Expression and processing analyses of wild type and p.R47H TREM2 variant in Alzheimer's disease brains.
Expression and processing analyses of wild type and p.R47H TREM2 variant in Alzheimer's disease brains.
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DOI：
10.1186/s13024-016-0137-9
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发表时间：
2016-11-25
影响因子：
15.1
通讯作者：
Sevlever D
中科院分区：
文献类型：
--
作者：
Ma L;Allen M;Sakae N;Ertekin-Taner N;Graff-Radford NR;Dickson DW;Younkin SG;Sevlever D
遗传分析表明，在髓样细胞2（TREM2）P.R47H变体中表达的触发受体增加了阿尔茨海默氏病的风险（AD）。
Quantitative real-time PCR (qPCR) was performed using 2 sets of primers one that detects all TREM2 mRNA isoforms and one specific for the alternative spliced isoform (TREM2alt) that encodes for the extracellular domain (soluble TREM2). Because in the brain TREM2 is expressed primary in microglial cells, we also assessed the levels of IBA1 to control for microglial variability across samples. For TREM2 protein quantitation and N-糖基化处理，RIPA脑提取物在EndoH和PNGASEF处理前后通过Western印迹分析。
与对照相比，我们在AD受试者的临时皮层中确定了统计学上的trem2转录水平。 P.R47H的载体中的TREM2对单个TREM2物种的分析发现，成熟和未成熟物种的相对量没有差异，而非载体和P.R47H样品之间的羧基末端碎片与非载体或P.R47H的trem2物种同样容易受到Endoh和P.R47H的影响。
我们的结果表明，AD中的Trem2表达增加了，我们提供的证据表明，P.R47H突变不会直接通过改变表达或间接影响蛋白质的处理来影响蛋白质的处理，以影响蛋白质的研究结果，这些发现表明P.R47H的变化会通过更改受体而不是表达的结合特性来影响TREM2。
Genetic analyses showed that the triggering receptor expressed in myeloid cells 2 (TREM2) p.R47H variant increases the risk for Alzheimer’s disease (AD). The question of whether the p.R47H mutation affects expression or function of the receptor remains unanswered. To address this question we quantified mRNA and analyzed protein profiles of WT and p.R47H TREM2 in human brains. Quantitative real-time PCR (qPCR) was performed using 2 sets of primers one that detects all TREM2 mRNA isoforms and one specific for the alternative spliced isoform (TREM2alt) that encodes for the extracellular domain (soluble TREM2). Because in the brain TREM2 is expressed primarily in microglial cells, we also assessed the levels of IBA1 to control for microglial variability across samples. For TREM2 protein quantitation and N-glycosylation processing, RIPA brain extracts were analyzed by Western blot before and after EndoH and PNGaseF treatments. We identified statistically significant increased levels of TREM2 transcripts in the temporal cortex of AD subjects when compared with controls; TREM2alt was likewise higher in AD cases, but was not significant after adjustment for covariates. Quantitative analysis of TREM2 protein confirmed qPCR results that showed higher levels in AD than in control brains. Among AD subjects, we observed a trend towards higher mRNA and protein TREM2 levels in carriers of the p.R47H risk allele. Analysis of individual TREM2 species found no difference in the relative amounts of mature and immature species, and carboxyl terminal fragments between non carriers and p.R47H samples. Furthermore, TREM2 species from either non carriers or p.R47H brains were equally susceptible to EndoH and PNGaseF treatments. Our results suggest that TREM2 expression is increased in AD. Furthermore, we provide evidence indicating that p.R47H mutation does not affect the levels of TREM2 either directly by altering expression or indirectly by affecting processing of the protein. Our data support previous findings that suggest that p.R47H variant affects TREM2 function by altering binding properties of the receptor rather than expression.
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DOI：
10.1056/nejmoa1211851
发表时间：
2013-01-10
期刊：
The New England journal of medicine
影响因子：
--
作者：
Guerreiro R;Wojtas A;Bras J;Carrasquillo M;Rogaeva E;Majounie E;Cruchaga C;Sassi C;Kauwe JS;Younkin S;Hazrati L;Collinge J;Pocock J;Lashley T;Williams J;Lambert JC;Amouyel P;Goate A;Rademakers R;Morgan K;Powell J;St George-Hyslop P;Singleton A;Hardy J;Alzheimer Genetic Analysis Group
通讯作者：
Alzheimer Genetic Analysis Group
影响因子：
30.8
作者：
Paloneva, J;Kestilä, M;Peltonen, L
通讯作者：
Peltonen, L
影响因子：
3.5
作者：
Jin, Sheng Chih;Benitez, Bruno A.;Cruchaga, Carlos
通讯作者：
Cruchaga, Carlos
影响因子：
12.7
作者：
Piccio L;Deming Y;Del-Águila JL;Ghezzi L;Holtzman DM;Fagan AM;Fenoglio C;Galimberti D;Borroni B;Cruchaga C
通讯作者：
Cruchaga C
影响因子：
4.4
作者：
Thrash, J. Cameron;Torbett, Bruce E.;Carson, Monica J.
通讯作者：
Carson, Monica J.