Mouse Mammary Tumors Express Elevated Levels of RNA Encoding the Murine Homolog of SKY, a Putative Receptor Tyrosine Kinase

doi:10.1074/jbc.270.12.6872

Mouse Mammary Tumors Express Elevated Levels of RNA Encoding the Murine Homolog of SKY, a Putative Receptor Tyrosine Kinase (*)

From the (1)Department of Microbiology and Immunology, University of California, San Francisco, California 94143,
(2)NCI, National Institutes of Health, Bethesda, Maryland 20892, and the
(3)Lawrence Berkeley Laboratory, University of California, Berkeley, California 94720

^¶¶ Formerly an American Cancer Society Research Professor. To whom correspondence should be addressed:
NCI, National Institutes of Health, Bldg. 49, Rm. 4A56, 9000 Rockville Pike, Bethesda, MD 20892.
Tel.: 301-496-0429; Fax: 301-496-0332.

Abstract

To gain insight into the signal transduction pathways utilized by the Wnt-1-responsive mammary epithelial cell line C57MG, we screened for non-src family member tyrosine kinases expressed in these cells using a polymerase chain reaction-based technique. We identified five cDNA clones encoding receptor tyrosine kinases for which the ligand is known (fibroblast growth factor receptor, platelet-derived growth factor receptor, epithelial growth factor receptor, insulin receptor, and insulin-like growth factor receptor), two putative receptor tyrosine kinases for which the ligand remains to be identified (the products of ryk and the mouse klg homolog), and a novel tyrosine kinase. We cloned cDNAs encoding both the murine and human homologs of this kinase, the sequences of which were subsequently published under the names sky (Ohashi, K., Mizuno, K., Kuma, K., Miyata, T., and Nakamura, T.(1994) Oncogene 9, 699-705) and rse (Mark, M. R., Scadden, D. T., Wang, Z., Gu, Q., Goddard, A., and Godowski, P. J.(1994) J. Biol. Chem. 269, 10720-10728). Mouse sky RNA levels are abundant in mammary tumors derived from transgenic mice that express wnt-1, fgf-3, or both oncogenes in their mammary glands. However, little or no expression of sky is detected in mammary glands from virgin animals or in preneoplastic mammary glands from wnt-1 transgenic mice. Moreover, we find that the human homolog of sky is expressed at elevated levels when normal human mammary epithelial cells are rendered tumorigenic by the introduction of two viral oncogenes. Transient transfection of the human SKY cDNA into the quail fibrosarcoma cell line QT6 reveals that SKY is an active tyrosine kinase that augments the level of cellular phosphotyrosine. Introduction of murine Sky into RatB1a fibroblasts by retrovirus-mediated gene transfer results in morphological transformation, growth in soft agar, and the formation of tumors in nude mice. These data raise the possibility that the Sky tyrosine kinase is involved in the development and/or progression of mammary tumors.

Footnotes

↵^§ A Damon Runyan-Walter Winchell Cancer Research Fund Fellow.
↵^¶ Contributed equally to this work.
↵** Supported by a Centennial Fellowship from the Medical Research Council of Canada.
↵^§§ Received support from National Institutes of Health Grants CA-54247 and CA-24844 as well as from the Office of Energy Research, Office of Health and Environmental Research, U. S. Dept. of Energy under Contract DE-AC0376SF00098.
↵* This work was supported by National Institutes of Health Grant CA-39832. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

PTKs

protein tyrosine kinases

PCR

polymerase chain reaction

HA

hemagglutinin

FBS

fetal bovine serum

MEF

mouse embryo fibroblasts

HMEC

human mammary epithelial cells

DME media

Dulbecco's minimal essential media

bp

base pair(s)

kb

kilobase(s)

MOPS

4-morpholinepropanesulfonic acid

PAGE

polyacrylamide gel electrophoresis

BSA

bovine serum albumin

GAPDH

glyceraldehyde-3-phosphate dehydrogenase.
- Received October 28, 1994.