Discovering differential protein expression caused by CagA-induced ERK pathway activation in AGS cells using the SELDI-ProteinChip platform

doi:10.3748/wjg.14.554

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World Journal of Gastroenterology

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1007-9327

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Baishideng Publishing Group Inc, 7041 Koll Center Parkway, Suite 160, Pleasanton, CA 94566, USA

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World J Gastroenterol. Jan 28, 2008; 14(4): 554-562
Published online Jan 28, 2008. doi: 10.3748/wjg.14.554

Table 1 Overview of biomarkers’ expression level in the 2 000-20 000 m/z mass range in wild-type cagA, phosphorylation site mutant cagA and blank vector transfected AGS cells

Array type	m/z	Peak intensity (mean ± SD)			Fold change		Comments
Array type	m/z	V	W	M	W vs V	M vs V	Comments
SAX2	2013	30.39 ± 7.19	8.82 ± 5.37	29.92 ± 9.53	-3.45^a	-1.02	CagA phosphorylation dependent
	4286	5.08 ± 0.85	2.25 ± 0.70	4.44 ± 0.58	-2.26^a	-1.14
	8564	9.73 ± 1.88	3.72 ± 0.42	10.86 ± 1.77	-2.62^a	1.12
	9084	2.61 ± 1.35	6.13 ± 1.74	2.75 ± 0.60	2.35^a	1.05
	9952	5.22 ± 2.07	2.51 ± 0.95	5.19 ± 1.30	-2.08^a	-1.01
	11 643	3.39 ± 0.98	1.32 ± 0.86	3.70 ± 1.10	-2.57^a	1.09
CM10	4229	4.29 ± 0.57	9.36 ± 0.32	6.13 ± 0.78	2.18^a	1.43
	5130	3.59 ± 0.72	6.93 ± 0.61	4.58 ± 0.46	1.93^a	1.28
	8162	4.26 ± 0.38	8.88 ± 1.06	4.97 ± 0.24	2.08^a	1.17
	11 087	3.37 ± 0.75	1.94 ± 0.46	3.19 ± 0.39	-1.74^a	-1.06
	13 804	2.97 ± 0.79	8.66 ± 0.48	4.14 ± 1.23	2.92^a	1.39
	14 021	3.06 ± 0.24	5.59 ± 0.41	3.66 ± 1.04	1.83^a	1.20
SAX2	6545	3.08 ± 0.88	6.95 ± 1.34	5.53 ± 0.69	2.26^a	1.80^a	CagA phosphorylation independent
CM10	4714	10.16 ± 1.26	17.95 ± 3.61	17.31 ± 1.41	1.77^a	1.70^a	CagA phosphorylation independent
	4729	11.11 ± 1.75	20.51 ± 4.02	21.92 ± 2.58	1.80^a	1.97^a
	6656	9.67 ± 2.45	22.83 ± 4.27	21.46 ± 1.92	2.36^a	2.22^a

Biomarkers were selected by these criteria: Statistical P-value among sample groups less than 0.05; signal to noise ratio higher than 5.0; peak intensity higher than 1.0. The particular fold change in average peak intensity of wild-type cagA transfectants (W) or phosphorylation site mutant cagA transfectants (M) was calculated compared with the intensity of blank vector transfectants (V).

^aP < 0.05. Statistical P-value was calculated with a non-parametrical Mann-Whitney test (n = 4).

Table 2 Comparison of biomarkers’ expression level under mitogen-activated protein kinase kinase (MEK) inhibitor free, treated and DMSO treated conditions

Array	m/z	Inhibitor free		Inhibitor treated		DMSO treated		Comments
type	m/z	Fold change (W vs V)	P-value	Fold change (W+I vs V+I)	P-value	Fold change (W+D vs V+D)	P-value	Comments
SAX2	9084	1.64	0.021	1.00	0.827	2.21	0.021	Caused by ERK1/2 activation
CM10	4229	2.25	0.021	1.07	1.000	2.44	0.021	Caused by ERK1/2 activation
	8162	1.71	0.021	-1.15	0.149	1.81	0.021
SAX2	2013	-1.99	0.021	1.16	0.149	1.03	0.773	Have no relationship with ERK1/2 activation
	4286	-3.50	0.021	-2.14	0.021	-1.95	0.021
	6545	2.86	0.021	2.39	0.021	2.00	0.021
	8564	-3.05	0.021	-2.96	0.021	-1.85	0.021
	9952	-3.35	0.021	-2.87	0.021	-2.62	0.021
	11 643	-1.71	0.021	-2.49	0.021	-2.38	0.021
CM10	4714	1.89	0.021	1.68	0.043	1.74	0.021
	4729	1.99	0.021	1.72	0.021	1.86	0.021
	5130	1.67	0.021	1.66	0.021	2.91	0.021
	6656	2.12	0.021	2.31	0.021	2.44	0.021
	11 087	-1.72	0.021	-1.85	0.021	-1.52	0.021
	13 804	2.23	0.021	2.56	0.021	2.60	0.021
	14 021	1.98	0.021	1.79	0.021	1.61	0.021

Biomarkers listed in top three lanes were identified as biomarkers induced by extracellular signal-regulated kinase 1/2 (ERK1/2) activation. Columns 3, 5, 7 give fold changes in peak intensity in wild-type cagA transfectants (W) compared with vector transfectants (V) as reference under MEK inhibitor U0126 free, MEK inhibitor U0126 treated (I) and DMSO treated (D) conditions. Statistical P-value were calculated using a non-parametrical Mann-Whitney test (n = 4).

Table 3 Swiss-Prot and TrEMBL database searching results of the three ERK related biomarkers

Biomarker	Meaningful matching information in Swiss-Prot and TrEMBL database			Comments
(Mw)	Protein Name	Mw	pI	Comments
9084 (SAX2)	BIM-alpha3 (BCL2-like 11 transcript variant 10, chain 1-79)	9081	5.42	Over-expression of BIM-alpha3 results in cell apoptosis
	EP2E (chain 1-80)	9091	7.55	Part of the protein sequences homologous to β-defensins, a family of antimicrobial peptides.
	UPF0197 protein (C11orf10 chain 1-79)	9079	5.58	Expressed in CD34+ hematopoietic stem/progenitor cells
4229 (CM10)	Chromogranin-A Precursor (ER-37, chain 420-456)	4233	4.59	Neuroendocrine and endocrine secretory granules, considered as biomarkers in some human cancers
8162 (CM10)	Small-inducible cytokine B6 Precursor (chain 40-114)	8162	9.74	Functional characterization as chemokines, chemotactic for neutrophil granulocytes
	Putative microRNA host gene protein 1 (chain 1-70)	8163	9.25	Highly expressed in B-cell lymphoma and lung cancer and enhanced cell proliferation
	PNAS-145 (Chain: 1-68)	8161	11.24	Human acute promyelocytic leukemia cell line NB4’s apoptosis/differentiation related genes

Searching was carried out using TagIdent tool with mass and the pI information (0-9.0 for SAX2, 4.0-14.0 for CM10).

Citation: Ge Z, Zhu YL, Zhong X, Yu JK, Zheng S. Discovering differential protein expression caused by CagA-induced ERK pathway activation in AGS cells using the SELDI-ProteinChip platform. World J Gastroenterol 2008; 14(4): 554-562
URL: https://www.wjgnet.com/1007-9327/full/v14/i4/554.htm
DOI: https://dx.doi.org/10.3748/wjg.14.554