An integrative multi-omics approach uncovers the regulatory role of CDK7 and CDK4 in autophagy activation induced by silica nanoparticles

Autophagy. 2021 Jun;17(6):1426-1447. doi: 10.1080/15548627.2020.1763019. Epub 2020 May 23.


Dysfunction of macroautophagy/autophagy has been postulated as a major cellular toxicological response to nanomaterials. It has been reported that excessive autophagy activation, induced by silica nanoparticles (SiNPs), contributes to autophagy dysfunction, whereas little is known how SiNPs trigger autophagy activation. Here, we treated normal rat kidney (NRK) cells using 3 different sizes of SiNPs (16, 29, and 51 nm) and observed that 16-nm SiNPs, with a final concentration of 60 μg/mL, dramatically induce autophagy activation without reducing cell viability. We further conducted a transcriptomic, proteomic, and phosphoproteomic profiling, and detected 23 autophagy-related (Atg) genes and 35 autophagy regulators regulated on at least one omic layer. To identify key regulators from the multi-omics data, we developed a new algorithm of computational prediction of master autophagy-regulating kinases (cMAK) to detect 21 candidates and revealed the CDK7-CDK4 cascade to be functional. The silence or inhibition of Cdk7 or Cdk4 significantly attenuated autophagic activation but not influenced autophagic flux blockage induced by 16-nm SiNPs. Further computational modeling indicated that the CDK7-CDK4 signaling axis potentially triggers autophagy activation by phosphorylating RB1 (RB transcriptional corepressor 1), activating two critical transcription factors, E2F1 (E2F transcription factor 1) and FOXO3 (forkhead box O3), and enhancing the transcriptional levels of at least 8 Atg genes and autophagy regulators in response to SiNPs. Our studies not only established a powerful method for predicting regulatory kinases from the multi-omics data but also revealed a potential mechanism of SiNP-triggered autophagy activation through modulating the CDK7-CDK4 cascade.Abbreviations: 3-MA: 3-methyladenine; Atg: autophagy-related; BECN1: beclin 1; CCK-8: cell counting kit-8; CDK4: cyclin dependent kinase 4; CDK7: cyclin dependent kinase 7; cMAK: computational prediction of master autophagy-regulating kinases; CQ: chloroquine; DMEM: Dulbecco's modified Eagle's medium; DMSO: dimethyl sulfoxide; E-ratio: enrichment ratio; E2F1: E2F transcription factor 1; EBSS: Earle's balanced salt solution; ER: endoplasmic reticulum; FOXO3: forkhead box O3; FPKM: fragments per kilobase of exon per million fragments mapped; GO: gene ontology; H2O2: hydrogen peroxide; iGPS: in vivo GPS; KEGG: Kyoto Encyclopedia of Genes and Genomes; LC-MS/MS: liquid chromatography-tandem mass spectrometry; LDH: lactate dehydrogenase; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; NRK: normal rat kidney; p-site: phosphorylation site; PBS: phosphate-buffered saline; PDI: polydispersity index; PTM: post-translational modification; QKS: quantitative kinase state; RB1: RB transcriptional corepressor 1; RBHs: reciprocal best hits; RNA-Seq: RNA sequencing; ROS: reactive oxygen species; rSiNPs: SiNPs fluorescently labeled with rhodamine B; SEM: scanning electronic microscopy; SiNPs: silica nanoparticles; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; ssKSR: site-specific kinase-substrate relation; TEM: transmission electron microscopy; tfLC3: mRFP-GFP tandem fluorescent-tagged LC3.

Keywords: Autophagy; CDK4; CDK7; phosphoproteomics; protein kinase; silica nanoparticles.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autophagy / physiology*
  • Cyclin-Dependent Kinase 4 / metabolism*
  • Cyclin-Dependent Kinase-Activating Kinase
  • Cyclin-Dependent Kinases / metabolism*
  • Humans
  • Lysosomes / metabolism
  • Nanoparticles* / metabolism
  • Proteomics / methods
  • Signal Transduction / physiology
  • Silicon Dioxide / metabolism
  • TOR Serine-Threonine Kinases / metabolism
  • Tandem Mass Spectrometry / methods


  • Silicon Dioxide
  • TOR Serine-Threonine Kinases
  • CDK4 protein, human
  • Cyclin-Dependent Kinase 4
  • Cyclin-Dependent Kinases
  • Cyclin-Dependent Kinase-Activating Kinase
  • CDK7 protein, human

Grants and funding

This work was supported by the Special Project on Precision Medicine under the National Key R&D Program [2017YFC0906600 and 2018YFC091050]; Natural Science Foundation of China [31930021, 31970633, 81701567, 21771148, 31671360 and 31801095]; Fundamental Research Funds for the Central Universities [20720180033, 20720200005, 2017KFXKJC001 and 2019kfyRCPY043]; China Postdoctoral Science Foundation [2018M642816, 2018M632870, and 2019T120648].