Department of Neuroscience, Spinal Cord and Brain Injury Research Center, and Ambystoma Genetic Stock Center, University of Kentucky, Lexington, KY, 40536, USA.
, 10496 Stevens Rd, Vevay, IN, 47043, USA.
Sci Rep. 2024 Sep 27;14(1):22399. doi: 10.1038/s41598-024-73283-1.
Several dozen Mendelian mutants have been discovered in axolotl (Ambystoma mexicanum) populations, including several that affect pigmentation. Four recessive mutants have been described in the scientific literature and genes for three of these have been identified. Here we describe and genetically dissect copper, a mutant with an albino-like phenotype known only from the pet trade. We performed a cross segregating copper and wildtype color phenotypes and used bulked segregant RNA-Seq to identify a region on chromosome 6 that was enriched for single-nucleotide polymorphisms (SNPs) between the color phenotypes. This region included Tyrosinase-like Protein 1 (Tyrp1), a melanin synthesis protein that when mutated, is associated with lighter than black melanin coloration in animal models and oculocutaneous albinism in humans. Inspection of RNA-Seq reads identified a single nucleotide deletion that is predicted to change the coding frame, introduce a premature stop codon in exon 6 and yield a truncated Tyrp1 protein in copper individuals. Using CRISPR-Cas9 editing, we show that wildtype Tyrp1 crispants exhibit copper pigmentation, thus confirming Tyrp1 as the copper locus. Our results suggest that commercial and hobbyist axolotl populations may harbor useful mutants for biological research.
已经在蝾螈(Ambystoma mexicanum)种群中发现了数十种孟德尔突变体,包括几种影响色素沉着的突变体。科学文献中已经描述了四个隐性突变体,其中三个已经鉴定出了基因。在这里,我们描述并遗传剖析了铜突变体,这是一种仅在宠物贸易中才知道的白化表型的突变体。我们进行了一个分离铜和野生型颜色表型的杂交实验,并使用 bulked segregant RNA-Seq 鉴定了一个在染色体 6 上的区域,该区域在颜色表型之间富集了单核苷酸多态性(SNP)。这个区域包括酪氨酸酶样蛋白 1(Tyrp1),这是一种黑色素合成蛋白,当突变时,与动物模型中比黑色黑色素颜色更浅以及人类的眼皮肤白化病有关。检查 RNA-Seq 读数发现了一个单核苷酸缺失,预计会改变编码框架,在 6 号外显子中引入一个过早的终止密码子,并在铜个体中产生一个截断的 Tyrp1 蛋白。使用 CRISPR-Cas9 编辑,我们表明野生型 Tyrp1 敲除个体表现出铜色素沉着,因此证实 Tyrp1 是铜基因座。我们的结果表明,商业和业余蝾螈种群可能拥有对生物研究有用的突变体。