The α-Helical D1 Domain of the Tobacco bZIP Transcription Factor BZI-1 Interacts with the Ankyrin-repeat Protein ANK1 and Is Important for BZI-1 Function, Both in Auxin Signaling and Pathogen Response

Abstract

The tobacco (Nicotiana tabacum) bZIP transcription factor BZI-1 is involved in auxin-mediated growth responses and in establishing pathogen defenses. Transgenic plants expressing a dominant-negative BZI-1-⌬N derivative, which lacks the N-terminal activation domain, showed altered vegetative growth. In particular auxin-induced rooting and formation of tobacco mosaic virus-induced hypersensitive response lesions are affected. BZI-1-related proteins described in various plant species share the conserved domains D1, D2, BD, and D4. To define those BZI-1 domains involved in transcription factor function, BZI-1 deletion derivatives were expressed in transgenic plants. The domains D1 or BD are crucial for BZI-1-⌬N function in planta. The basic BD domain is mediating DNA binding of BZI-1. Yeast twohybrid and in vitro binding studies reveal the ankyrinrepeat protein ANK1, which specifically interacts with a part of the BZI-1 protein (amino acids 73-222) encoding the D1 domain. ANK1 does not bind DNA or act as a co-activator of BZI-1-mediated transcription. Moreover, green fluorescence protein localization studies propose that ANK1 is acting mainly inside the cytosol. Transcription analysis reveals that ANK1 is ubiquitously expressed, but after pathogen attack transcription is transiently down-regulated. Along these lines, ANK1 homologous proteins in Arabidopsis thaliana have been reported to function in pathogen defense. We therefore propose that the D1 domain serves as an interaction surface for ANK1, which appears to regulate BZI-1 function in auxin signaling as well as pathogen response.The tobacco bZIP protein BZI-1 displays all the characteristic features of a transcription factor. It binds DNA, in particular ACGT containing cis-elements (ACEs), 1 it is localized inside the nucleus, and its N-terminal domain acts as a trans-activation domain in plant cells (1). Like CPRF2, a highly homologous bZIP protein from parsley, BZI-1 has been isolated by virtue of its in vitro binding to chalcone synthase promoter cis-elements (1-4). However, using various transgenic approaches that modulate the amount or the activation potential of BZI-1, we were not able to show any influence on transcription of phenylpropanoid pathway genes, such as chalcone synthase or phenylalanine ammonia lyase in vivo (1).Functional analysis has been performed in transgenic plants expressing a dominant-negative BZI-1 derivative lacking the N-terminal activation domain (BZI-1-⌬N). These plants display reduced internodes, small curly leaves, enhanced lateral shoot formation, and flowers that are reduced in size. In particular auxin responses appeared to be reduced with respect to auxininduced rooting and regulation of a GH3 target gene. Moreover, BZI-1 transcription is up-regulated in response to pathogen attack and pathogen-induced phosphorylation of BZI-1-related proteins has been described (1, 3, 4).BZI-1-related transcription factors have been isolated from various plant species, e.g. CPRF2, (2), OHP1/2 (5), BLZ1 (6), or bZIP63 (7). Apart from the N...

The Arabidopsis NPR1 Disease Resistance Protein Is a Novel Cofactor That Confers Redox Regulation of DNA Binding Activity to the Basic Domain/Leucine Zipper Transcription Factor TGA1

“…Enhancements of DNA binding activity without changes in complex mobility also have been reported for the human viral protein TAX, which stimulates the DNA binding of bZIP proteins (Wagner and Green, 1993), and the Arabidopsis OBP1 protein, which stimulates the DNA binding of TGA4 and TGA5 to the as-1 element (Zhang et al, 1995). Furthermore, it was reported recently that the tobacco ankyrin repeat protein ANK1 inhibits the binding of the bZIP factor BZI-1 to its cognate promoter element in electrophoretic mobility shift assays without altering the complex mobility (Kuhlmann et al, 2003).…”

Altered expression of an ankyrin-repeat protein results in leaf abnormalities, necrotic lesions, and the elaboration of a systemic signal

“…GBP2 (? ), which lacks the ankyrin-repeat domain, is 98.2% identical in sequence to the tobacco protein AF352797 independently reported as HBP1, TIP1, and ANK1 (Fridborg et al, 2003;Kuhlmann et al, 2003). Therefore, we believe that GBP2 identified in the yeast two-hybrid screen is an incomplete cDNA clone representing a full-length ortholog of tobacco HBP1/TIP1/ANK1.…”

“…Tobacco HBP1/ANK1/TIP1 interacts with the bZIP transcription factor BZI-1 blocking the binding of this transcription factor to DNA. Studies with dominant negative constructs suggest that BZI-1 is a negative regulator of HR and a positive regulator of auxin responses of roots (Kuhlmann et al, 2003). The Arabidopsis PANK AKR2 interacts with both ascorbate peroxidase 3 (APX3), which is believed to be an H 2 O 2 scavenger, and the 14-3-3 protein GF 14k, which also interacts with APX3.…”

Functional roles of the pepper pathogen-induced bZIP transcription factor, CAbZIP1, in enhanced resistance to pathogen infection and environmental stresses

“…A variety of bZIP transcription factors play essential roles in biological processes, such as plant development and morphogenesis (Heinekamp et al 2004). The tobacco bZIP factor, BZI-1, exhibited alterations of vegetative and generative growth, as well as an altered responsiveness to auxin treatment and pathogen infection (Heinekamp et al 2002;Kuhlmann et al 2003). The ultimate understanding of bZIP transcription factors will help reveal regulatory systems of plant defense and development.…”