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Cell stemness is maintained upon concurrent expression of RB and the mitochondrial ribosomal protein S18-2.

ABSTRACT: Stemness encompasses the capability of a cell for self-renewal and differentiation. The stem cell maintains a balance between proliferation, quiescence, and regeneration via interactions with the microenvironment. Previously, we showed that ectopic expression of the mitochondrial ribosomal protein S18-2 (MRPS18-2) led to immortalization of primary fibroblasts, accompanied by induction of an embryonic stem cell (ESC) phenotype. Moreover, we demonstrated interaction between S18-2 and the retinoblastoma-associated protein (RB) and hypothesized that the simultaneous expression of RB and S18-2 is essential for maintaining cell stemness. Here, we experimentally investigated the role of S18-2 in cell stemness and differentiation. Concurrent expression of RB and S18-2 resulted in immortalization of Rb1 ^-/- primary mouse embryonic fibroblasts and in aggressive tumor growth in severe combined immunodeficiency mice. These cells, which express both RB and S18-2 at high levels, exhibited the potential to differentiate into various lineages in vitro, including osteogenic, chondrogenic, and adipogenic lineages. Mechanistically, S18-2 formed a multimeric protein complex with prohibitin and the ring finger protein 2 (RNF2). This molecular complex increased the monoubiquitination of histone H2A^Lys119, a characteristic trait of ESCs, by enhanced E3-ligase activity of RNF2. Furthermore, we found enrichment of KLF4 at the S18-2 promoter region and that the S18-2 expression is positively correlated with KLF4 levels. Importantly, knockdown of S18-2 in zebrafish larvae led to embryonic lethality. Collectively, our findings suggest an important role for S18-2 in cell stemness and differentiation and potentially also in cancerogenesis.

SUBMITTER: Mushtaq M

PROVIDER: S-EPMC7355020 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Cell stemness is maintained upon concurrent expression of RB and the mitochondrial ribosomal protein S18-2.

Mushtaq Muhammad M Kovalevska Larysa L Darekar Suhas S Abramsson Alexandra A Zetterberg Henrik H Kashuba Vladimir V Klein George G Arsenian-Henriksson Marie M Kashuba Elena E

Proceedings of the National Academy of Sciences of the United States of America 20200622 27

Stemness encompasses the capability of a cell for self-renewal and differentiation. The stem cell maintains a balance between proliferation, quiescence, and regeneration via interactions with the microenvironment. Previously, we showed that ectopic expression of the mitochondrial ribosomal protein S18-2 (MRPS18-2) led to immortalization of primary fibroblasts, accompanied by induction of an embryonic stem cell (ESC) phenotype. Moreover, we demonstrated interaction between S18-2 and the retinobla ...[more]

PMID: 32571933

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Project description:Using proteomics analysis, we previously compared MCF7 breast cancer cells grown as 3D tumor spheres, with the same cell line grown as monolayers. Our results indicated that during 3D anchorage-independent growth, the cellular machinery associated with i) mitochondrial biogenesis and ii) ribosomal biogenesis, were both significantly increased. Here, for simplicity, we refer to these two new oncogenic hallmarks as "mito-stemness" and "ribo-stemness" features. We have now applied this same type of strategy to begin to understand how fibroblasts and MCF7 breast cancer cells change their molecular phenotype, when they are co-cultured together. We have previously shown that MCF7-fibroblast co-cultures are a valuable model of resistance to apoptosis induced by hormonal therapies, such as Tamoxifen and Fulvestrant. Here, we directly show that these mixed co-cultures demonstrate the induction of mito-stemness and ribo-stemness features, likely reflecting a mechanism for cancer cells to increase their capacity for accumulating biomass. In accordance with the onset of a stem-like phenotype, KRT19 (keratin 19) was induced by ~6-fold during co-culture. KRT19 is a well-established epithelial CSC marker that is used clinically to identify metastatic breast cancer cells in sentinel lymph node biopsies. The potential molecular therapeutic targets that we identified by label-free proteomics of MCF7-fibroblast co-cultures were then independently validated using a bioinformatics approach. More specifically, we employed publically-available transcriptional profiling data derived from primary tumor samples from breast cancer patients, which were previously subjected to laser-capture micro-dissection, to physically separate breast cancer cells from adjacent tumor stroma. This allowed us to directly validate that the proteins up-regulated in this co-culture model were also transcriptionally elevated in patient-derived breast cancer cells in vivo. This powerful approach for target identification and translational validation, including the use of patient outcome data, can now be applied to other tumor types as well, to validate new therapeutic targets that are more clinically relevant, for patient benefit. Moreover, we discuss the therapeutic implications of these findings for new drug development, drug repurposing and Tamoxifen-resistance, to effectively target mito-stemness and ribo-stemness features in breast cancer patients. We also discuss the broad implications of this "organelle biogenesis" approach to cancer therapy.

Dataset Information

Cell stemness is maintained upon concurrent expression of RB and the mitochondrial ribosomal protein S18-2.

Publications

Cell stemness is maintained upon concurrent expression of RB and the mitochondrial ribosomal protein S18-2.

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