Comparative proteome analysis of wild-type and klotho-knockout mouse kidneys using a combination of MALDI-IMS and LC-MS/MS.
Comparative proteome analysis of wild-type and klotho-knockout mouse kidneys using a combination of MALDI-IMS and LC-MS/MS.
复制标题
使用 MALDI-IMS 和 LC-MS/MS 组合对野生型和 klotho 敲除小鼠肾脏进行比较蛋白质组分析。
DOI：
10.1002/prca.201600095
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发表时间：
2017
期刊：
影响因子：
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通讯作者：
Yoshiko Y.
中科院分区：
文献类型：
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作者：
Fujino Y;Minamizaki T;Hayashi I;Kawakami A;Miyaji T;Sakurai K;Yoshioka H;Kozai K;Okada M;Yoshiko Y.
目的研究小鼠klothogene基因的突变，诱发类似人类加速衰老的综合征。然而，Klotho在肾脏中的作用的证据有限。我们对野生型（WT）和klotho基因敲除（kl−/−）小鼠肾脏进行了比较蛋白质组分析，以确定与Klotho缺陷相关的蛋白质。实验设计使用7周龄雄性WT和kl −/−小鼠冷冻肾脏切片的MALDI成像MS（MALDI-IMS）来确定MS分布的基因型特异性差异。蛋白质独特地分布inkl−/−肾脏被确定通过随后的分析相邻的胰蛋白酶化切片MALDI-IMS结合LC-MS/MS。免疫组织化学和蛋白质印迹法采用定性和定量analysis. ResultsNine ‐ 7和69蛋白质LC-MS/MS鉴定匹配的MALDI-IMS光谱WT和kl −/−小鼠肾脏，分别。在匹配的蛋白质类型中，核酸结合蛋白最丰富，其次是酶。我们鉴定了分泌颗粒蛋白-1（SCG 1），它主要分布在kl −/−小鼠肾脏的肾小球和肾小管中。结论SCG 1可能是Klotho缺陷症的候选蛋白。虽然需要进一步的研究来研究SCG 1在肾脏中的作用，但我们显示了MALDI-IMS与LC-MS/MS相结合的有用性。
PurposeMutation of theklothogene in mice elicits a syndrome resembling accelerated human aging. However, there is limited evidence for the role of Klotho in the kidney. We conducted a comparative proteome analysis of wild‐type (WT) andklotho‐knockout (kl−/−) mouse kidneys to identify proteins involved in Klotho deficiency.Experimental designMALDI imaging MS (MALDI‐IMS) of frozen kidney sections from 7‐wk‐old male WT andkl−/−mice was used to determine genotype‐specific differences in the MS distribution. Proteins uniquely distributed inkl−/−kidneys were identified by subsequent analysis of adjacent trypsinized sections by MALDI‐IMS in combination with LC‐MS/MS. Immunohistochemistry and western blotting were adopted in qualitative and quantitation analysis.ResultsNinety‐seven and 69 proteins identified by LC‐MS/MS were matched to the MALDI‐IMS spectra in WT andkl−/−mouse kidneys, respectively. Among protein types matched, nucleic acid binding proteins were most abundant, followed by enzymes. We identified secretogranin‐1 (SCG1), which was predominately distributed in the glomeruli and renal tubules ofkl−/−mouse kidneys. Immunohistochemistry for SCG1 mirrored images of MALDI‐IMS.ConclusionsSCG1 may be a candidate protein involved in Klotho deficiency. Although further research is needed to investigate the role of SCG1 in the kidney, we show the usefulness of MALDI‐IMS combined with LC‐MS/MS.