👤 Guadalupe Reyes-Cruz

Calcium-sensing receptor (CaSR), a G protein-coupled receptor, is overexpressed in certain breast cancer tumors where it drives cell migration and secretion of chemotactic agonists, likely contributing to metastatic dissemination. Since CaSR activates breast cancer cell migration via the Gβγ-PI3K-mTORC2/Rac-1 pathway, we hypothesized that PKCζ and perhaps other protein kinase C (PKC) isoforms, known as mTORC2-regulated effectors, are involved in migratory and invasive signaling elicited by CaSR. We analyzed the effect of PKC inhibitors and siRNAs which pointed to PKCζ as effector of CaSR in cell migration and invasion. In breast cancer phosphoproteomic CPTAC datasets, we identified a group of Luminal A subtype cancer patients having active PKCζ, according to its phosphorylation status at the turn motif. In addition, various phosphorylated RacGEFs, including TRIO, ARHGEF26, DOCK1 and DOCK7, clustered as phosphoproteins with active PKCζ. We therefore introduce atypical PKCζ as an effector component of the CaSR-Gβγ-PI3K-mTORC2 pathway in the activation of the promigratory small GTPase Rac. These results support ongoing initiatives to establish critical elements of the CaSR signaling pathway as potential targets in metastatic breast cancer. Show less

Rho guanine nucleotide exchange factors (RhoGEFs) integrate cell signaling inputs into morphological and functional responses. However, little is known about the endothelial repertoire of RhoGEFs and their regulation. Thus, we assessed the expression of 81 RhoGEFs (70 homologous to Dbl and 11 of the DOCK family) in endothelial cells. Further, in the case of DH-RhoGEFs, we also determined their responses to VEGF exposure in vitro and in the context of tumors. A phylogenetic analysis revealed the existence of four groups of DH-RhoGEFs and two of the DOCK family. Among them, we found that the most abundant endothelial RhoGEFs were: Tuba, FGD5, Farp1, ARHGEF17, TRIO, P-Rex1, ARHGEF15, ARHGEF11, ABR, Farp2, ARHGEF40, ALS, DOCK1, DOCK7 and DOCK6. Expression of RASGRF2 and PREX2 increased significantly in response to VEGF, but most other RhoGEFs were unaffected. Interestingly murine endothelial cells isolated from tumors showed that all four phylogenetic subgroups of DH-RhoGEFs were altered when compared to non-tumor endothelial cells. In summary, our results provide a detailed assessment of RhoGEFs expression profiles in the endothelium and set the basis to systematically address their regulation in vascular signaling. Show less