Zengjin Yuan

Adhesion G-protein-coupled receptors (GPCR), which contain adhesion domains in their extracellular region, have been found to play important roles in cell adhesion, motility, embryonic development, and immune response. Because most adhesion molecules with adhesion domains have vital roles in cancer metastasis, we speculated that adhesion GPCRs are potentially involved in cancer metastasis. In this study, we identified GPR116 as a novel regulator of breast cancer metastasis through expression and functional screening of the adhesion GPCR family. We found that knockdown of GPR116 in highly metastatic (MDA-MB-231) breast cancer cells suppressed cell migration and invasion. Conversely, ectopic GPR116 expression in poorly metastatic (MCF-7 and Hs578T) cells promoted cell invasion. We further showed that knockdown of GPR116 inhibited breast cancer cell metastasis in two mammary tumor metastasis mouse models. Moreover, GPR116 modulated the formation of lamellipodia and actin stress fibers in cells in a RhoA- and Rac1-dependent manner. At a molecular level, GPR116 regulated cell motility and morphology through the Gαq-p63RhoGEF-RhoA/Rac1 pathway. The biologic significance of GPR116 in breast cancer is substantiated in human patient samples, where GPR116 expression is significantly correlated with breast tumor progression, recurrence, and poor prognosis. These findings show that GPR116 is crucial for the metastasis of breast cancer and support GPR116 as a potential prognostic marker and drug target against metastatic human breast cancer.

The fourth member of the leucine-rich repeat-containing GPCR family (LGR4, frequently referred to as GPR48) and its cognate ligands, R-spondins (RSPOs) play crucial roles in the development of multiple organs as well as the survival of adult stem cells by activation of canonical Wnt signaling. Wnt/β-catenin signaling acts to regulate breast cancer; however, the molecular mechanisms determining its spatiotemporal regulation are largely unknown. In this study, we identified LGR4 as a master controller of Wnt/β-catenin signaling-mediated breast cancer tumorigenesis, metastasis, and cancer stem cell (CSC) maintenance. LGR4 expression in breast tumors correlated with poor prognosis. Either Lgr4 haploinsufficiency or mammary-specific deletion inhibited mouse mammary tumor virus (MMTV)- PyMT- and MMTV- Wnt1-driven mammary tumorigenesis and metastasis. Moreover, LGR4 down-regulation decreased in vitro migration and in vivo xenograft tumor growth and lung metastasis. Furthermore, Lgr4 deletion in MMTV- Wnt1 tumor cells or knockdown in human breast cancer cells decreased the number of functional CSCs by ∼90%. Canonical Wnt signaling was impaired in LGR4-deficient breast cancer cells, and LGR4 knockdown resulted in increased E-cadherin and decreased expression of N-cadherin and snail transcription factor -2 ( SNAI2) (also called SLUG), implicating LGR4 in regulation of epithelial-mesenchymal transition. Our findings support a crucial role of the Wnt signaling component LGR4 in breast cancer initiation, metastasis, and breast CSCs.-Yue, Z., Yuan, Z., Zeng, L., Wang, Y., Lai, L., Li, J., Sun, P., Xue, X., Qi, J., Yang, Z., Zheng, Y., Fang, Y., Li, D., Siwko, S., Li, Y., Luo, J., Liu, M. LGR4 modulates breast cancer initiation, metastasis, and cancer stem cells.

Many methods, including parametric, nonparametric, and Bayesian methods, have been used for detecting differentially expressed genes based on the assumption that biological systems are linear, which ignores the nonlinear characteristics of most biological systems. More importantly, those methods do not simultaneously consider means, variances, and high moments, resulting in relatively high false positive rate. To overcome the limitations, the SWang test is proposed to determine differentially expressed genes according to the equality of distributions between case and control. Our method not only latently incorporates functional relationships among genes to consider nonlinear biological system but also considers the mean, variance, skewness, and kurtosis of expression profiles simultaneously. To illustrate biological significance of high moments, we construct a nonlinear gene interaction model, demonstrating that skewness and kurtosis could contain useful information of function association among genes in microarrays. Simulations and real microarray results show that false positive rate of SWang is lower than currently popular methods (T-test, F-test, SAM, and Fold-change) with much higher statistical power. Additionally, SWang can uniquely detect significant genes in real microarray data with imperceptible differential expression but higher variety in kurtosis and skewness. Those identified genes were confirmed with previous published literature or RT-PCR experiments performed in our lab.