PSMC5 knockout HeLa cell line

补充内容

PSMC5即26S蛋白酶调节亚基8，也称为26S蛋白酶AAA-ATPase亚基Rpt6，是由PSMC5基因在人类中编码的酶。作为蛋白质编码基因，PSMC5在人体中衍生出重要的生物学功能，参与细胞内蛋白质降解并促进结肠癌细胞的增殖和转移。它作为一种新型的药物靶点或生物标志物，被广泛应用于药物研发领域。PSMC5在19S蛋白酶体复合物中是必需的亚基之一，与多种疾病有关，如Cockayne综合症和Ogden综合症。通过功能性实验和临床数据分析，揭示了PSMC5在CRC中的重要作用和机制。PSMC5的研究为理解细胞内蛋白质降解过程以及疾病发生发展提供了重要线索和参考。

1. 设计 gRNA：选择目标 PSMD5 基因的特定区域，并设计适当的 gRNA (guide RNA) 序列，用于导向 CRISPR/Cas9 系统在目标基因上引发双链断裂。

2. 选择细胞系：选择适当的细胞系作为研究对象，通常选择易于转染且能够长期培养的细胞株。

3. 转染 CRISPR/Cas9 系统：将设计好的 gRNA 和 Cas9 蛋白共转染入目标细胞系中，使其形成 Cas9/gRNA 复合物，针对 PSMD5 基因发挥作用。

4. 筛选阳性细胞克隆：通过细胞筛选技术，筛选出成功敲除 PSMD5 基因的阳性细胞克隆，确保其基因敲除效率和稳定性。

5. 鉴定 PSMD5 基因敲除细胞系：利用 PCR、Western blot 或测序等技术验证目标 PSMD5 基因的敲除效果，确保已成功构建 PSMD5 基因敲除细胞系。

6. 功能鉴定和研究：进一步对 PSMD5 基因敲除细胞系进行功能研究，探究 PSMD5 基因在细胞生物学和疾病发生发展中的功能和机制。

Species

Gene ID

GenBank Accession

Transcript

Human (Homo sapiens)

PSMC5

5705

NM_002805.6

NP_002796.4

PRS8

Proteasome 26S Subunit, ATPase 5

HR55, MSS1, PRS8

17q23.3

protein-coding

A8K3Z3, A8K763, O35051, O43208, P47210, P52915, P52916, P62195

PSMC5 participates in the proteasome pathway

The 26S proteasome is a multicatalytic proteinase complex with a highly ordered structure composed of 2 complexes, a 20S core and a 19S regulator. The 20S core is composed of 4 rings of 28 non-identical subunits; 2 rings are composed of 7 alpha subunits and 2 rings are composed of 7 beta subunits. The 19S regulator is composed of a base, which contains 6 ATPase subunits and 2 non-ATPase subunits, and a lid, which contains up to 10 non-ATPase subunits. Proteasomes are distributed throughout eukaryotic cells at a high concentration and cleave peptides in an ATP/ubiquitin-dependent process in a non-lysosomal pathway. An essential function of a modified proteasome, the immunoproteasome, is the processing of class I MHC peptides. This gene encodes one of the ATPase subunits, a member of the triple-A family of ATPases which have a chaperone-like activity. In addition to participation in proteasome functions, this subunit may participate in transcriptional regulation since it has been shown to interact with the thyroid hormone receptor and retinoid X receptor-alpha. Two transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Nov 2010]