Exon skipping of exonuclease 1 in MRL/MpJ mice is caused by a nucleotide substitution of the branchpoint sequence in intron eight.
In MRL/MpJ mice, there is a genetic mutation of exonuclease 1 (Exo1), in which the exon 9 is sometimes deleted. In the present study, to check the generation of the spliced exons, exon 8-intron 8-exon 9 (pCX/Ex/EIE/B and pCX/Ex/EIE/M) plasmids were temporally transfected in vitro into BALB 3T3 cells, and RT-PCR using appropriate primer pair was carried out 1 day after transfection. In these constructions, pCX/Ex/EIE/B was derived from genomic sequence of C57BL/6 mice, and pCX/Ex/EIE/M was from MRL/MpJ. A spliced band was detected in pCX/Ex/EIE/B, but was present little or very weakly in pCX/Ex/EIE/M. Next, the same spliced band was demonstrated in the pCX/Ex/EIE/M(T) plasmid, in which the branchpoint sequence (BPS) of pCX/Ex/EIE/M including the exon 9 was changed into that of pCX/Ex/EIE/B. The splicing did not occur in the dell1/B mutant, in which 1960 nucleotides of the intron 8 were deleted, whereas it was detected in the del2/B plasmid deleted 1036 nucleotides in its middle region. These results suggest that the nucleotide T to A mutation of the BPS in the intron 8 is at least a sufficient for generation of splice variants (tr-1 and tr-2 Exo1).
Namiki
Yuka
Y
Laboratory of Experimental Animal Science, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan.
Kon
Yasuhiro
Y
Sasaki
Nobuya
N
Agui
Takashi
T
Endoh
Daiji
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005090:Exodeoxyribonucleases / Q000235:genetics*; D005091:Exons / Q000235:genetics*; D007438:Introns / Q000235:genetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D019463:Mice, Inbred MRL lpr; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D017354:Point Mutation / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014162:Transfection
D005090:Exodeoxyribonucleases; C028450:exodeoxyribonuclease I