Splicing of scorpion toxin gene BmKK2 in HEK 293T cells.
Using GFP as a reporter gene, splicing of scorpion toxin gene BmKK2 was investigated in cultured HEK 293T cells. The results of RT-PCR and western blotting showed that BmKK2's intron could be recognized and spliced in cultured HEK 293T cells. At the same time, a cryptic splicing site of BmKK2 gene was found at the 91st nucleotide site of the second exon, which is a typical form of alternative splicing. For the first time, alternative splicing would partially explain the diversity of scorpion toxins at the gene level. Moreover, replacing BmKK2's intron with BmP03's intron (an artificial BmKK2-BmP03 mosaic gene) did not affect the intron's recognition and splicing, but increased the expression of the toxin-GFP fusion protein by fluorescence imaging, which indicated that both introns may regulate the expression of toxin-GFP fusion protein. The artificial BmKK2-BmP03 mosaic gene was also spliced into two kinds of mRNA molecules, which showed that sequence of intron was not absolutely conserved. The results suggested that introns of scorpion toxin genes BmKK2 and BmP03 increase the diversity of scorpion toxins and regulate the expression of their genes.
Zhijian
Cao
C
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, 430072, People's Republic of China.
Chao
Dai
D
Shijin
Yin
Y
Yingliang
Wu
W
Jiqun
Sheng
S
Yonggang
Sha
S
Wenxin
Li
L
D002460:Cell Line; D006801:Humans; D007438:Introns / Q000235:genetics; D022821:RNA Splice Sites / Q000235:genetics*; D012326:RNA Splicing / Q000235:genetics*; D012604:Scorpion Venoms / Q000235:genetics*
C106379:BmP03 toxin; C445369:KK toxin, Buthus martensi; D022821:RNA Splice Sites; D012604:Scorpion Venoms