SENSITIVE TO PROTON RHIZOTOXICITY1, CALMODULIN BINDING TRANSCRIPTION ACTIVATOR2, and Other Transcription Factors Are Involved inALUMINUM-ACTIVATED MALATE TRANSPORTER1Expression  

Abstract

In Arabidopsis (Arabidopsis thaliana) the root apex is protected from aluminum (Al) rhizotoxicity by excretion of malate, an Al chelator, by ALUMINUM-ACTIVATED MALATE TRANSPORTER1 (AtALMT1). AtALMT1 expression is fundamentally regulated by the SENSITIVE TO PROTON RHIZOTOXICITY1 (STOP1) zinc finger protein, but other transcription factors have roles that enable Alinducible expression with a broad dynamic range. In this study, we characterized multiple cis-elements in the AtALMT1 promoter that interact with transcription factors. In planta complementation assays of AtALMT1 driven by 59 truncated promoters of different lengths showed that the promoter region between -540 and 0 (the first ATG) restored the Al-sensitive phenotype of atalm1 and thus contains ciselements essential for AtALMT1 expression for Al tolerance. Computation of overrepresented octamers showed that eight regions in this promoter region contained potential cis-elements involved in Al induction and STOP1 regulation. Mutation in a position around -297 from the first ATG completely inactivated AtALMT1 expression and Al response. In vitro binding assays showed that this region contained the STOP1 binding site, which accounted for the recognition by four zinc finger domains of the protein. Other positions were characterized as cis-elements that regulated expression by repressors and activators and a transcription factor that determines root tip expression of AtALMT1. From the consensus of known cis-elements, we identified CALMODULIN-BINDING TRANSCRIPTION ACTIVATOR2 to be an activator of AtALMT1 expression. Al-inducible expression of AtALMT1 changed transcription starting sites, which increased the abundance of transcripts with a shortened 59 untranslated region. The present analyses identified multiple mechanisms that regulate AtALMT1 expression.

Malate-dependent Fe accumulation is a critical checkpoint in the root developmental response to low phosphate

“…2C). The first zinc finger domain of STOP1 is critical to bind to the promoter of its target genes (27). Mapping by sequencing of the lpi6 mutant revealed a 13-base pair deletion in the first exon of ALMT1 starting at position +757 (Fig.…”

The Responses of Arabidopsis Early Light-Induced Protein2 to Ultraviolet B, High Light, and Cold Stress Are Regulated by a Transcriptional Regulatory Unit Composed of Two Elements

“…A key factor in this success is the accuracy and sensitivity of promoter prediction. Previously, we developed a unique methodology of promoter prediction using microarray data (Yamamoto et al, 2011) and used the method to identify unique promoter elements that regulate the response of a malate transporter gene, ALUMINUM-ACTIVATED MALATE TRANSPORTER1 (Tokizawa et al, 2015), to soil stresses. In this report, we utilized promoter prediction as the basis for design of synthetic promoters to identify cis-regulatory elements that can conduct environmental stress responses of ELIP2.…”

“…Total RNA was extracted from shoots of stress-treated plants of the wild type (ecotype Columbia) and transfer DNA-tagged HY5 disruptants (SALK_096651c; Alonso et al, 2003) and subjected to real-time qRT-PCR analysis as described previously (Tokizawa et al, 2015). Primers used for RT-PCR are ELIP2_F (59-TATTGACTACACGCAACATCAGAA), ELIP2_R (59-GTTTTCTCCCTTT-GATAACTCCAT-39), UBQ1_F (59-TCGTAAGTACAATCAGGATAAGATG-39), and UBQ1_R (59-CACTGAAACAAGAAAAACAACCCT-39).…”

“…Our method is simple but offers considerable advantages in terms of the accuracy and sensitivity of predictions compared with established methods such as Gibbs Sampling and Multiple Expectation Maximization for Motif Elicidation that are based upon extraction of consensus sequences . We have successfully used this method to identify previously unknown transcriptional regulatory elements for the responses of Arabidopsis to aluminum (Tokizawa et al, 2015). We have used this method to identify putative promoter sequences for ELIP2 and then tested the function of these putative regulatory elements by creating synthetic promoters attached to a luciferase reporter.…”

The roles of STOP1-like transcription factors in aluminum and proton tolerance

“…Recent study has indicated that these C 2 H 2 zinc finger domains are only responsible for directly binding to the cis-acting element GTGCCCAA in the promoter of AtALMT1, but not responsible for transcriptional activation of target genes. 23 We have examined the transcriptional activity of VuSTOP1 via a yeast expression system. Interestingly, we found that both N and C regions are responsible for transactivation.…”