Journal of Jilin University(Medicine Edition) ›› 2021, Vol. 47 ›› Issue (1): 66-72.doi: 10.13481/j.1671-587x.20210109

• Research in basic medicine • Previous Articles     Next Articles

Screening and identification of target proteins interacting with GINS2 in human promyelocytic leukemia cDNA library

Xi ZHANG1,Yuqing PAN2,Xin YU1,Yating ZHENG1,Chengbin GUO1,Na XU1,Liangxiao WANG1,Liang HE1(),Li YANG1()   

  1. 1.Department of Clinical Laboratory,Caner Hospital of Yunnan Province,Third Affiliated Hospital,Kunming Medical University,Kunming 650118,China
    2.Department of Clinical Laboratory,First Affiliated Hospital,Kunming Medical University,Yunnan Institute of Experimental Diagnosis,Yunnan Key Laboratory of Laboratory Medicine,Kunming 650032,China
  • Received:2020-06-07 Online:2021-01-28 Published:2021-01-27
  • Contact: Liang HE,Li YANG E-mail:2464606392@qq.com;mrzhh@foxmail.com

Abstract:

Objective: To screen the target proteins interacting with human GINS2(go-ichi-ni-san) by yeast two hybrid system, and to elucidate its pathogenesis in acute promyelocytic leukemia (APL) and related mechanism.

Methods

The constructed bait plasmid pGBKT7-GINS2 and human promyelocyte cDNA library plasmids were simultaneously transformed into the yeast AH109 cells by heat shock method to select the library proteins interacting with GINS2 coding proteins. Then, multiple reporter positive clones detection and DNA sequencing were used for bioinformatics analysis of the genes encoding the interacting proteins.

Results

A total of 15 initial positive clones were successfully screened from the yeast two hybrid library, among which 10 clones could activate ADE2 and HIS3 reporter genes,and 13 clones could activate LacZ reporter gene. After DNA sequencing and BLAST analysis of 10 positive clones which could grow in SD/-Trp/-Leu/-His/-Ade defective culture medium were performed, except for the sequencing failure of No. 8 and No. 9, the remaining 8 positive clones belonged to the eight different protein-coding genes, namely RNA binding motif protein 39(RBM39), ubiquitin like modifier activating enzyme 1(UBA1),metallothionein 2A (MT2A),mitochondrial calcium uptake 1(MICU1),N-ethylmaleimide sensitive fusion protein cofactor 1(NSFL1C), fatty acyl-CoA reductase 1(FAR1), dual specificity phosphatase 1(DUSP1) and cell surface antigen 14(CD14).

Conclusion

A total of eight functional proteins that interact with GINS2 are successfully screened, and it is speculated that the target proteins may play an important role in cell damage, transcriptional regulation, proliferation and metabolism.

Key words: leukemia, human promyelocytic cDNA library, yeast two hybrid, GINS2

CLC Number: 

  • R733.7