Constitutive activation of NF-κB inducing kinase (NIK) in the mesenchymal lineage using Osterix (Sp7)- or Fibroblast-specific protein 1 (S100a4)-Cre drives spontaneous soft tissue sarcoma.

Jennifer L Davis,Fei Wan,Heather M Zannit,Andre J Van Wijnen,Biancamaria Ricci,Deborah J Veis,Roman Thaler,Linda Cox,Roberta Faccio,Amel Dudakovic,Jung-Eun Kim

doi:10.1371/journal.pone.0254426

Abstract

Aberrant NF-κB signaling fuels tumor growth in multiple human cancer types including both hematologic and solid malignancies. Chronic elevated alternative NF-κB signaling can be modeled in transgenic mice upon activation of a conditional NF-κB-inducing kinase (NIK) allele lacking the regulatory TRAF3 binding domain (NT3). Here, we report that expression of NT3 in the mesenchymal lineage with Osterix (Osx/Sp7)-Cre or Fibroblast-Specific Protein 1 (FSP1)-Cre caused subcutaneous, soft tissue tumors. These tumors displayed significantly shorter latency and a greater multiple incidence rate in Fsp1-Cre;NT3 compared to Osx-Cre;NT3 mice, regardless of sex. Histological assessment revealed poorly differentiated solid tumors with some spindled patterns, as well as robust RelB immunostaining, confirming activation of alternative NF-κB. Even though NT3 expression also occurs in the osteolineage in Osx-Cre;NT3 mice, we observed no bony lesions. The staining profiles and pattern of Cre expression in the two lines pointed to a mesenchymal tumor origin. Immunohistochemistry revealed that these tumors stain strongly for alpha-smooth muscle actin (αSMA), although vimentin staining was uniform only in Osx-Cre;NT3 tumors. Negative CD45 and S100 immunostains precluded hematopoietic and melanocytic origins, respectively, while positive staining for cytokeratin 19 (CK19), typically associated with epithelia, was found in subpopulations of both tumors. Principal component, differential expression, and gene ontology analyses revealed that NT3 tumors are distinct from normal mesenchymal tissues and are enriched for NF-κB related biological processes. We conclude that constitutive activation of the alternative NF-κB pathway in the mesenchymal lineage drives spontaneous sarcoma and provides a novel mouse model for NF-κB related sarcomas.

Highlights

NF-κB signaling is best known for its pivotal roles in innate and adaptive immunity [1]
In the course of characterizing the bone phenotype of the Osx-Cre;NT3 mice [26], we observed spontaneous soft tissue tumors. Based on their subcutaneous location and histological appearance, we considered that a mesenchymal origin was likely, and crossed the NT3 transgenic mice with a Fibroblast-Specific Protein 1 (FSP1)-Cre (S100a4) line [31], which predominantly shows expression in the fibroblast population of multiple organs but has been documented in the myeloid lineage [31, 32, 43,44,45]
We did observe a rare population of Tdt+ cells in the dermis as well as in cultured dermal fibroblasts from Osx-Cre;Tdt mice (S3 Fig). These findings suggest that NT3 transgene recombination occurs in dermal fibroblasts of both strains, with the greater extent in FSP1-Cre;NT3 mice corresponding to a greater number and earlier onset of tumors

Summary

Introduction

NF-κB signaling is best known for its pivotal roles in innate and adaptive immunity [1] This transcription factor family can regulate both physiologic and pathologic processes outside of the immune system such as in bone and skin, and has been implicated in tumorigenesis [1,2,3,4]. The alternative NF-κB pathway is triggered exclusively through NF-κB-inducing kinase (NIK) which phosphorylates IKKα to mobilize RelB/p52-containing dimers into the nucleus [1, 6]. Both RelB and Nfkb, the gene encoding p52, have κB binding sites in their promoters and are transcriptionally upregulated by classical NF-κB stimuli. While activation of either the classical or the alternative NF-κB program is highly stimulus-dependent, significant crosstalk exists between these two pathways [7]

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Results

Conclusion