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J Mol Biol. 2005 Sep 23;352(3):585-96.

Functional dissection of the plant-specific SBP-domain: overlap of the DNA-binding and nuclear localization domains.

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1
Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829 Cologne, Germany.

Abstract

SBP-domain proteins are plant-specific putative transcription factors. They all contain the highly conserved 76 amino acid residue SBP-domain, shown to bind specifically to related motifs in the Antirrhinum majus SQUA promoter and the orthologous Arabidopsis thaliana AP1 promoter. The structural basis for this sequence-specific binding of DNA are two Zn-finger like structures formed by the coordination of two zinc ions by conserved cysteine and histidine residues. Amino acid exchanges of the cysteine residues involved revealed that each of the Zn(2+)-coordinating structures is essential for DNA binding. By random target-site selection studies, it is shown that the palindromic GTAC core motif is essential for efficient DNA binding with additional nucleotides preferred by different SBP-domain proteins. Despite their different functions and origin from plants at different evolutionary distances, the mode of DNA binding is conserved from the single-cell algae Chlamydomonas reinhardtii to the moss Physcomitrella patens and higher plants. At the C-terminal end of the SBP-domain, a putative bipartite nuclear localization signal is located, which overlaps with the DNA-binding domain, in particular with the second Zn(2+)-binding structure. By immunolocalization of SPL3 and transient expression of SBP-green fluorescent protein fusion proteins in plant cells, it is shown that this nuclear localization signal is functional. Exchange of a highly conserved serine next to the nuclear localization signal by aspartate, which may mimic phosphorylation, resulted in a decreased nuclear import (SPL8), while DNA binding in vitro was abolished completely. In contrast, exchange by alanine increased nuclear import and left DNA binding intact. This suggests that the function of SBP-domain proteins is also regulated by post-translational modification on the levels of nuclear import and DNA binding.

PMID:
16095614
DOI:
10.1016/j.jmb.2005.07.013
[Indexed for MEDLINE]
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