Musashi-2 (MSI2) supports TGF-β signaling and inhibits claudins to promote non-small cell lung cancer (NSCLC) metastasis

Abstract

Non-small cell lung cancer (NSCLC) has a 5-y survival rate of ∼16%, with most deaths associated with uncontrolled metastasis. We screened for stem cell identity-related genes preferentially expressed in a panel of cell lines with high versus low metastatic potential, derived from NSCLC tumors of Kras LA1/+ ;P53 R172HΔG/+ (KP) mice. The Musashi-2 (MSI2) protein, a regulator of mRNA translation, was consistently elevated in metastasis-competent cell lines. MSI2 was overexpressed in 123 human NSCLC tumor specimens versus normal lung, whereas higher expression was associated with disease progression in an independent set of matched normal/primary tumor/lymph node specimens. Depletion of MSI2 in multiple independent metastatic murine and human NSCLC cell lines reduced invasion and metastatic potential, independent of an effect on proliferation. MSI2 depletion significantly induced expression of proteins associated with epithelial identity, including tight junction proteins [claudin 3 (CLDN3), claudin 5 (CLDN5), and claudin 7 (CLDN7)] and down-regulated direct translational targets associated with epithelial-mesenchymal transition, including the TGF-β receptor 1 (TGFβR1), the small mothers against decapentaplegic homolog 3 (SMAD3), and the zinc finger proteins SNAI1 (SNAIL) and SNAI2 (SLUG). Overexpression of TGFβRI reversed the loss of invasion associated with MSI2 depletion, whereas overexpression of CLDN7 inhibited MSI2-dependent invasion. Unexpectedly, MSI2 depletion reduced E-cadherin expression, reflecting a mixed epithelialmesenchymal phenotype. Based on this work, we propose that MSI2 provides essential support for TGFβR1/SMAD3 signaling and contributes to invasive adenocarcinoma of the lung and may serve as a predictive biomarker of NSCLC aggressiveness.on-small cell lung cancer (NSCLC) is the leading cause of cancer-related deaths in the world (1). Approximately 7% of individuals born in the United States in 2013 will ultimately be diagnosed with lung cancer, and 160,000 die from this disease each year (1). The 5-y survival rate for lung cancer is around 16% of diagnosed cases (2). Much of the lethality of lung cancer is due to frequent diagnosis of the malignancy at the metastatic stage, when fundamental changes in tumor biology cause the disease to be refractory to many treatments. A better understanding of the biological processes that promote NSCLC metastasis promises to further improve clinical care of the patients. Kras LA1/+ ;P53 R172HΔG/+ (KP) mice provide a useful and wellvalidated model for the study of NSCLC metastasis. These mice combine a mutant p53 allele (p53 R175HΔG) with an activating KrasG12D allele (Kras LA1 ) (3), leading to development of adenocarcinomas resembling human NSCLC, which are often characterized by mutation of KRAS (∼30%) (4) and loss of TP53 (∼60%) (5). Many of the KP tumors metastasize to sites commonly seen in NSCLC patients (3). These features make the KP murine model a useful tool with which to evaluate factors that underlie NSCLC metastasis. Among the pathways activated...

Musashi RNA-Binding Proteins as Cancer Drivers and Novel Therapeutic Targets

“…Two elegant 2015 studies expanded on the role of Musashi proteins in the intestinal epithelium, demonstrating that either MSI1 or MSI2 can act as an oncogenic driver and is induced by the loss of the tumor suppressor Apc, and identifying these proteins as regulators of the central PDK-AKT-mTORC1 signaling axis, which was defined as essential for Musashi-dependent transformation (37,55). In other work in solid tumors, MSI2 (but not MSI1) was identified as upregulated in, and a driver of, metastasis in non-small cell lung cancer (NSCLC), with activity in directly supporting expression of the TGFβ receptor TGFBR1, and TGFβ effector SMAD3, and indirectly downregulating tight junction–associated claudins to increase tumor cell invasion and metastasis (26). MSI2 targets were found to include many genes associated with cell adhesion and motility and Musashi proteins to promote cell migration in multiple cell types (45,56).…”

“…However, while Musashi proteins repressed NUMB consistently in CNS tumors and some hematologic malignancies, HSCs lacking Msi2 have unchanged levels of the Numb protein(13). Katz et al did not identify significant MSI1-dependent changes in NUMB RNA expression by ribosome profiling in neural stem cells upon MSI1 manipulation (54), and no consistent pattern of change in NUMB protein levels was detected upon MSI2 overexpression or depletion in human and murine NSCLC cells (26). …”

Stem Cells, Cancer, and MUSASHI in Blood and Guts

“…Msi2-deficient HSCs in the mouse had reduced total (and phosphorylated) SMAD3 and increased TGFBR1, which may contribute to the attenuated TGF-β output and lower levels of p57 expression 19 . Of note, MSI2 can also activate TGF-β signaling in non-small cell lung cancer and its metastases 31 . Additional Msi2 targets were associated with the MLL gene expression program that is likely to be shared between normal HSCs and LSCs, including transcripts encoding IKZF2, HOXA9 and MYC 32 .…”

Msi2 Maintains Quiescent State of Hair Follicle Stem Cells by Directly Repressing the Hh Signaling Pathway

“…Inactivation of MSI2 in hematopoietic stem cells (HSCs) impairs their competitive repopulation ability upon transplantation (Hope et al , 2010; Ito et al , 2010; Kharas et al , 2010). Recently, MSI2 was shown to play important roles in several types of cancers (Kang et al , 2016; Katz et al , 2014; Kudinov et al , 2016; Wang et al , 2015). Although Msi2 was shown to be expressed in HF bulge, sHG and IRS (Sugiyama-Nakagiri et al , 2006), its role in HF development and regenerative cycling remains unknown.…”