Human CUL1 forms an evolutionarily conserved ubiquitin ligase complex (SCF) with SKP1 and an F-box protein
Human CUL1 forms an evolutionarily conserved ubiquitin ligase complex (SCF) with SKP1 and an F-box protein
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DOI：
10.1073/pnas.95.13.7451
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发表时间：
1998-06-23
影响因子：
11.1
通讯作者：
Deshaies, RJ
中科院分区：
文献类型：
--
作者：
Lyapina, SA;Correll, CC;Deshaies, RJ
芽殖酵母的SCF泛素连接酶复合物通过催化S期细胞周期蛋白依赖性激酶抑制剂SIC 1的泛素化来触发DNA复制。SCF由三种蛋白质组成-ySKP 1，CDC 53（Cullin）和F-box蛋白CDC 4-它们从酵母到人类都是保守的。作为鉴定推定的人SCF复合物的组分和底物的努力的一部分，我们在与hCUL 1（与CDC 53最接近的人同源物）的双杂交筛选中分离了hSKP 1。在这里，我们表明hCUL 1在体内与hSKP 1缔合，并且在体外直接与hSKP 1和人F-bos蛋白SKP 2相互作用，形成SCF样颗粒。hCUL 1补充酵母cdc 53（ts）突变体的生长缺陷，与人细胞提取物中的泛素化促进活性相关，并且可以与酵母SCF组分组装成功能性嵌合泛素连接酶复合物。综上所述，这些数据表明，hCUL 1功能的SCF泛素连接酶复合物在人类细胞中的一部分。类似于本文所述的生化测定的进一步应用现在可用于鉴定基于hCUL 1的SCF复合物的调节剂/组分，以确定hCUL 2-hCUL 5蛋白是否也是人细胞中泛素连接酶复合物的组分，并筛选调节hSKP 1和hCUL 1蛋白活性的化合物。
The SCF ubiquitin ligase complex of budding yeast triggers DNA replication by catalyzing ubiquitination of the S phase cyclin-dependent kinase inhibitor SIC1. SCF is composed of three proteins-ySKP1, CDC53 (Cullin), and the F-box protein CDC4-that are conserved from yeast to humans. As part of an effort to identify components and substrates of a putative human SCF complex, we isolated hSKP1 in a two-hybrid screen with hCUL1, the closest human homologue of CDC53, Here, we show that hCUL1 associates with hSKP1 in vivo and directly interacts with both hSKP1 and the human F-bos protein SKP2 in vitro, forming an SCF-like particle, Moreover, hCUL1 complements the growth defect of yeast cdc53(ts) mutants, associates with ubiquitination-promoting activity in human cell extracts, and can assemble into functional, chimeric ubiquitin ligase complexes with yeast SCF components. Taken together, these data suggest that hCUL1 functions as part of an SCF ubiquitin ligase complex in human cells. Further application of biochemical assays similar to those described here can now be used to identify regulators/components of hCUL1-based SCF complexes, to determine whether the hCUL2-hCUL5 proteins also are components of ubiquitin ligase complexes in human cells, and to screen for chemical compounds that modulate the activities of the hSKP1 and hCUL1 proteins.