Abstract
While transcription as regulated by histones and their post-translational modifications has been well described, the function of histone variants in this process remains poorly characterized. Potentially important insight into this process pertain to the frequently occurring mutations of H3.3, leading to G34 substitutions in childhood glioblastoma and giant cell tumor of the bone (GCTB). In this study, we have established primary cell lines from GCTB patients and used them to uncover the influence of H3.3 G34W substitutions on cellular growth behavior, gene expression, and chromatin compaction. Primary cell lines with H3.3 G34W showed increased colony formation, infiltration and proliferation, known hallmarks of tumor development. Isogenic cell lines with H3.3 G34W recapitulated the increased proliferation observed in primary cells. Transcriptomic analysis of primary cells and tumor biopsies revealed slightly more downregulated gene expression, perhaps by increased chromatin compaction. We identified components related to splicing, most prominently hnRNPs, by immunoprecipitation and mass spectrometry that specifically interact with H3.3 G34W in the isogenic cell lines. RNA-sequencing analysis and hybridization-based validations further enforced splicing aberrations. Our data uncover a role for H3.3 in RNA processing and chromatin modulation that is blocked by the G34W substitution, potentially driving the tumorigenic process in GCTB.
Highlights
Central to cancer progression is the deterioration of function and integrity of tumorigenic cells previously in a structured relationship with tissues and organs in the organism[1]
Since stratification of giant cell tumor of the bone (GCTB) samples based on mutational status remains uncharted, we wanted to determine if there is a difference between locations of tumor and mutational incidence
We found that the distal femur and proximal tibia of the lower long bones are the areas where GCTB are the most frequent, especially in the German cohort
Summary
Central to cancer progression is the deterioration of function and integrity of tumorigenic cells previously in a structured relationship with tissues and organs in the organism[1]. Gain-of-function characteristics in cancer driver genes caused by genetic aberrations can readily overthrow this order. Histones, with their key and multifunctional properties, are central components of the cell vulnerable to these forces[2]. The leading example is a mutation of H3.3 that produces K27M substitution (hereafter referred to as H3.3K27M) which sterically bind and block the function of the polycomb repressive complex 26–8. Mutations in the very same gene have been identified in giant cell tumor of the bone (GCTB), not in children but in younger adults[9]. GCTB with H3.3G34W poses a proliferative and infiltrative advantage that uncover important aspects of H3.3 in cancer and opens possibilities for new stratification regiments and treatments
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