A regulator of nutritional copper signaling in Chlamydomonas is an SBP domain protein that recognizes the GTAC core of copper response element

Abstract

The CRR1 (Copper Response Regulator) locus, required for both activating and repressing target genes of a copper-and hypoxiasensing pathway in Chlamydomonas, encodes a 1,232-residue candidate transcription factor with a plant-specific DNA-binding domain named SBP, ankyrin repeats, and a C-terminal Cys-rich region, with similarity to a Drosophila metallothionein. The recombinant SBP domain of Crr1 shows zinc-dependent binding to functionally defined copper-response elements associated with the CYC6 and CPX1 promoters that contain a critical GTAC core sequence. Competition experiments indicate equivalent selectivity for copper-response elements from either promoter and 10-fold greater selectivity for the wild-type sequence vs. a sequence carrying a single mutation in the GTAC core. The SBP domain of Chlamydomonas Crr1 binds also to a related GTAC-containing sequence in the Arabidopsis AP1 promoter that is the binding site of a defining member of the SBP family of DNA-binding proteins. Chlamydomonas Crr1 is most similar to a subset of the Arabidopsis SBP domain proteins, which include SPL1, SPL7, and SPL12. The abundance of the CRR1 mRNA is only marginally copper-responsive, and although two mRNAs that differ with respect to splicing of the first intron are detected, there is no indication that the splicing event is regulated by metal nutrition or hypoxia. It is likely that the dramatic copper-responsive action of Crr1 occurs at the level of the polypeptide.acclimation ͉ copper homeostasis ͉ hypoxia ͉ metal ͉ transcription factor

Identification of a Consensus DNA-Binding Site for the Arabidopsis thaliana SBP Domain Transcription Factor, AtSPL14, and Binding Kinetics by Surface Plasmon Resonance

“…The highly conserved SBP DNA-binding domain is a Cys-and His-rich region (consensus: CX 4 CX 13 HX 5 HX-15 CQQCX 3 HX 11 C) found only in proteins from photosynthetic organisms (12,13,18,28). The founding members of the SBP domain gene family (SBP1 and SBP2 from A. majus) were identified by their ability to bind to the upstream regulatory region of the SQUAMOSA floral meristem identity gene (29).…”

“…The Kinetic Binding Parameters of SBP Domains Differ -The SBP domain DNA binding is dependent on the presence of Zn 2+ ions (18,28). The NMR-resolved structures of SBP domains revealed that the SBP domain forms a novel two Zn structure DNA-binding domain (26).…”

“…Moreover, we monitored the kinetic features of the AtSPL14 SBP domain−DNA binding by SPR. We further compare and contrast the target sequences and the equilibrium binding constants we determined for AtSPL14 with those of other SBP domain proteins (18,28,29).…”

“…To date, only a few functions for SBP domain proteins have been reported, perhaps due to genetic redundancy of closely related family members. In all cases, SBP domain proteins have been implicated in various aspects of plant growth and development, including metal sensing in algae and directing development of leaves, embryos, and floral organs in higher plants (9,(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). Some of these SBP do-main proteins may also be posttranscriptionally regulated by noncoding microRNAs to control their spatial and temporal expression (23)(24)(25).…”

Genetically Programmed Changes in Photosynthetic Cofactor Metabolism in Copper-deficient Chlamydomonas

“…In Chlamydomonas, this switch is controlled by copper response elements associated with the CYC6 gene encoding Cyt c 6 and a transcription factor, copper response regulator 1 (CRR1) (4 -6). CRR1 has a SQUAMOSA promoter binding protein (SBP) domain that is conserved in the green lineage (6). Its homologue in Arabidopsis, SPL7, controls nutritional copper homeostasis as well (7,8).…”