Small interference RNA-mediated knockdown of sperm associated antigen 9 having structural homology with c-Jun N-terminal kinase-interacting protein.
Recently, we reported a novel testis-specific sperm associated antigen 9 (SPAG9) protein, a new member of the JNK-interacting protein family, having a functional role in sperm-egg fusion [N. Jagadish, R. Rana, R. Selvi, D. Mishra, M. Garg, S. Yadav, J.C. Herr, K. Okumura, A. Hasegawa, K. Koyama, A. Suri, Biochem. J. 389 (2005) 73-82]. NCBI Blast searches revealed SPAG9 nucleotide sequence similarities with ESTs of various cancerous tissues. In the present study, we compared the efficiency of two independent SPAG9 specific small interfering RNA (siRNA) constructs, BS/U6/spag9 and BS/U6/spag9-I, to ablate the SPAG9 expression in mammalian cells. A positive correlation between the ratio of target gene versus siRNA and the suppression of SPAG9 expression was observed. Further, the cotransfection of BS/U6/spag9 with pcDNA-SPAG9 and pFlag-CMV2-JNK-3 resulted in specific suppression of SPAG9 without affecting JNK-3 expression. The present investigation will eventually extend the application of SPAG9 siRNA in in vivo targeting experiments that aim to define the SPAG9 functional genomics in tumor and reproductive biology.
Rana
Ritu
R
Genes and Proteins Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, J.N.U. Campus, New Delhi-110 067, India.
Jagadish
Nirmala
N
Garg
Manoj
M
Mishra
Deepshikha
D
Dahiya
Neetu
N
Chaurasiya
Dipak
D
Suri
Anil
A
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D020868:Gene Silencing; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D034741:RNA, Small Interfering / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D048868:Adaptor Proteins, Signal Transducing; D008565:Membrane Proteins; D034741:RNA, Small Interfering; C111047:SPAG9 protein, human; D048031:JNK Mitogen-Activated Protein Kinases