Transcription Factor RAP2.2 and Its Interacting Partner SINAT2: Stable Elements in the Carotenogenesis of Arabidopsis Leaves

Abstract

The promoter of phytoene synthase, the first specific enzyme of carotenoid biosynthesis, shows two main regulatory regions: a G-box-containing region located near the TATA box, and a TATA box distal region containing the cis-acting element ATCTA, which mediates strong basal promoter activity. This second element was also present in the promoter of phytoene desaturase, the next step of the carotenoid pathway, suggesting a common regulatory mechanism. In this work, we demonstrate that AtRAP2.2, a member of the APETALA2 (AP2)/ethylene-responsive element-binding protein transcription factor family, binds to the ATCTA element. In Arabidopsis (Arabidopsis thaliana) leaves, AtRAP2.2 transcript and protein levels were tightly controlled as indicated by unchanged transcript and protein levels in T-DNA insertion mutants in the AtRAP2.2 promoter and 5# untranslated region and the lack of change in AtRAP2.2 protein levels in lines strongly overexpressing the AtRAP2.2 transcript. Homozygous loss-of-function mutants could not be obtained for the AtRAP2.2 5# untranslated region T-DNA insertion line indicating a lethal phenotype. In AtRAP2.2 overexpression lines, modest changes in phytoene synthase and phytoene desaturase transcripts were only observed in root-derived calli, which consequently showed a reduction in carotenoid content. The RING finger protein SEVEN IN ABSENTIA OF ARABIDOPSIS2 (SINAT2) was identified as an AtRAP2.2 interaction partner using a two-hybrid approach. The structure of SINAT2 and related proteins of Arabidopsis show homology to the SEVEN IN ABSENTIA protein of Drosophila that is involved in proteasome-mediated regulation in a variety of developmental processes. The action of SINAT2 may explain the recalcitrance of AtRAP2.2 protein levels to change by altering AtRAP2.2 transcription.

Comprehensive Profiling of Ethylene Response Factor Expression Identifies Ripening-Associated ERF Genes and Their Link to Key Regulators of Fruit Ripening in Tomato

“…Another subclass E member, SlERF.E4 (named SlERF6 in Lee et al, 2012), has been reported to play an important role in fruit ripening by integrating ethylene and carotenoid pathways. This is in agreement with a previous report on RAP2.2, a subclass E Arabidopsis ERF, shown to regulate the expression of carotenoid biosynthesis genes via binding to the ATCTA cis-element in the promoter regions of PSY and PDS (Welsch et al, 2007). Moreover, correlation analysis revealed that the expression of SlERF.F1 is positively correlated with a-carotene accumulation, suggesting the involvement of subclass F members in controlling fruit ripening through the regulation of carotenoid accumulation (Lee et al, 2012).…”

Direct regulation of phytoene synthase gene expression and carotenoid biosynthesis by phytochrome-interacting factors

“…The identification of a common cis element (ATCTA) in the promoters of PSY and other Arabidopsis genes involved in photosynthesis and photoprotection (18) suggested a simple mechanism for the coordinated control of these two critical processes based on the existence of common cis and trans factors. However, the only trans factor found to bind to this motif does not appear to be instrumental for the control of PSY expression or carotenoid synthesis (37). In contrast, the results reported here validate the existence of such a mechanism for a safe transition of etiolated (heterotrophic) seedlings to photomorphogenic (photoautotrophic) development based on direct or indirect regulation of the expression of key genes of both the carotenoid and the chlorophyll pathways by PIF1 and other PIFs in response to light signals.…”

Carotenoids in nature: insights from plants and beyond

“…Very recently, the DELLA proteins were shown to function in part through the regulation of PIF activity, by a DELLA-PIF complex that coordinates the levels of POR, chlorophyll precursors and carotenoids in cotyledons of dark-grown seedlings (Cheminant et al 2011). PSY is strongly lightinduced in greening seedlings and a transcription factor, RAP2.2, (AP2/EREBP family) which binds to the PSY promoter was able to slightly perturb pigment alterations in Arabidopsis root calli (Welsch et al 2007).…”