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Journal of Peking University(Health Sciences) ›› 2019, Vol. 51 ›› Issue (2): 197-205. doi: 10.19723/j.issn.1671-167X.2019.02.001

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Chronic phosphoproteomic in temporal lobe epilepsy mouse models induced by kainic acid

Zhi-ming SUN1,Qian CHEN1,Ming-hua LI1,Wei-ning MA2,Xu-yang ZHAO1,3,∆(),Zhuo HUANG1,∆()   

  1. 1. Institute of Systems Biomedicine, State Key Laboratory of Natural and Biomimetic Drugs, Department of Molecular and Cellular Pharmacology, Peking University School of Pharmaceutical Science, Beijing 100191, China
    2. Department of Neurosurgery, Sheng Jing Hospital affiliated to China Medical University, Shenyang 110000, China
    3. Beijing Key Laboratory of Tumor Systems Biology, Peking University School of Basic Medical Science, Beijing 100191, China
  • Received:2017-04-27 Online:2019-04-18 Published:2019-04-26
  • Contact: Xu-yang ZHAO,Zhuo HUANG E-mail:zhao-xu-yang@163.com;huangz@hsc.pku.edu.cn
  • Supported by:
    the National Basic Research Program of China(973 Program);the National Basic Research Program of China(2015CB559200);National Science Foundation of China(81371432);National Science Foundation of China(31400695)

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Abstract:

Objective: To investigate functions of proteins and signaling pathways involved in epileptogenesis during the chronic stage of temporal lobe epilepsy in mouse models.Methods: Kainic acid-induced temporal lobe epilepsy models were conducted, when reaching stage 4 using racine scale, the mice of experimental group were supposed to be successfully established. Pentobarbital sodium was injected to stop epileptic seizure in case of death. Twenty-eight days after the kainic acid injection, when the experimental group generally turned into chronic spontaneous seizures, mice hippocampal tissues were extracted from the control and the experimental groups respectively for phosphoproteomic. Enriched phosphorylated proteins were detected using mass spectrometry, only the proteins whose density was greater than 10 6 were analyzed by matching the Gene Ontology (GO) database, Kyoto Encyclopedia of Genes and Genomes (KEGG) database and STRING database to detect proteins involved in epileptogenesis in protein functions, signaling pathways and protein-protein interaction respectively. After that, literatures were reviewed about the key proteins.Results: (1) Total of 12 697 phosphorylation sites of enriched proteins were detected by mass spectrometry, and there were 159 sites whose phosphorylation levels were significantly different from the control (P<0.001). (2) GO database showed that 35.7% of the 159 sites were about “catalytic activity”, 39.5% were about “binding” and 20.8% were about “cell communication”, and the 159 proteins also participated in many biological processes, such as “primary metabolic process” “response to stimulus” “developmental process” “localization” and “phosphate-containing compound metabolic process”. (3) KEGG database showed that the 159 protein sites mainly involved in 10 signaling pathways: glutamatergic synapse, Ras signaling pathway, African trypanosomiasis, Cocaine addiction, Circadian entrainment, Amyotrophic lateral sclerosis (ALS), Long-term potentiation, Endocytosis, Gap junction, Nicotine addiction. (4) STRING database showed that the protein-protein interaction network formed by the 159 proteins was focused on Grin1/Dlg3, Arhgef 2/Arhgap33/Tiam1 and Sptnb1/3/4/Add3/Ank2 protein group respectively. (5) Phosphorylation levels of Grin1, Arhgef 2, Arhgap33, Tiam1, Sptbn1/2/4 and Ank2 in experimental group were significantly higher than in the control (P<0.001).Conclusion: Phosphoproteomic illustrated integral distribution of phosphorylated proteins at the chronic stage of temporal lobe epilepsy in the mouse model. Literatures showed that most key proteins were closely related to epileptogenesis, suggesting that some proteins or signaling pathways may play a role in epileptogenesis, such as dopamine and Kir3.1.

Key words: Temporal lobe epilepsy, Phosphoproteomic, Epileptogenesis

CLC Number: 

  • R393

Table 1

Summary of phosphoproteomic data"

Items Phospho site Phospho protein Up-regulated phospho site Up-regulated phospho protein Down-regulated phospho site Down-regulated
phospho protein
Total 12 697 3 435 4 597 2 084 3 531 1 791
P<0.05 159 147 118 112 41 40

Figure 1

Summary of functions of phosphorylated proteins by matching GO database A, biological adhesion; B, biological regulation; C, cellular process; D, cellular component organization or biogenesis; E, developmental process; F, immune system process; G, localization; H, locomotion; I, metabolic process; J, reproduction; K, multicellular organismal process; L, response to stimulus; M, binding; N, catalytic activity; O, receptor activity; P, signal transducer activity; Q, structural molecule activity; R, translation regulator activity; S, transporter activity."

Table 2

Summary of signaling pathways involved in epileptogensis"

Signaling pathways Number of genes Percentage/% P value Benjamini
Glutamatergic synapse 7 4.7 <0.001 0.03
Ras signaling pathway 8 5.4 <0.001 0.11
African trypanosomiasis 4 2.7 <0.001 0.08
Cocaine addiction 4 2.7 <0.001 0.17
Circadian entrainment 5 3.4 <0.001 0.15
Amyotrophic lateral sclerosis (ALS) 4 2.7 <0.001 0.13
Long-term potentiation 4 2.7 <0.001 0.21
Endocytosis 7 4.7 <0.001 0.26
Gap junction 4 2.7 <0.001 0.31
Nicotine addiction 3 2 <0.001 0.35

Figure 2

Summary of protein-protein interactions by searching STRING database"

Figure 3

Changes of phosphorylation level of proteins involved in epileptogenesis *P<0.05, #P<0.01."

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