Abstract
The ability to integrate environmental and developmental signals with physiological responses is critical for plant survival. How this integration is done, particularly through posttranscriptional control of gene expression, is poorly understood. Previously, it was found that the 30 kD subunit of Arabidopsis cleavage and polyadenylation specificity factor (AtCPSF30) is a calmodulin-regulated RNA-binding protein. Here we demonstrated that mutant plants (oxt6) deficient in AtCPSF30 possess a novel range of phenotypes – reduced fertility, reduced lateral root formation, and altered sensitivities to oxidative stress and a number of plant hormones (auxin, cytokinin, gibberellic acid, and ACC). While the wild-type AtCPSF30 (C30G) was able to restore normal growth and responses, a mutant AtCPSF30 protein incapable of interacting with calmodulin (C30GM) could only restore wild-type fertility and responses to oxidative stress and ACC. Thus, the interaction with calmodulin is important for part of AtCPSF30 functions in the plant. Global poly(A) site analysis showed that the C30G and C30GM proteins can restore wild-type poly(A) site choice to the oxt6 mutant. Genes associated with hormone metabolism and auxin responses are also affected by the oxt6 mutation. Moreover, 19 genes that are linked with calmodulin-dependent CPSF30 functions, were identified through genome-wide expression analysis. These data, in conjunction with previous results from the analysis of the oxt6 mutant, indicate that the polyadenylation factor AtCPSF30 is a regulatory hub where different signaling cues are transduced, presumably via differential mRNA 3′ end formation or alternative polyadenylation, into specified phenotypic outcomes. Our results suggest a novel function of a polyadenylation factor in environmental and developmental signal integration.
Highlights
The 39-end processing of eukaryotic mRNA, that entails cleavage and polyadenylation of a precursor RNA, is a critical step of gene expression [1, 2]
Given that rapid calcium fluxes seem to contribute to the responses of plants to treatments that involve the generation of reactive oxygen species, it seemed possible that the role of AtCPSF30 in responses to oxidative stress may involve its interaction with calmodulin
To elucidate the responses of these transgenic lines to oxidative stress, the lengths of roots of seedlings grown on defined media containing 50 nM methyl viologen (MV) were determined; this assay is the same as that used in the previous characterization of the mutant [16]
Summary
The 39-end processing of eukaryotic mRNA, that entails cleavage and polyadenylation of a precursor RNA, is a critical step of gene expression [1, 2]. 39-end processing is directed by polyadenylation signal sequences at 39-end of precursor mRNA, and involves collaborations of a large number of protein factors. Poly(A) polymerase (PAP), symplekin, and the nuclear poly(A) binding protein (PABPN1) play vital roles to 39 end processing of mRNA [2, 4, 5, 6]. All those factors are required for cleavage reaction, but only CPSF, PAP, and PABP are necessary for adding poly(A) tails. CPSF recognizes the polyadenylation signal (typically AAUAAA) located in 10,30 bp upstream of cleavage site [7]; recognition of G/U rich domain in downstream of cleavage site is mediated by CstF [8]
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