Genetic variation in the enzymes that catalyze posttranslational modification of proteins is a potentially important source of phenotypic variation during evolution. Ubiquitination is one such modification that affects turnover of virtually all of the proteins in the cell in addition to roles in signaling and epigenetic regulation. UBE2D3 is a promiscuous E2 enzyme, which acts as an ubiquitin donor for E3 ligases that catalyze ubiquitination of developmentally important proteins. We have used protein sequence comparison of UBE2D3 orthologs to identify a position in the C-terminal α-helical region of UBE2D3 that is occupied by a conserved serine in amniotes and by alanine in anamniote vertebrate and invertebrate lineages. Acquisition of the serine (S138) in the common ancestor to modern amniotes created a phosphorylation site for Aurora B. Phosphorylation of S138 disrupts the structure of UBE2D3 and reduces the level of the protein in mouse embryonic stem cells (ESCs). Substitution of S138 with the anamniote alanine (S138A) increases the level of UBE2D3 in ESCs as well as being a gain of function early embryonic lethal mutation in mice. When mutant S138A ESCs were differentiated into extraembryonic primitive endoderm, levels of the PDGFRα and FGFR1 receptor tyrosine kinases were reduced and primitive endoderm differentiation was compromised. Proximity ligation analysis showed increased interaction between UBE2D3 and the E3 ligase CBL and between CBL and the receptor tyrosine kinases. Our results identify a sequence change that altered the ubiquitination landscape at the base of the amniote lineage with potential effects on amniote biology and evolution.

中文翻译：

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羊膜特异性机制的进化，该机制通过E2酶UBE2D3的磷酸化调节泛素信号传导。

催化蛋白质翻译后修饰的酶的遗传变异是进化过程中表型变异的潜在重要来源。泛素化是一种这样的修饰，除了在信号传导和表观遗传调控中起作用外，它实际上影响细胞中所有蛋白质的更新。UBE2D3是混杂的E2酶，充当E3连接酶的泛素供体，E3连接酶催化重要的蛋白质泛素化。我们已经使用UBE2D3直系同源物的蛋白质序列比较来确定UBE2D3的C末端α螺旋区域中的位置，该位置由羊膜中的保守丝氨酸以及羊膜脊椎动物和无脊椎动物谱系中的丙氨酸占据。在现代祖先的共同祖先中获得丝氨酸（S138）产生了Aurora B的磷酸化位点。S138的磷酸化破坏了UBE2D3的结构，并降低了小鼠胚胎干细胞（ESCs）中蛋白质的水平。用羊膜丙氨酸（S138A）替代S138可提高ESC中UBE2D3的水平，并获得小鼠早期胚胎致死突变的功能。当突变体S138A ESCs分化为胚外原始内胚层时，PDGFRα和FGFR1受体酪氨酸激酶的水平降低，原始内胚层分化受到损害。邻近连接分析显示，UBE2D3与E3连接酶CBL之间以及CBL与受体酪氨酸激酶之间的相互作用增加。我们的结果确定了序列变化，该序列变化改变了羊膜谱系基础上的泛素化景观，并对羊膜生物学和进化有潜在影响。