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Augurin

Human Augurin

(A) ECRG4 encodes conserved, potentially secreted proteins. Homologies between species of Ecrg4 encoded proteins
(B) Candidate proteins generated from Ecrg4 gene expression. Eight potential protein products are predicted by algorithms to be generated from the single gene: intact Ecrg4(1-148); its leader sequence, augurin, argilin and ecilin and their δ-16C-terminal cleaved homologs: C-δ-16-augurin, C-δ-16-argilin and the δ-16 peptide itself.

 

HPLC Analysis of Human Augurin

Mass Spec Analysis of Human Augurin

 

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Cell surface localization and release of the candidate tumor suppressor Ecrg4 from polymorphonuclear cells and monocytes activate macrophages.

We identified fresh human leukocytes as an abundant source of the candidate epithelial tumor suppressor gene, Ecrg4, an epigenetically regulated gene, which unlike other tumor suppressor genes, encodes an orphan-secreted, ligand-like protein. In human cell lines, Ecrg4 gene expression was low, Ecrg4 protein undetectable, and Ecrg4 promoter hypermethylation high (45-90%) and reversible by the methylation inhibitor 5-AzaC. In contrast, Ecrg4 gene expression in fresh, normal human PBMCs and PMNs was 600-800 times higher than in cultured cell lines, methylation of the Ecrg4 promoter was low (<3%), and protein levels were readily detectable in lysates and on the cell surface. Flow cytometry, immunofluorescent staining, and cell surface biotinylation established that full-length, 14-kDa Ecrg4 was localized on PMN and monocyte cell surfaces, establishing that Ecrg4 is a membrane-anchored protein. LPS treatment induced processing and release of Ecrg4, as detected by flow and immunoblotting, whereas an effect of fMLF treatment on Ecrg4 on the PMN cell surface was detected on the polarized R2 subpopulation of cells. This loss of cell surface Ecrg4 was associated with the detection of intact and processed Ecrg4 in the conditioned media of fresh leukocytes and was shown to be associated with the inflammatory response that follows severe, cutaneous burn injury. Furthermore, incubation of macrophages with a soluble Ecrg4-derived peptide increased the P-p65, suggesting that processing of an intact sentinel Ecrg4 on quiescent circulating leukocytes leads to processing from the cell surface following injury and macrophage activation.

Baird A, et al., J Leukoc Biol. 2012 Mar 6.

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Processing of proaugurin is required to suppress proliferation of tumor cell lines.

Ozawa et al. Molecular Endocrinology May 1, 2011 vol. 25 no. 5 776-784

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Ecrg4 expression and its product augurin in the choroid plexus: impact on fetal brain development, cerebrospinal fluid homeostasis and neuroprogenitor cell response to CNS injury

Gonzalez et al. Fluids Barriers CNS. 2011 Jan 18;8(1):6.

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Peptides from prepro-Augurin identified by MALDI-TOF mass spectrometry

Ozawa A et. Al. Mol Endocrinol. 2011 May;25(5):776-84. Epub 2011 Mar 24.

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Augurin stimulates the hypothalamo-pituitary-adrenal axis via the release of corticotrophin-releasing factor in rats.

Tadross et al. Br J Pharmacol. 2010 Apr;159(8):1663-71. Epub 2010 Mar 8.

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The bioactive peptide, E42-R70, has been synthesized as Phoenix´s catalog numbers 012-19 (human) and 012-20 (mouse).

Gonzalez et al. Fluids and Barriers of the CNS 2011 8:6

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Identification of novel peptide hormones in the human proteome by hidden Markov model screening

Mirabeau et al. Genome Res. 2007 Mar;17(3):320-7. Epub 2007 Feb 6.

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Colocalization of Spexin and Augurin with Insulin in endocrine cells.

Mirabeau et al. Genome Res. 2007 Mar;17(3):320-7. Epub 2007 Feb 6.

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Identification of spexin and augurin in cell supernatants.

Mirabeau et al. Genome Res. 2007 Mar;17(3):320-7. Epub 2007 Feb 6.

Human Augurin Prohormone Schematic and Sequence

 

Alignment of Human, Rat and Mouse Augurin

alignment of human rat mouse augurin

Mapping in Human Pancreas Tissue (H-012-22)

Tissue Sample

Human pancreas tissue

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

N/A

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Rabbit Anti-Augurin, Prepro (71-107) (H) Antiserum (Catalog No.: H-012-22)

Optimal Dilution

1:500, 1 hour at RT

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

Western Blot Analysis by WBK-012-22

Lane #1: Porcine Pancreas Extracts
Lane #2: Bovine Hypothalamus Extracts Lane
#3: Bovine Pituitary Extracts Lane
#4: Porcine Liver Extracts
The major band in the porcine pancreas is 14.2 kDa augurin, but the bovine hypothalamus and pituitary show augurin derived peptides with the size at 14.1, 9.6 kDa and 6.5 kDa which can be detected by anti-augurin (71-107) antibody.

Standard Curve of Augurin, Prepro (71-107) (human) EIA Kit (EK-012-22)

Sensitivity

Minimum Detectable Concentration: 0.16ng/ml

Range

0.01-100ng/ml

Linear Range

0.16-1.46ng/ml

Precision

Intra-assay variation: <10%
Inter-assay variation:<15%

Cross-Reactivity
Prepro-Augurin (71-107) (H) 100
Prepro-Augurin (108-132) (H)
0
∆16 / Prepro-Augurin (133-148) (H)

0

augurin RIA;new_augurin;32-EPS-Augurin-Poster

%Augurin%


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