Expression of Ppp3cb and Ppm1g in the hippocampus of NHE1 gene knockout rats based on proteomics

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Expression of Ppp3cb and Ppm1g in the hippocampus of NHE1 gene knockout rats based on proteomics

Basic Medicine | 更新时间：2026-01-09

- Expression of Ppp3cb and Ppm1g in the hippocampus of NHE1 gene knockout rats based on proteomics
- Journal of Army Medical University Vol. 46, Issue 11, Pages: 1244-1253(2024)
- 作者机构：
  
  1.550000 贵阳，贵州医科大学临床医学院
  2.550025 贵阳，贵州医科大学基础医学院：分子医学工程研究中心
  3.生物教研室
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(8186050199);the General Program of National Natural Science Foundation of China(8236050283)
- DOI：10.16016/j.2097-0927.202309029
  CLC： R338.26;R394-33;R394.2
- Received：06 September 2023，
  
  Revised：2023-10-12，
  
  Published：15 June 2024
- 稿件说明：
移动端阅览
Pengfei MA, Pan YANG, Qian ZHENG, et al. Expression of Ppp3cb and Ppm1g in the hippocampus of NHE1 gene knockout rats based on proteomics[J]. Journal of Army Medical University, 2024, 46(11): 1244-1253.
DOI：

Pengfei MA, Pan YANG, Qian ZHENG, et al. Expression of Ppp3cb and Ppm1g in the hippocampus of NHE1 gene knockout rats based on proteomics[J]. Journal of Army Medical University, 2024, 46(11): 1244-1253. DOI： 10.16016/j.2097-0927.202309029.

摘要

目的

2

通过NHE1基因敲除模型鼠的海马组织差异蛋白质组学分析，发现并明确Ppp3cb和Ppm1g的表达特征。

方法

2

① 选取6只2周龄NHE1基因敲除模型鼠作为模型组，同周龄野生型小鼠6只作为对照组，采用琼脂糖凝胶电泳检测其基因型；应用旷场实验和强迫游泳实验对模型组和对照组小鼠进行行为学评估，并按照Racine评分标准对模型鼠进行癫痫发作分级；②通过串联质谱分析技术对模型组和对照组的海马组织进行差异蛋白筛选，基因本体论(Gene Ontology Analysis

GO)分析差异蛋白并进行注释和富集，蛋白网络数据库(search tool for the retrieval of interesting genes

STRING)分析差异蛋白之间的蛋白相互作用(protein-protein interaction

PPI)；③应用qPCR和Western blot检测Ppp3cb和Ppm1g的转录和翻译水平，应用免疫组织化学技术分别观察其在组织中的表达量。

结果

2

① 模型组小鼠NHE1基因未见表达，旷场实验中模型鼠的运动总距离较对照组减少(

P

=0.007 3)，跨越的格子数比对照组显著减少(

P

＜0.000 1)。强迫游泳实验结果显示，模型鼠不动的时间明显延长(

P

＜0.000 1)；②以表达倍数(FC)≥1.2倍且

P

<

0.05为筛选标准，检测到海马组织中845个差异表达的蛋白质点，其中有9个蛋白表达上调，7个蛋白表达下调。其中Ppp3cb下调，Ppm1g上调。GO功能注释结果表明，NHE1敲除后，分子功能(MF)富集在蛋白丝氨酸/苏氨酸磷酸酶活性的差异最显著，细胞成分(CC)富集在质膜部分的差异蛋白数量最多，生物过程(BP)富集在负向调节生物过程、免疫系统过程的差异蛋白数量最多。STRING分析显示差异蛋白Ppp3cb和Slc9a1直接作用，Ppm1g通过Ppp3cb和Slc9a1间接作用，Ppp3cb和Ppm1g之间相互作用。③Ppp3cb的转录和翻译水平减少，在组织中的表达量下降，而Ppm1g转录和翻译水平增加，在组织中的表达量上升(

P

＜0.05)。

结论

2

本研究确定了NHE1基因敲除小鼠海马组织中差异蛋白Ppp3cb表达下调，而Ppm1g表达上调，为进一步研究Ppp3cb和Ppm1g参与癫痫发病机制提供了依据。

Abstract

Objective

2

To investigate and validate the expression profiles of Ppp3cb and Ppm1g through differential proteomic analysis of hippocampal tissue in NHE1 gene knockout mice with proteomic analysis.

Method

2

① Six 2-week-old NHE1 knockout mice were selected as the model group

and 6 wild-type mice of the same age served as the control group

and their genotypes were detected by agar-gel electrophoresis. Open field test and forced swimming test were used to evaluate the behaviors of mice in the model group and control group

and epileptic seizure was graded according to Racine scoring. ② Tandem mass spectrometry was employed to screen the differential proteins in the hippocampus tissues from the model group and the control group. Then the obtained differential proteins were annotated and enriched in the Gene Ontology (GO) database. Search tool for the retrieval of interesting genes (STRING) database was used to analyze protein-protein interaction (PPI) among different proteins. ③ The transcriptional and translational levels of Ppp3cb and Ppm1g were detected by qPCR and Western blotting

respectively

and their expression levels in the tissues were observed with immunohistochemistry.

Results

2

① NHE1 was not expressed in the model group. The mice of the model group had shorter total movement distance (

P

=0.007 3) and less crossing cells (

P

<

0.000 1) in open field test

and longer period of immobility in forced swimming test (

P

<

0.000 1) when compared with those from the control group. ② When fold change ≥1.2 times and

P

<

0.05 were set as the significant threshold for differential expression

845 differentially expressed protein sites were detected in the hippocampus

among which 9 proteins (including Ppm1g) were up-regulated and 7 ones (including Ppp3cb) were down-regulated. Gene Ontology (GO) functional analysis showed that after NHE1 knockout

the most significant differences were observed in the concentration of molecular function (MF) related to protein serine/threonine phosphatase activity

concentration of cellular component (CC) related to the plasma membrane

and con

centration of biological process (BP) related to negative regulation of biological processes and immune system processes. STRING analysis indicated that the differential proteins Ppp3cb and Slc9a1 directly acted

Ppm1g indirectly acted through Ppp3cb and Slc9a1

and Ppp3cb and Ppm1g interacted. ③The transcriptional and translational levels of Ppp3cb were decreased

and its expression level was reduced in the tissues

while those of Ppm1g were increased

and its expression was elevated in the tissues (

P

<

0.05).

Conclusion

2

In the hippocampus of NHE1 gene knockout mice

the expression of differential protein Ppp3cb is down-regulated and that of Ppm1g is up-regulated

which provide a basis for further study on their involvement in the pathogenesis of epilepsy.

关键词

Keywords

references

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