Molecular basis underlying histone H3 lysine–arginine methylation pattern readout by Spin/Ssty repeats of Spindlin1

Abstract

Histone modification patterns and their combinatorial readout have emerged as a fundamental mechanism for epigenetic regulation. Here we characterized Spindlin1 as a histone effector that senses a cis-tail histone H3 methylation pattern involving trimethyllysine 4 (H3K4me3) and asymmetric dimethylarginine 8 (H3R8me2a) marks. Spindlin1 consists of triple tudor-like Spin/Ssty repeats. Cocrystal structure determination established concurrent recognition of H3K4me3 and H3R8me2a by Spin/Ssty repeats 2 and 1, respectively. Both H3K4me3 and H3R8me2a are recognized using an ''insertion cavity'' recognition mode, contributing to a methylation statespecific layer of regulation. In vivo functional studies suggest that Spindlin1 activates Wnt/b-catenin signaling downstream from protein arginine methyltransferase 2 (PRMT2) and the MLL complex, which together are capable of generating a specific H3 ''K4me3-R8me2a'' pattern. Mutagenesis of Spindlin1 reader pockets impairs activation of Wnt target genes. Taken together, our work connects a histone ''lysine-arginine'' methylation pattern readout by Spindlin1-to-Wnt signaling at the transcriptional level.

Kinetic and high-throughput profiling of epigenetic interactions by 3D-carbene chip-based surface plasmon resonance imaging technology

“…This result agrees reasonably with that determined by ITC (K D = 9.4 μM) (20). In addition, this affinity agrees well with that determined using the oriented immobilization strategy [K D = 8.2 ± 0.7 μM, C-terminal biotin labeled H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4me3 peptide] (SI Appendix, Fig. S3).…”

“…First, we checked the positive controls in this peptide array. The H3K4me3 readers binds to H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4me3 instead of H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4un peptides as expected (Fig. 4B).…”

“…Empirically, the real positive result should have a RC value less than 10, which means that unregenerable signal contributes less than 10% to the overall signal. We chose H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4me3 and H3 K9crK27cr peptides as examples to analyze their respective signal response to CHD1. The result showed that the regenerable signal of H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4me3 peptide contributes to 99.8% of the overall signal while this value of H3 K9crK27cr peptide is only 53.3% (SI Appendix, Fig.…”

“…The 3D-carbene SPRi platform makes high-throughput kinetic analysis of histone "mark-reader" pairs possible. We analyzed kinetic curves of four peptides [H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4un, H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4ac, H3 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) K4me1, and H3 (1)(2)(3)(4)(5)(6)(7)…”

“…To complicate the matter further, the recognition of marks by reader proteins is often either synergized or antagonized by other marks in close proximity (8). For instance, Spindlin1 recognizes histone H3 trimethylation of Lys-4 (H3K4me3) and asymmetrically dimethylation of Arg-8 (H3R8me2a) in concert, while phosphorylation of Ser-10 on H3 (H3S10ph) completely abrogates the binding of H3 Lys-9 trimethylation (H3K9me3) by HP1 protein (9,10). BRDT protein binds neither histone H4 Lys-5 acetylation (H4K5ac) nor Lys-8 acetylation (H4K8ac) but engages H4 peptide bearing both marks with decent affinity (K D = 22 μM) (11).…”