Differential CRE Expression in Lhrh-cre and GnRH-cre Alleles and the Impact on Fertility in Otx2-Flox Mice
Differential CRE Expression in Lhrh-cre and GnRH-cre Alleles and the Impact on Fertility in Otx2-Flox Mice
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DOI：
10.1159/000497791
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发表时间：
2019-01-01
影响因子：
4.1
通讯作者：
Mellon, Pamela L.
中科院分区：
文献类型：
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作者：
Hoffmann, Hanne M.;Larder, Rachel;Mellon, Pamela L.
通过Cre重组的条件性基因缺失，利用细胞特异性基因缺失的研究有增加的趋势。尽管有许多优点，但这一策略也有局限性，如异位Cre表达和生殖系重组。两种常用的促性腺激素释放激素(GnRH)驱动的Cre表达的小鼠都以GnRH神经元为靶标。然而，靶向细胞及其表型结果的直接比较尚未公布。为了比较重组发生在哪里，我们用rosa26-lacZ报告鼠杂交了GnRH-cre和LHRH-cre线。LHRH-cre允许在大约700个细胞中重组rosa26-lacZ基因，这与GnRH神经元群体相当。令人惊讶的是，在成年GnRH-cre：rosa26-lacZ中，表达LacZ的细胞比LHRH-cre：rosa26-lacZ多20倍。GnRH-cre和LHRH-cre在靶向上的最大差异是在隔核、视交叉上核和下丘脑隔区。这种靶向细胞的差异从胚胎第12天就出现了。之前一项使用GnRH-cre删除转录因子OTX2的研究发现，GnRH神经元较少，导致男性和女性亚生育。为了总结这项研究，我们对OTX2：LHRH-cre小鼠进行了生育能力测试。我们证实了OTX2：LHRH-cre雄性对GnRH神经元的发育、生育和促性腺激素的正确释放都需要OTX2，但这种不育性比OTX2：GnRH-cre弱，而在雌性OTX2：LHRH-cre中没有。这表明GnRH-cre的异位表达与观察到的生殖表型有关。最后，当Cre等位基因从任何一个亲本传播时，会导致FLOX等位基因的种系重组，产生致命的胚胎敲除后代，产生更小的活后代。
There is an increasing trend in studies utilizing cell-specific deletion of genes through conditional gene deletion by CRE recombination. Despite numerous advantages, this strategy also has limitations such as ectopic CRE-expression and germline recombination. Two commonly used gonadotropin-releasing hormone (Gnrh)-driven CRE-expressing mice both target GnRH neurons. However, a direct comparison of the cells targeted and their phenotypic outcome have not yet been presented. To compare where recombination takes place, we crossed the Gnrh-cre and Lhrh-cre lines with the Rosa26-LacZ reporter mouse. Lhrh-cre allowed recombination of the Rosa26-LacZ gene in approximate to 700 cells, which is comparable to the GnRH neuronal population. Surprisingly, there were > 20 times more LacZ expressing cells in the adult Gnrh-cre:Rosa26-LacZ than the Lhrh-cre:Rosa26-LacZ brain. The greatest differences in targeting of the Gnrh-cre and Lhrh-cre lines were found in the septum, the suprachiasmatic nucleus, and the septohypothalamic area. This difference in cells targeted was present from embryonic day 12. A prior study using the Gnrh-cre to delete the transcription factor Otx2 found fewer GnRH neurons, leading to male and female subfertility. To recapitulate this study, we performed a fertility assay in Otx2:Lhrh-cre mice. We confirmed the requirement for Otx2 in GnRH neuron development, fertility and correct gonadotropin hormone release in Otx2:Lhrh-cre males, but the subfertility was more modest than in Otx2:Gnrh-cre and absent in female Otx2:Lhrh-cre. This suggests that ectopic expression of Gnrh-cre contributes to the reproductive phenotype observed. Finally, the Cre alleles caused germline recombination of the flox allele when transmitted from either parent, generating embryonic lethal knock-out offspring, producing smaller live litters.