Objectives

Glucose-stimulated insulin secretion is a critical function in the regulation of glucose homeostasis, and its deregulation is associated with the development of type 2 diabetes. Here, we performed a genetic screen using islets isolated from the BXD panel of advanced recombinant inbred (RI) lines of mice to search for novel regulators of insulin production and secretion.

Methods

Pancreatic islets were isolated from 36 RI BXD lines and insulin secretion was measured following exposure to 2.8 or 16.7 mM glucose with or without exendin-4. Islets from the same RI lines were used for RNA extraction and transcript profiling. Quantitative trait loci (QTL) mapping was performed for each secretion condition and combined with transcriptome data to prioritize candidate regulatory genes within the identified QTL regions. Functional studies were performed by mRNA silencing or overexpression in MIN6B1 cells and by studying mice and islets with beta-cell-specific gene inactivation.

Results

Insulin secretion under the 16.7 mM glucose plus exendin-4 condition was mapped significantly to a chromosome 2 QTL. Within this QTL, RNA-Seq data prioritized Crat (carnitine O-acetyl transferase) as a strong candidate regulator of the insulin secretion trait. Silencing Crat expression in MIN6B1 cells reduced insulin content and insulin secretion by ∼30%. Conversely, Crat overexpression enhanced insulin content and secretion by ∼30%. When islets from mice with beta-cell-specific Crat inactivation were exposed to high glucose, they displayed a 30% reduction of insulin content as compared to control islets. We further showed that decreased Crat expression in both MIN6B1 cells and pancreatic islets reduced the oxygen consumption rate in a glucose concentration-dependent manner.

Conclusions

We identified Crat as a regulator of insulin secretion whose action is mediated by an effect on total cellular insulin content; this effect also depends on the genetic background of the RI mouse lines. These data also show that in the presence of the stimulatory conditions used the insulin secretion rate is directly related to the insulin content.

中文翻译：

---

遗传筛选将Crat鉴定为胰腺β细胞胰岛素分泌的调节剂。

目标

葡萄糖刺激的胰岛素分泌是调节葡萄糖稳态的关键功能，其失调与2型糖尿病的发生有关。在这里，我们使用从小鼠的高级重组近交（RI）系的BXD面板中分离的胰岛进行了基因筛选，以寻找胰岛素产生和分泌的新型调节剂。

方法

从36个RI BXD品系中分离出胰岛，并在暴露于2.8或16.7 mM葡萄糖（有或没有exendin-4的情况下）后测量胰岛素分泌。来自相同RI系的胰岛用于RNA提取和转录谱分析。对每种分泌条件进行定量性状基因座（QTL）作图，并与转录组数据结合以对已鉴定的QTL区域内的候选调控基因进行优先排序。通过在MIN6B1细胞中进行mRNA沉默或过表达以及通过研究具有β细胞特异性基因失活的小鼠和胰岛来进行功能性研究。

结果

在16.7 mM葡萄糖加exendin-4条件下的胰岛素分泌显着定位于2号染色体QTL。在此QTL中，RNA-Seq数据将Crat（肉碱O-乙酰基转移酶）列为胰岛素分泌性状的强有力候选调节剂。在MIN6B1细胞中沉默Crat表达可使胰岛素含量和胰岛素分泌减少约30％。相反，Crat的过度表达使胰岛素含量和分泌增加了约30％。当来自具有β细胞特异性Crat失活的小鼠的胰岛暴露于高葡萄糖时，与对照胰岛相比，它们的胰岛素含量降低了30％。我们进一步表明，降低的Crat MIN6B1细胞和胰岛中的蛋白表达以葡萄糖浓度依赖性方式降低了耗氧率。

结论

我们将Crat鉴定为胰岛素分泌的调节剂，其作用是通过对总细胞胰岛素含量的影响来介导的。这种作用还取决于RI小鼠品系的遗传背景。这些数据还表明，在存在刺激条件的情况下，胰岛素分泌速率与胰岛素含量直接相关。