Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP

REACTIVITY:

H M R

SENSITIVITY:

Endogenous

MW (kDa):

60

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q14457

Entrez-Gene Id:

8678

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:200

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb recognizes endogenous levels of Beclin-1 protein only when phosphorylated at Ser30.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser30 of human Beclin-1 protein.

Background

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of proteins activated in response to nutrient deprivation and in neurodegenerative conditions (1). One of the proteins critical to this process is Beclin-1, the mammalian orthologue of the yeast autophagy protein Apg6/Vps30 (2). Beclin-1 can complement defects in yeast autophagy caused by loss of Apg6 and can also stimulate autophagy when overexpressed in mammalian cells (3). Mammalian Beclin-1 was originally isolated in a yeast two-hybrid screen for Bcl-2 interacting proteins and has been shown to interact with Bcl-2 and Bcl-xL, but not with Bax or Bak (4). While Beclin-1 is generally ubiquitously expressed, research studies have shown it is monoallelically deleted in 40-75% of sporadic human breast and ovarian cancers (5). Beclin-1 is localized within cytoplasmic structures including the mitochondria, although overexpression of Beclin-1 reveals some nuclear staining and CRM1-dependent nuclear export (6). Investigators have demonstrated that Beclin-1-/- mice die early in embryogenesis and Beclin-1-/+ mice have a high incidence of spontaneous tumors. Stem cells from the null mice demonstrate an altered autophagic response, although responses to apoptosis appeared normal (7). Researchers have also found that overexpression of Beclin-1 in virally infected neurons in vivo resulted in significant protection against Sindbis virus-induced disease and neuronal apoptosis (4).
ULK1 phosphorylates Beclin-1 at Ser30 to induce autophagy in response to amino acid starvation, mTORC1 inhibition, and hypoxia (8).

  1. Reggiori, F. and Klionsky, D.J. (2002) Eukaryot Cell 1, 11-21.
  2. Kametaka, S. et al. (1998) J Biol Chem 273, 22284-91.
  3. Liang, X.H. et al. (1999) Nature 402, 672-6.
  4. Liang, X.H. et al. (1998) J Virol 72, 8586-96.
  5. Aita, V.M. et al. (1999) Genomics 59, 59-65.
  6. Liang, X.H. et al. (2001) Cancer Res 61, 3443-9.
  7. Yue, Z. et al. (2003) Proc Natl Acad Sci USA 100, 15077-82.
  8. Park, J.M. et al. (2018) Autophagy 14, 584-597.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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Revision 1
#35955

Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb

Western Blotting Image 1: Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb Expand Image
Western blot analysis of extracts from HCT 116 cells, untreated (-) or treated with Torin 1 #14379 (250 nM, 2 hr; +) and/or the ULK1 inhibitor SBI-0206965 #29089 (50 μM, 2 hr; +) as indicated, using Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb (upper), Beclin-1 (D40C5) Rabbit mAb #3495 (middle), and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Western Blotting Image 2: Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb Expand Image
Western blot analysis of extracts from C2C12 and H-4-II-E cells, untreated (-) or treated with Torin 1 #14379 (250 nM, 2 hr; +), using Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb (upper), Beclin-1 (D40C5) Rabbit mAb #3495 (middle), and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Western Blotting Image 3: Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb Expand Image
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with constructs expressing Myc/DDK-tagged full-length human Beclin-1 protein (hBeclin-1-Myc/DDK; +) and/or mouse ULK1 protein (mULK1; +), using Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Immunoprecipitation Image 1: Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb Expand Image
Immunoprecipitation of phospho-Beclin-1 (Ser30) from HCT 116 cell extracts treated with Torin 1 #14379 (250 nM, 2 hr). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb. Western blot analysis was performed using Phospho-Beclin-1 (Ser30) (E1C4X) Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L29A9) mAb (HRP Conjugate) #5127 was used as a secondary antibody.