Involvement of Lyn and the Atypical Kinase SgK269/PEAK1 in a Basal Breast Cancer Signaling Pathway

Abstract

Basal breast cancer cells feature high expression of the Src family kinase Lyn that has been implicated in the pathogenicity of this disease. In this study, we identified novel Lyn kinase substrates, the most prominent of which was the atypical kinase SgK269 (PEAK1). In breast cancer cells, SgK269 expression associated with the basal phenotype. In primary breast tumors, SgK269 overexpression was detected in a subset of basal, HER2-positive, and luminal cancers. In immortalized MCF-10A mammary epithelial cells, SgK269 promoted transition to a mesenchymal phenotype and increased cell motility and invasion. Growth of MCF-10A acini in three-dimensional (3D) culture was enhanced upon SgK269 overexpression, which induced an abnormal, multilobular acinar morphology and promoted extracellular signal-regulated kinase (Erk) and Stat3 activation. SgK269 Y635F, mutated at a major Lyn phosphorylation site, did not enhance acinar size or cellular invasion. We show that Y635 represents a Grb2-binding site that promotes both Stat3 and Erk activation in 3D culture. RNA interferencemediated attenuation of SgK269 in basal breast cancer cells promoted acquisition of epithelial characteristics and decreased anchorage-independent growth. Together, our results define a novel signaling pathway in basal breast cancer involving Lyn and SgK269 that offers clinical opportunities for therapeutic intervention.

Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

“…This suggests that these proteins are paralogs, and the three homologous regions undertake critical conserved functions. Previously, we demonstrated that overexpression of SgK269 in MCF-10A-immortalized mammary epithelial cells perturbed cell phenotype and signaling in a manner consistent with SgK269 functioning as a breast cancer oncogene (11). To interrogate the function of specific regions of SgK269, we generated a series of deletion mutants lacking the N-terminal segment (⌬N), the CH region (⌬CH), and the pseudokinase domain (⌬PK) (Fig.…”

“…SgK269 Y1188 binds Shc1 and mediates a key switch in EGF receptor signal output over time, from mitogenic/survival signaling to that regulating morphogenesis (10). In addition, SgK269 couples to the Ras signaling pathway via Tyr-635-mediated binding to the Grb2 SH2 domain (11). Overexpression of SgK269 in mammary epithelial cells promotes a partial epithelial-mesenchyme transition and aberrant growth and morphogenesis in 3D Matrigel culture, identifying SgK269 as a potential breast cancer oncogene (11).…”

“…In addition, SgK269 couples to the Ras signaling pathway via Tyr-635-mediated binding to the Grb2 SH2 domain (11). Overexpression of SgK269 in mammary epithelial cells promotes a partial epithelial-mesenchyme transition and aberrant growth and morphogenesis in 3D Matrigel culture, identifying SgK269 as a potential breast cancer oncogene (11). A likely oncogenic role for this pseudokinase is further underscored by its overexpression in breast, pancreatic, and colon cancers (9,11,12).…”

“…Overexpression of SgK269 in mammary epithelial cells promotes a partial epithelial-mesenchyme transition and aberrant growth and morphogenesis in 3D Matrigel culture, identifying SgK269 as a potential breast cancer oncogene (11). A likely oncogenic role for this pseudokinase is further underscored by its overexpression in breast, pancreatic, and colon cancers (9,11,12). Interestingly, SgK269 exhibits a conserved molecular architecture and sequence similarity with a second pseudokinase, SgK223, the human ortholog of rat Pragmin (4,13), suggesting that SgK269 and SgK223 are paralogs.…”

Phosphoproteomic Profiling Identifies Focal Adhesion Kinase as a Mediator of Docetaxel Resistance in Castrate-Resistant Prostate Cancer

“…Quantitative tyrosine phosphorylation profiling of docetaxel-sensitive and resistant cells was undertaken by immunoaffinity purification followed by liquid chromatography/tandem mass spectrometry (LC/MS-MS) in combination with stable isotope labeling with amino acids in cell culture (SILAC), as previously described (19,20).…”