Mutations in cis-regulatory elements play important roles for phenotypic changes during evolution. Eye degeneration in the blind mole rat (BMR; Nannospalax galili) and other subterranean mammals is significantly associated with widespread divergence of eye regulatory elements, but the effect of these regulatory mutations on eye development and function has not been explored. Here, we investigate the effect of mutations observed in the BMR sequence of a conserved non-coding element upstream of Tdrd7, a pleiotropic gene required for lens development and spermatogenesis. We first show that this conserved element is a transcriptional repressor in lens cells and that the BMR sequence partially lost repressor activity. Next, we recapitulated evolutionary changes in this element by precisely replacing the endogenous regulatory element in a mouse line by the orthologous BMR sequence with CRISPR-Cas9. Strikingly, this repressor replacement caused a more than two-fold up-regulation of Tdrd7 in the developing lens; however, increased mRNA level does not result in a corresponding increase in TDRD7 protein nor an obvious lens phenotype, possibly explained by buffering at the posttranscriptional level. Our results are consistent with eye degeneration in subterranean mammals having a polygenic basis where many small-effect mutations in different eye-regulatory elements collectively contribute to phenotypic differences.

中文翻译：

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在单个顺式调控元件中概括进化差异足以引起晶状体基因Tdrd7的表达变化。

顺式调控元件的突变对于进化过程中的表型变化起重要作用。盲mole鼠（BMR；Nannospalax galili）和其他地下哺乳动物的眼睛退化与眼睛调节元件的广泛散布显着相关，但是尚未探讨这些调节突变对眼睛发育和功能的影响。在这里，我们调查了在Tdrd7上游保守的非编码元件的BMR序列中观察到的突变的影响，是晶状体发育和精子形成所需的多效基因。我们首先显示该保守元件是晶状体细胞中的转录阻遏物，而BMR序列部分失去了阻遏物活性。接下来，我们通过用CRISPR-Cas9用直系同源BMR序列精确替换小鼠系中的内源性调控元件，概括了该元件的进化变化。令人惊讶的是，这种阻遏物的替代引起Tdrd7的两倍以上上调在显影镜头中；然而，增加的mRNA水平不会导致TDRD7蛋白相应的增加，也不会导致明显的晶状体表型，这可能是由于转录后水平的缓冲所致。我们的结果与具有多基因基础的地下哺乳动物的眼部退化一致，在该基础上，不同眼部调节元件中的许多小效应突变共同促成表型差异。