Cloning and expression of JE, a gene inducible by platelet-derived growth factor and whose product has cytokine-like properties.

Abstract

The platelet-derived growth factor-inducible gene JE has been widely used as a molecular marker for the cellular response to growth factors, antimitogenic agents, and other biological response modifiers; however, the structure of the JE gene and the nature of its encoded protein have not been previously described. We present here structural and regulatory features of the JE gene and its product that link it to a family of cytokines, including macrophage colony-stimulating factor, interferon a, interleukin 6 (also known as interferon I2, B-cell-stimulatory factor 2, 26-kDa protein, and hybridoma/ plasmacytoma growth factor), and interleukin 2. Just as T lymphocytes secrete interleukins as a component of their response to mitogens, it appears that fibroblasts secrete cytokines as a component of their response to platelet-derived growth factor.In confluent growth-arrested BALB/c 3T3 cells, plateletderived growth factor (PDGF) stimulates expression of a family of single-copy genes. The protooncogenes c-myc and c-fos are members of this gene family (1-5). However the PDGF-inducible genes detected and characterized as such were the "competence" genes, JE, KC, and JB. Partial cDNA clones for these three genes were isolated by differential screening of a cDNA library from PDGF-treated fibroblasts (6). The c-myc (7-12) and c-fos (13,14) bovine calf serum and antibiotics. Confluent density-arrested monolayer cell cultures were prepared as previously described (20) and then transferred to fresh DMEM supplemented with 5% platelet-poor plasma, which is free of PDGF (21). Cells were kept in platelet-poor plasma for 16 hr prior to analysis.Due to growing safety concerns about processing thousands of units of clinically outdated human platelets each year, our laboratory now uses medium conditioned by v-sis-transformed NRK (normal rat kidney) cells (sisconditioned medium) as the routine source of PDGF (B chain homodimer). All experimental results generated with sisconditioned medium have been confirmed with either homogeneous PDGF from human platelets or recombinant v-sis protein (Amgen Biologicals, Thousand Oaks, CA). Recombinant human interleukin la (IL-la) was obtained from Genzyme (Norwalk, CT).Gene and cDNA Cloning. For the isolation of JE genomic DNA, 106 plaques from the BALB/c germ line DNA library of Davis et al. (22) were transferred to nitrocellulose and probed as described (23). The probe was the 750-base-pair (bp) Pst I fragment of pBC-JE (6) nick-translated to 108 cpm/pg. Four hybridizing clones (AJE-1, AJE-2, AJE-3, and AJE-4) were identified and subcloned in pGEM-1 (Promega Biotec, Madison, WI) for sequence analysis. For the isolation of JE cDNA, poly(A)-selected RNA was prepared from BALB/c 3T3 cells that had been treated with PDGF (v-sis conditioned medium) for 2 hr. cDNA was synthesized by the RNase H method (24) and cloned in AgtlO. About 106 plaques were screened as above. Seventy-eight hybridizing plaques were identified, three of which (cJE-1, cJE-2, and cJE-3) were subcloned in pGEM-1. Sequence ...

Human monocyte chemoattractant protein‐1 (MCP‐1) Full‐length cDNA cloning, expression in mitogen‐stimulated blood mononuclear leukocytes, and sequence similarity to mouse competence gene JE

“…TCA 3: from ConA-stimulated mouse T cell line [23]. JE: from platelet-derived growth factor (PDGF)-stimulated mouse fibroblast line [24]. MIP: from culture fluid of LPS-stimulated mouse macrophage cell line [25,26].…”

Regulation of connective tissue growth factor gene expression in human skin fibroblasts and during wound repair.

“…One category of these genes includes c-fos and c-myc, whose products are nuclear proteins binding to DNA and regulating the transcription of other genes controlling for cell growth and replication (Kelley et al, 1983;Greenberg and Ziff, 1984;Kruijer et al, 1984;Muller et al, 1984). The other group encodes secreted peptides with cytokine characteristics such as the KC/gro or JE genes Rollins et al, 1988;Oquendo et al, 1989). The CTGF gene belongs to the latter category because its protein product is secreted and possesses chemotactic and mitogenic activities for connective tissue cells.…”

Low-Level Monocyte Chemoattractant Protein-1 Stimulation of Monocytes Leads to Tumor Formation in Nontumorigenic Melanoma Cells

“…Expression has been reported for melanoma (13), glioma (14,15), sarcoma (16,17), leukemia (18), hemangioma (19), and carcinomas of breast (20), cervix (21,22), and ovary (23). The malignant cells express MCP-1, apparently due to the constitutive production of activating growth factors and cytokines such as IL-1 (24), TGF-␤ (25), and platelet-derived growth factor (26,27). MCP-1 can be protective in some tumor models but destructive in others; murine colon carcinoma cells expressing MCP-1 fail to metastasize when injected into mice (28), whereas other carcinoma cells show enhanced metastasis (29).…”