Pirh2 is an E3 ligase belonging to the RING-H2 family and shown to bind, ubiquitinate and downregulate p73 tumor suppressor function without altering p73 protein levels. AIP4, an E3 ligase belonging to the HECT domain family, has been reported to be a negative regulatory protein that promotes p73 ubiquitination and degradation. Herein, we found that Pirh2 is a key regulator of AIP4 that inhibits p73 function. Pirh2 physically interacts with AIP4 and significantly downregulates AIP4 expression. This downregulation is shown to involve the ubiquitination of AIP4 by Pirh2. Importantly, we demonstrated that the ectopic expression of Pirh2 inhibits the AIP4-p73 negative regulatory pathway, which was restored when depleting endogenous Pirh2 utilizing Pirh2-siRNAs. We further observed that Pirh2 decreases AIP4-mediated p73 ubiquitination. At the translational level and specifically regarding p73 cell cycle arrest function, Pirh2 still ensures the arrest of p73-mediated G1 despite AIP4 expression. Our study reveals a novel link between two E3 ligases previously thought to be unrelated in regulating the same effector substrate, p73. These findings open a gateway to explain how E3 ligases differentiate between regulating multiple substrates that may belong to the same family of proteins, as it is the case for the p53 and p73 proteins.

中文翻译：

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Eir连接酶Pirh2通过调节AIP4表达和泛素化来调节AIP4-p73调节途径。

Pirh2是一种E3连接酶，属于RING-H2家族，在不改变p73蛋白水平的情况下，它可以结合，泛素化和下调p73肿瘤抑制功能。据报道，AIP4（一种属于HECT域家族的E3连接酶）是一种负调控蛋白，可促进p73泛素化和降解。在本文中，我们发现Pirh2是抑制p73功能的AIP4的关键调节因子。Pirh2在物理上与AIP4相互作用，并显着下调AIP4的表达。显示该下调涉及Pirh2对AIP4的泛素化。重要的是，我们证明了Pirh2的异位表达抑制了AIP4-p73负调控途径，当利用Pirh2-siRNA消耗内源性Pirh2时，该途径得以恢复。我们进一步观察到，Pirh2降低了AIP4介导的p73泛素化。在翻译水平，特别是关于p73细胞周期阻滞功能，尽管AIP4表达，Pirh2仍可确保p73介导的G1的阻滞。我们的研究揭示了以前认为与调节同一效应物底物p73不相关的两种E3连接酶之间的新颖联系。这些发现为解释E3连接酶如何在调节可能属于同一蛋白质家族的多种底物之间进行区分开辟了途径，p53和p73蛋白质就是这种情况。