Molecular Characterization of the PEND Protein, a Novel bZIP Protein Present in the Envelope Membrane That Is the Site of Nucleoid Replication in Developing Plastids

Abstract

Plastid nucleoids are known to bind to the envelope membrane in developing chloroplasts. Here, plastid DNA is extensively replicated. We previously detected a DNA binding protein in the inner envelope membranes of developing plastids in pea and named it PEND (for plastid envelope DNA binding) protein. In this study, we report on the structure and molecular characterization of a cDNA for the PEND protein. As a result of screening cDNA libraries in gt11 with one of the target sequences of the PEND protein as a probe, we obtained a clone (PD2) for a novel DNA binding protein consisting of 633 amino acid residues. Analysis of the N-terminal sequence of the purified PEND protein indicated that the transit peptide is just 16 residues long. The PEND protein was detected specifically in the plastid envelope membrane of young unopened leaf buds by immunoblot analysis. The PEND protein consists of a basic region plus zipper region, an unprecedented sextuple repeat region, and a putative membrane-spanning region. The basic region with a zipper region seems to have diverged from that of other plant transcription factors. In addition, the PEND protein could be a distant homolog of the trans -Golgi network integral membrane proteins. The PEND protein is therefore a novel type of DNA binding protein that binds to the membrane as an intrinsic membrane protein.

Continuous expression in tobacco leaves of a Brassica napus PEND homologue blocks differentiation of plastids and development of palisade cells

“…A search of the databases revealed homology to GSBF1, a cDNA sequence from B. napus corresponding to a presumed transcription factor (Waldmüller et al. , 1996), PD2, from Pisum sativum encoding PEND (Sato et al. , 1998), and cDNA sequences from Arabidopsis thaliana (accession no. )…”

“…The association of plastid DNA with the inner envelope membrane has been studied in some detail by Sato et al. (1993, 1998) who purified an integral protein from the envelope membranes of pea chloroplasts (PEND; plastid envelope DNA‐binding protein).…”

“…(1993, 1998) who purified an integral protein from the envelope membranes of pea chloroplasts (PEND; plastid envelope DNA‐binding protein). It was suggested that the PEND protein is involved in the transcriptional regulation of chloroplast genes during the early phase of chloroplast development, in the translation of chloroplast mRNA or in replication (Sato and Ohta, 2001; Sato et al. , 1998).…”

The Etched1 gene of Zea mays (L.) encodes a zinc ribbon protein that belongs to the transcriptionally active chromosome (TAC) of plastids and is similar to the transcription factor TFIIS

“…According to Sato's model of discontinuous evolution of plastid nucleoids (Sato, 2001), the plastid genetic machinery has lost many prokaryotic proteins and has acquired nuclear‐encoded eukaryotic‐type proteins, e.g. the PEND protein, which is belonging to the bZIP proteins (Sato, 2001; Sato et al ., 1998). Another example for a protein of eukaryotic origin with a function in plastid nucleoids is the coiled‐coil DNA‐binding protein MFP1, which seems to function as a nucleoid anchor at the interface to the thylakoid membranes (Jeong et al ., 2003).…”

Evolutionary aspects of plastid proteins involved in transcription: The transcription of a tiny genome is mediated by a complicated machinery

“…PEND is composed of a cbZIP domain and a C-terminal hydrophobic domain. The cbZIP domain is involved in dimerization of PEND and sequence-specific DNA binding, whereas the C-terminal hydrophobic domain is required for targeting the PEND protein to the chloroplast envelope membrane 28 . In contrast to PEND, MFP1 has been shown to be localized to the thylakoid membranes and its C-terminal domain has DNA binding activity 29 .…”