A calpain-6/YAP axis in sarcoma stem cells that drives the outgrowth of tumors and metastases

Sarcomas include cancer stem cells, but how these cells contribute to local and metastatic relapse is largely unknown. We previously showed the pro-tumor functions of calpain-6 in sarcoma stem cells. Here, we use an osteosarcoma cell model, osteosarcoma tissues and transcriptomic data from human tumors to study gene patterns associated with calpain-6 expression or suppression. Calpain-6 modulates the expression of Hippo pathway genes and stabilizes the hippo effector YAP. It also modulates the vesicular trafficking of β-catenin degradation complexes. Calpain-6 expression is associated with genes of the G2M phase of the cell cycle, supports G2M-related YAP activities and up-regulated genes controlling mitosis in sarcoma stem cells and tissues. In mouse models of bone sarcoma, most tumor cells expressed calpain-6 during early steps of tumor out-growth. YAP inhibition prevented the neoformation of primary tumors and metastases but had no effect on already developed tumors. It could even ac..., Total RNAs was extracted from osteosarcoma cells by using the ISOLATE II RNA Mini Kit (Bioline). 
RNA purity/quality (RIN â‰¥ 7) was assessed with the 2100 Agilent Bioanalyzer by using the Agilent small RNA kit. RNAseq involved using the Integragen platform. Libraries were prepared with the Ultra II Directional RNA Library Prep Kit for Illumina protocol according to the supplier recommendations. Briefly, the key stages of this protocol are successive for the purification of PolyA-containing mRNA molecules using poly-T oligo attached magnetic beads from 1µg total RNA (with the Magnetic mRNA Isolation Kit from NEB), fragmentation by using divalent cations under elevated temperature to obtain approximately 300 bp pieces, double-strand cDNA synthesis and finally Illumina adapters ligation and cDNA library amplification by PCR for sequencing. Sequencing was then carried out on Paired End 100b reads of Illumina NovaSeq. Image analysis and base calling involved using Illumina Real..., , , # Data for: A calpain-6/YAP axis in sarcoma stem cells that drives the outgrowth of tumors and metastases

Dataset DOI: 10.5061/dryad.z34tmpghd

## Description of the data and file structure

The 143B human osteosarcoma cell line that was modified to express GFP under control of the CAPN6 regulatory sequence to sort calpain-6 expressing cells (GFP+) from the other tumor cells (GFP-). These cells were compared to a homogeneous population with calpain-6 expression stably inhibited by a specific shRNA (Calp6shRNA cells). A homogeneous population of 143B cells that overexpressed calpain-6 by using a lentiviral vector (Calp6 + cells) was also included in the analysis. Cells were confluent and cultured in 3% O2 in DMEM + 10% FCS

Total RNAs was extracted from 3 independent cultures by using the ISOLATE II RNA Mini Kit (Bioline). Reverse transcription was performed with random primers with the High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems). RNA purity/quality (RIN = 7) was a...