, also acts as a regulator of translation and stability in the cytoplasm (PubMed:24335285). Component of the CYFIP1-EIF4E-FMR1 complex which binds to the mRNA cap and mediates translational repression: in the complex, EIF4E mediates the binding to the mRNA cap (By similarity). Component of a multiprotein complex that sequesters and represses translation of proneurogenic factors during neurogenesis (By similarity). In P-bodies, component of a complex that mediates the storage of translationally inactive mRNAs in the cytoplasm and prevents their degradation (PubMed:24335285). May play an important role in spermatogenesis through translational regulation of stage-specific mRNAs during germ cell development (By similarity). As well as its roles in translation, also involved in mRNA nucleocytoplasmic transport (By similarity). Its role in mRNA export from the nucleus to the cytoplasm relies on its ability to bind the m7G cap of RNAs and on the presence of the 50-nucleotide EIF4E sensitivity element (4ESE) in the 3'UTR of sensitive transcripts (By similarity). Interaction with the 4ESE is mediated by LRPPRC which binds simultaneously to both EIF4E and the 4ESE, thereby acting as a platform for assembly for the RNA export complex (By similarity). EIF4E-dependent mRNA export is independent of ongoing protein or RNA synthesis and is also NFX1-independent but is XPO1-dependent with LRPPRC interacting with XPO1 to form an EIF4E-dependent mRNA export complex (By similarity). Alters the composition of the cytoplasmic face of the nuclear pore to promote RNA export by reducing RANBP2 expression, relocalizing nucleoporin NUP214 and increasing expression of RANBP1 and RNA export factors DDX19 and GLE1 (By similarity). Promotes the nuclear export of cyclin CCND1 mRNA (By similarity). Promotes the nuclear export of NOS2/iNOS mRNA (PubMed:23471078). Promotes the nuclear export of MDM2 mRNA (PubMed:22684010). Promotes the export of additional mRNAs, including others involved in the cell cycle (By similarity). In the nucleus, binds to capped splice factor-encoding mRNAs and stimulates their nuclear export to enhance splice factor production by increasing their cytoplasmic availability to the translation machinery (By similarity). May also regulate splicing through interaction with the spliceosome in an RNA and m7G cap-dependent manner (By similarity). Also binds to some pre-mRNAs and may play a role in their recruitment to the spliceosome (By similarity). Promotes steady-state capping of a subset of coding and non-coding RNAs by mediating nuclear export of capping machinery mRNAs including RNMT, RNGTT and RAMAC to enhance their translation (By similarity). Stimulates mRNA 3'-end processing by promoting the expression of several core cleavage complex factors required for mRNA cleavage and polyadenylation, and may also have a direct effect through its interaction with the CPSF3 cleavage enzyme (By similarity). Rescues cells from apoptosis by promoting activation of serine/threonine-protein kinase AKT1 through mRNA export of NBS1 which potentiates AKT1 phosphorylation and also through mRNA export of AKT1 effectors, allowing for increased production of these proteins (By similarity).
EIF4EL1, EIF4F, EIF4E, Eukaryotic translation initiation factor 4E, eIF-4E, eIF4E, eIF-4F 25 kDa subunit, mRNA cap-binding protein
Rabbit Polyclonal eIF4E phospho S209 antibody. Suitable for WB, ICC/IF and reacts with Rat, Human samples. Cited in 1 publication. Immunogen corresponding to Synthetic Peptide within Human EIF4E phospho S209.
pH: 7.3
Preservative: 0.05% Sodium azide
Constituents: PBS, 50% Glycerol (glycerin, glycerine), 1% BSA
This phosphorylation site specific antibody is selective for eIF-4E containing a phosphate on serine 209.
Purified from rabbit serum by sequential epitope-specific chromatography. The antibody has been negatively preadsorbed using a non-phosphopeptide corresponding to the site of phosphorylation to remove antibody that is reactive with non-phosphorylated eIF-4E. The final product is generated by affinity chromatography using an eIF-4E-derived peptide that is phosphorylated at serine 209.
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The eIF4E protein also known as the eukaryotic translation initiation factor 4E functions mainly in the initiation phase of mRNA translation. It plays a role by binding to the 7-methylguanylate cap structure at the 5’ end of mRNA facilitating ribosome assembly on mRNA. The molecular weight of eIF4E is approximately 25 kDa and it is ubiquitously expressed in various tissues across eukaryotic organisms. Additionally phosphorylation of eIF4E affects its activity and is an important post-translational modification.
EIF4E regulates gene expression at the translational level and is a part of the eIF4F complex which also includes eIF4A and eIF4G. This complex is essential for the recruitment of ribosomes to the mRNA impacting the rate of protein synthesis. Changes in eIF4E levels can alter the translational efficiency of specific mRNAs thereby influencing cellular growth and proliferation.
EIF4E directly involves in the mTOR signaling pathway which regulates cell growth proliferation and survival. Within this pathway eIF4E interacts with proteins such as 4EBP1 which binds to eIF4E and inhibits its function in the absence of phosphorylation by mTOR. This interaction illustrates the control mechanism of protein synthesis under different cellular conditions connecting it to various signaling cascades.
EIF4E has been linked to cancer and neurodegenerative conditions. In cancer overexpression of eIF4E leads to increased translation of oncogenes facilitating tumorigenesis. The mTOR pathway proteins like mTOR and 4EBP1 are often dysregulated in these cases. Conversely its role in neurological disorders involves the aberrant regulation of synaptic proteins like PSD95 impacting neuronal communication and function.
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Immunofluorescence analysis using 70% confluent log phase U-87 MG cells labeled for Phospho-eIF4E pSer209 using ab4774. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with ab4774 at 1:250 dilution in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate at a dilution of 1:2000 for 45 minutes at room temperature (Panel A: green). Nuclei (Panel B: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI. F-actin (Panel C: red) was stained with Rhodamine Phalloidin (1:300). Panel D is a merged image showing cytoplasmic localization. Panel E is a no primary antibody control. The images were captured at 60X magnification.
Western blot analysis was performed on HeLa, HEK-293 (human cell lines) and L6 (rat cell line) cell lysates with various treatments, blotted for eIF4E (pS209) using ab4774. Two bands ~ 25 and 28 kDa band corresponding to eIF4E (pS209) was observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel, XCell SureLock™ Electrophoresis System and Novex® Sharp Pre-Stained Protein Standard. Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System. The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate
All lanes: Western blot - Anti-eIF4E (phospho S209) antibody (ab4774)
Lane 1: HeLa whole cell extracts at 20 µg
Lane 2: Serum starved HeLa at 20 µg
Lane 3: HeLa Serum Starved for overnight followed by Serum Released at 20 µg
Lane 4: HEK-293 at 20 µg
Lane 5: HEK-293 treated for 30 minutes with 25 µg/mL of Anisomycin at 20 µg
Lane 6: L6 at 20 µg
Lane 7: L6 treated for 10 minutes with 200 ng/mL of Insulin at 20 µg
All lanes: Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate at 0.4 µg/mL
Predicted band size: 25 kDa
Extracts of HeLa cells were resolved by SDS-PAGE on a 4-20% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer for one hour at room temperature, either left untreated (1-4) or treated with lambda (?) phosphatase (5), and then incubated with the
eIF4E (phospho S209) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1,5), the nonphosphopeptide
corresponding to the phosphopeptide immunogen (2), a
generic phosphoserine-containing peptide (3), or the phosphopeptide immunogen (4). After washing, the membrane was incubated with goat F(ab’)2 anti-rabbit IgG alkaline phosphatase and signals were detected using the Pierce SuperSignal™ method.
The data show that only the peptide corresponding to eIF4E (phospho S209) blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody i
All lanes: Western blot - Anti-eIF4E (phospho S209) antibody (ab4774)
Predicted band size: 25 kDa
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