Differential Expression of Ccn4 and Other Genes Between Metastatic and Non-metastatic EL4 Mouse Lymphoma Cells

Abstract

EL4 is a murine lymphoma cell line developed in 1945 by treating C57 black mice with 9:10-dimethyl-1:2-benzanthracene (1). The cells were originally propagated in animal hosts, prior to adaptation to cell culture. EL4 cells were used for many years as a source of interleukin-2 (IL2), which they secrete when treated with phorbol 12-myristate 13-acetate (PMA).The EL4 cell line originally provided to us, by a colleague at the University of Washington, was heterogeneous with respect to PMA response. We, therefore, developed and characterized EL4 sub-lines. Wild-type (WT)-derived cell lines, which are PMA responsive, grow readily in suspension culture as did the original strain. Variant (V)-derived (PMA-resistant) sub-lines were selected for enhanced adhesion to tissue culture plastic (2). Clonal lines were developed from both cell types by limiting dilution (3). The PMA sensitivity of WT-derived cells, as reflected by PMA-induced IL2 production and mitogen-activated protein kinase (ERK1/2) activation, has been attributed to expression of Ras guanyl nucleotide releasing protein 1 (RasGRP1), which binds PMA and directly activates Ras (4); V-derived cells do not express RasGRP1.Clonal EL4 cell lines were used for experimental metastasis studies in syngeneic mice, with WT2 and V7 as prototypes (5, 6). WT2 cells do not form tumors (nonmetastatic), while V7 cells form liver tumors (metastatic) after tail vein injection. 'Metastasis' more strictly refers to tumorigenesis in this model, since circulating EL4 cells home to the liver to form tumors (5). The clonal C5 cell line was developed after stably overexpressing human hemagllutinin (HA)-tagged phospholipase D2 (PLD2) in V7 cells in order to characterize the signaling role of PLD2 using a cell line expressing little or no endogenous PLD2 (5). As compared to V7, C5 cells exhibit increased cell migration (7) and tumor growth (5), providing an early example of the positive role of PLD2 in tumorigenesis.This report presents data from microarray analyses comparing transcripts expressed by these three EL4 cell types, with the goal of further defining their phenotypes.

The role of CCN4/WISP-1 in the cancerous phenotype

“…In 2016, we published the results of an Affymetrix microarray analysis comparing gene expression between metastatic and nonmetastatic EL4 cell lines. 60 Some of the results recapitulated those we had already published previously studying the levels of expression of signal transduction proteins. 57 , 58 However, to our surprise, WISP-1 emerged as a new protein of interest, being one of the top-ranked genes with respect to differences in mRNA expression.…”

“…The meta-static/adherent phenotype is characterized by expression of FAK, a tyrosine kinase that regulates focal adhesion function. In 2016, we published the results of an Affymetrix microarray analysis comparing gene expression between metastatic and nonmetastatic EL4 cell lines 60. Some of the results recapitulated those we had already published previously studying the levels of expression of signal transduction proteins 57,58.…”

Type 2 Cystatins and Their Roles in the Regulation of Human Immune Response and Cancer Progression

“…Upregulation of CST7 causes a decrease in the cytotoxicity of NK cells to tumor cells [168]. In contrast, multiple studies suggest that CST7 downregulation is associated with higher invasiveness of tumors, more metastatic events, and tumor progression in prostate cancer, lung cancer, pancreatic cancer, and lymphoma [170][171][172][173]. Since CST7 downregulation is correlated with metastasis and tumor progression, this indicates that CST7, like other type 2 cystatin family members, regulates the cathepsin B-L metastatic axis.…”

Tp53 Mutation Inhibits Ubiquitination and Degradation of WISP1 via Down-Regulation of Siah1 in Pancreatic Carcinogenesis

“…WISP1 is a matricellular protein and plays a significant role in regulation of cellular signaling networks ( Berschneider and Konigshoff, 2011 ). Recently, abnormal expression of WISP1 has been proven in various types of human malignancies ( Gurbuz and Chiquet-Ehrismann, 2015 ; Chahal et al, 2016 ; Wu et al, 2016 ; Jing et al, 2017 ). A previous study demonstrated that WISP1 protects human lung and breast cancer cells from p53-dependent cell death, suggesting that there could be a crosstalk between Tp53 and WISP1 signaling pathways ( Su et al, 2002 ).…”