Purification of the human SMN-GEMIN2 complex and assessment of its stimulation of RAD51-mediated DNA recombination reactions

Motoki Takaku, Takashi Tsujita, Naoki Horikoshi, Yoshimasa Takizawa, Yong Qing, Kouji Hirota, Masae Ikura, Tsuyoshi Ikura, Shunichi Takeda, Hitoshi Kurumizaka

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Abstract

A deficiency in the SMN gene product causes the motor neuron degenerative disease spinal muscular atrophy. GEMIN2 was identified as an SMN-interacting protein, and the SMN-GEMIN2 complex constitutes part of the large SMN complex, which promotes the assembly of the spliceosomal small nuclear ribonucleoprotein (snRNP). In addition to its splicing function, we previously found that GEMIN2 alone stimulates RAD51-mediated recombination in vitro, and functions in DNA double-strand-break (DSB) repair through homologous recombination in vivo. However, the function of SMN in homologous recombination has not been reported. In the present study, we successfully purified the SMN-GEMIN2 complex as a fusion protein. The SMN-GEMIN2 fusion protein complemented the growth-defective phenotype of GEMIN2-knockout cells. The purified SMN-GEMIN2 fusion protein enhanced the RAD51-mediated homologous pairing much more efficiently than GEMIN2 alone. SMN-GEMIN2 possessed DNA-binding activity, which was not observed with the GEMIN2 protein, and significantly stimulated the secondary duplex DNA capture by the RAD51-single-stranded DNA complex during homologous pairing. These results provide the first evidence that the SMN-GEMIN2 complex plays a role in homologous recombination, in addition to spliceosomal snRNP assembly.

Original languageEnglish
Pages (from-to)6797-6805
Number of pages9
JournalBiochemistry
Volume50
Issue number32
DOIs
Publication statusPublished - 2011 Aug 16

ASJC Scopus subject areas

  • Biochemistry

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    Takaku, M., Tsujita, T., Horikoshi, N., Takizawa, Y., Qing, Y., Hirota, K., Ikura, M., Ikura, T., Takeda, S., & Kurumizaka, H. (2011). Purification of the human SMN-GEMIN2 complex and assessment of its stimulation of RAD51-mediated DNA recombination reactions. Biochemistry, 50(32), 6797-6805. https://doi.org/10.1021/bi200828g