Chemical modifications on histones and DNA/RNA constitute a fundamental mechanism for epigenetic regulation. These modifications often function as docking marks to recruit or stabilize cognate “reader” proteins. So far, a platform for quantitative and high-throughput profiling of the epigenetic interactome is urgently needed but still lacking. Here, we report a 3D-carbene chip-based surface plasmon resonance imaging (SPRi) technology for this purpose. The 3D-carbene chip is suitable for immobilizing versatile biomolecules (e.g., peptides, antibody, DNA/RNA) and features low nonspecific binding, random yet function-retaining immobilization, and robustness for reuses. We systematically profiled binding kinetics of 1,000 histone “reader–mark” pairs on a single 3D-carbene chip and validated two recognition events by calorimetric and structural studies. Notably, a discovery on H3K4me3 recognition by the DNA mismatch repair protein MSH6 in Capsella rubella suggests a mechanism of H3K4me3-mediated DNA damage repair in plant. Significance In the era of functional proteomics, a myriad of new interactions, notably those modification-dependent ones, are widely suggested by advanced proteomic approaches and bioinformatic analysis. Therefore, there exists an urgent need to develop a technology for high-throughput mapping and quantitative characterization of biomolecular binding events. This study achieved the immobilization and kinetic detection of various biomacromolecules (including modified peptides and modified nucleic acids) in high throughput through the 3D-carbene chip-based surface plasmon resonance imaging (SPRi) technology. Modified histone peptides and nucleic acids, which are key epigenetic marks, could be efficiently probed by this platform. We envision that the 3D-carbene SPRi technology described here will have wide appeal in profiling and discovering biological recognitions in and beyond epigenetics.

Objective: Endoplasmic reticulum (ER) stress and mitochondrial function affected intramuscular fat accumulation. However, there is no clear evident on the effect of the regulation of ER stress and mitochondrial function by Angiotensin-converting enzyme 2 (ACE2) on the prevention of intramuscular fat metabolism. We investigated the effects of ACE2 on ER stress and mitochondrial function in skeletal muscle lipid metabolism. Methods: The triglyceride (TG) content in skeletal muscle of ACE2 knockout mice and Ad-ACE2-treated db/db mice were detected by assay kits. Meanwhile, the expression of lipogenic genes (ACCα, SREBP-1c, LXRα, CPT-1α, PGC-1α and PPARα), ER stress and mitochondrial function related genes (GRP78, eIF2α, ATF4, BCL-2, and SDH6) were analyzed by RT-PCR. Lipid metabolism, ER stress and mitochondrial function related genes were analyzed by RT-PCR in ACE2-overexpression C2C12 cell. Moreover, the IKKβ/NFκB/IRS-1 pathway was determined using lysate sample from skeletal muscle of ACE2 knockout mice. Results: ACE2 deficiency in vivo is associated with increased lipid accumulation in skeletal muscle. The ACE2 knockout mice displayed an elevated level of ER stress and mitochondrial dysfunctions in skeletal muscle. In contrast, activation of ACE2 can ameliorate ER stress and mitochondrial function, which slightly accompanied by reduced TG content and down-regulated the expression of skeletal muscle lipogenic proteins in the db/db mice. Additionally, ACE2 improved skeletal muscle lipid metabolism and ER stress genes in the C2C12 cells. Mechanistically, endogenous ACE2 improved lipid metabolism through the IKKβ/NFκB/IRS-1 pathway in skeletal muscle. Conclusions: ACE2 was first reported to play a notable role on intramuscular fat regulation by improving endoplasmic reticulum and mitochondrial function. This study may provide a strategy for treating insulin resistance in skeletal muscle.