Lee R C, Walters J A, Reyland M E, Anderson S M
Department of Pathology, School of Medicine, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
J Biol Chem. 1999 Apr 9;274(15):10024-34. doi: 10.1074/jbc.274.15.10024.
The ability to induce the oncogenic activation of the human prolactin receptor (PRLR) was examined by deleting 178 amino acids of the extracellular ligand-binding domain. Expression of this deletion mutant in the interleukin-3 (IL-3)-dependent murine myeloid cell line 32Dcl3 resulted in the induction of growth factor-independent proliferation. Parental 32Dcl3 cells proliferated only in the presence of exogenous murine IL-3 (mIL-3), while 32Dcl3 cells transfected with the long form of the human PRLR were able to proliferate in response to mIL-3, ovine prolactin, or human PRL. Cells expressing the Delta178 deletion mutant contained numerous phosphotyrosine-containing proteins in the absence of stimulation with either mIL-3 or ovine prolactin. Growth factor stimulation increased the number of proteins phosphorylated and the intensity of phosphorylation. These proteins included constitutively phosphorylated Janus kinase 2, signal transducer and activator of transcription 5, and SHC. Activated extracellular signal-regulated kinases 1 and 2 (ERK1 and ERK2) were observed in unstimulated 32Dcl3 cells expressing the Delta178 mutant. Likewise, transfection of Nb2 cells with the Delta178 deletion mutant induced growth factor-independent proliferation and constitutive activation of Janus kinase 2, ERK1, and ERK2. In addition to the induction of a growth factor-independent state, the expression of the Delta178 deletion mutant also suppressed the apoptosis that occurs when 32Dcl3 cells are cultured in the absence of growth factors such as IL-3. These data suggest that the constitutive activation of the PRLR can be achieved by deletion of the ligand binding domain and that this mutation leads to the oncogenic activation of the receptor as determined by the ability of the receptor to induce growth factor-independent proliferation of factor-dependent hematopoietic cells.
