Excess PLAC8 promotes an unconventional ERK2-dependent EMT in colon cancer

Abstract

The epithelial-to-mesenchymal transition (EMT) transcriptional program is characterized by repression of E-cadherin (CDH1) and induction of N-cadherin (CDH2), and mesenchymal genes like vimentin (VIM). Placenta-specific 8 (PLAC8) has been implicated in colon cancer; however, how PLAC8 contributes to disease is unknown, and endogenous PLAC8 protein has not been studied. We analyzed zebrafish and human tissues and found that endogenous PLAC8 localizes to the apical domain of differentiated intestinal epithelium. Colon cancer cells with elevated PLAC8 levels exhibited EMT features, including increased expression of VIM and zinc finger E-box binding homeobox 1 (ZEB1), aberrant cell motility, and increased invasiveness. In contrast to classical EMT, PLAC8 overexpression reduced cell surface CDH1 and upregulated P-cadherin (CDH3) without affecting CDH2 expression. PLAC8-induced EMT was linked to increased phosphorylated ERK2 (p-ERK2), and ERK2 knockdown restored cell surface CDH1 and suppressed CDH3, VIM, and ZEB1 upregulation. In vitro, PLAC8 directly bound and inactivated the ERK2 phosphatase DUSP6, thereby increasing p-ERK2. In a murine xenograft model, knockdown of endogenous PLAC8 in colon cancer cells resulted in smaller tumors, reduced local invasion, and decreased p-ERK2. Using MultiOmyx, a multiplex immunofluorescence-based methodology, we observed coexpression of cytosolic PLAC8, CDH3, and VIM at the leading edge of a human colorectal tumor, supporting a role for PLAC8 in cancer invasion in vivo.

PLAC8 Localizes to the Inner Plasma Membrane of Pancreatic Cancer Cells and Regulates Cell Growth and Disease Progression through Critical Cell-Cycle Regulatory Pathways

“…Moreover, PLAC8 overexpression resulted in induction, rather than inhibition, of apoptosis in both primary and transformed human lymphocytes (11), and decreased PLAC8 expression was reported to be associated with oncogenesis in the liver (34). Further evidence for context-dependent differential roles of PLAC8 comes from a study by Li and colleagues (13), who demonstrated that PLAC8 overexpression caused decreased migration of mesenchymal cells in zebrafish embryos, while resulting in enhanced migration and features reminiscent of epithelial-to-mesenchymal transition in cultured colon cancer cells.…”

“…Although evolutionarily highly conserved, the protein does not contain any known functional domains, and its molecular function remains enigmatic. Moreover, the cellular role of PLAC8 seems to be highly contextdependent, resulting in strongly variable, and sometimes contradictory, effects of PLAC8 modulation in different cell types, ranging from apoptosis protection in fibroblasts (10) versus apoptosis induction in human lymphocytes (11) to inhibition of cell differentiation in primary acute myeloid leukemia cells (12), and induction of epithelial-to-mesenchymal transition in cultured colon cancer cells (13).…”

Zinc finger protein X-linked is overexpressed in colorectal cancer and is associated with poor prognosis

“…In fact, in these genes, signaling pathways along with certain other pathways, including wnt/β-catenin, play essential roles in CRC progression (22)(23)(24)(25)(26). The correlation between ZFX and these genes and pathways in CRC progression requires further study.…”

Opportunities and Challenges in Implementation of Multiparameter Single Cell Analysis Platforms for Clinical Translation

“…Mutually exclusive phosphorylation patterns of the two canonical substrates of mTORC1 (S6K1 and 4E‐BP1) were observed in individual cells, large regions of most tumors, and in distinct cell lineages, demonstrating differential pathway activation. In the research setting, MxIF has also been applied to evaluate signaling pathway changes in mouse salivary glands,26 estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), and Ki67 colocalization in breast cancer,27 and other single‐cell analyses 28, 29…”

“…Multiplex protein expression (4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37) …”