ID #470 Histone crotonylation is a novel subgroup difference in atypical teratoid rhabdoid tumors
Aspen Hirsch, Stefania Tocci, Tessa House, Angela Gushue, Payton Zarceno, Shawna Larsen, Whitaker Cohn, Jessica TsaiAbstract
Background
Atypical teratoid rhabdoid tumors (ATRTs) are aggressive tumors of the central nervous system with a marked predilection for young children. While all ATRTs are driven by aberrant chromatin remodeling resulting from SWI/SNF inactivation, the three molecular subgroups- ATRT-TYR, ATRT-SHH, and ATRT-MYC- are defined by transcriptomic and DNA methylation profiling, suggesting a unique oncogenic mechanism. Crotonylation is a novel post-translational modification first discovered on histones that has emerged as a major regulator of gene expression implicated in many cancer types.
Methods
We utilized in vitro patient-derived ATRT neurosphere models, crotonylation immunoblotting, functional genomics, and high-resolution mass spectrometry to determine whether crotonylation differs between ATRT subgroups.
Results
ATRT-SHH cells harbor increased crotonylation at lysine (K) residues K14, K18, K23, and K27 in histone H3 compared to ATRT-MYC cells. Analysis of RNA expression across pediatric brain tumors demonstrates that five genes associated with known histone crotonyltransferases are significantly upregulated ATRT-SHH compared to ATRT-MYC. Together, these findings suggest that crotonylation is enhanced in ATRT-SHH and regulates gene expression to drive ATRT-SHH signaling pathways. We then individually deleted the five crotonyltransferase genes in ATRT-SHH cells to determine if they are necessary for crotonylation in ATRT-SHH and if crotonylation is required for ATRT-SHH cell viability. To establish if sodium crotonate (NaCr), a crotonylation precursor, is sufficient to enhance histone crotonylation in ATRT-MYC, we cultured ATRT-MYC cells in medium containing NaCr and monitored for increased histone crotonylation. Transcriptomic profiling, currently underway, will determine if the increase in histone crotonylation is sufficient to induce the ATRT-SHH signaling program. Lastly, to determine if crotonylation is globally increased in ATRT-SHH cells, all crotonylated proteins will be identified via mass spectrometry with a pan-KCr antibody.
Conclusion
Histone crotonylation is a subtype difference in ATRT and represents a driver of ATRT-SHH pathogenesis which may be amenable to subtype specific therapeutic interventions.