Reversion of the exon - trap Arabidopsis rpn 12 a - 1 mutation by an intragenic suppressor that weakens the chimeric 5 ’ splice site

In the 26S proteasome mutant , an Background: Arabidopsis rpn12a-1 exon-trap T-DNA is inserted 531 base pairs downstream of the STOP RPN12a codon. We have previously shown that this insertion activates a STOP codon-associated latent 5' splice site that competes with the polyadenylation signal during processing of the pre-mRNA. As a result of this dual input from splicing and polyadenylation in the mutant, two transcripts rpn12a-1 RPN12a are produced and they encode the wild-type RPN12a and a chimeric RPN12a-NPTII protein. Both proteins form complexes with other proteasome subunits leading to the formation of wild-type and mutant proteasome versions. The net result of this heterogeneity of proteasome particles is a reduction of total cellular proteasome activity. One of the consequences of reduced proteasomal activity is decreased sensitivity to the major plant hormone cytokinin. We performed ethyl methanesulfonate mutagenesis of and Methods: rpn12a-1 isolated revertants with wild-type cytokinin sensitivity. We describe the isolation and analyses of suppressor of ( Results: rpn12a-1 ). The mutation is intragenic and located at the fifth position of the sor1 sor1 chimeric intron. This mutation weakens the activated 5' splice site associated with the STOP codon and tilts the processing of the mRNA back RPN12a towards polyadenylation. These results validate our earlier interpretation of the unusual Conclusions: nature of the mutation. Furthermore, the data show that optimal 26S rpn12a-1 proteasome activity requires RPN12a accumulation beyond a critical threshold. Finally, this finding reinforces our previous conclusion that proteasome function is critical for the cytokinin-dependent regulation of plant growth. Referee Status: