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目的了解Pax7的重要生物学功能,筛选并验证与其相互作用的Ubc9基因编码蛋白,并明确Pax7与小泛素相关修饰蛋白(SUMO)的关系,为进一步研究Pax7的作用机制奠定基础。方法酵母双杂交系统筛选出与Pax7相互作用的基因Ubc9;GST-pull down方法验证Pax7与Ubc9在体外相互作用;Western blot方法检测Pax7的SUMO化状态;免疫共沉淀方法验证Pax7与SUMO的关系。结果 Pax7和Ubc9在酵母中存在相互作用;Pax7和Ubc9在体外可特异性结合;在Ubc9存在的条件下,Pax7呈现SUMO化状态,且可与SUMO相互作用;Pax7在鸡胚胎中也呈现SUMO化状态。结论 Pax7可以与Ubc9相互作用,且这种作用与SUMO有关。
Objective To understand the important biological functions of Pax7, screen and validate the interaction of Ubc9 with its protein, clarify the relationship between Pax7 and SUMO, and lay a foundation for further study on the mechanism of Pax7. Methods The yeast two-hybrid system was used to screen out the gene Ubc9 that interacted with Pax7. The interaction between Pax7 and Ubc9 was verified by GST-pull down assay. The SUMO status of Pax7 was detected by Western blot. The co-immunoprecipitation assay was used to verify the relationship between Pax7 and SUMO. Results Pax7 and Ubc9 showed interaction in yeast. Pax7 and Ubc9 could specifically bind in vitro. In the presence of Ubc9, Pax7 showed SUMO status and interacted with SUMO. Pax7 also showed SUMO in chicken embryos status. Conclusion Pax7 interacts with Ubc9 and this effect is related to SUMO.