Characterization of a Chloroplast Inner Envelope K+ Channel

Mi, Feng; Peters, Jeanne S.; Berkowitz, Gerald A.

doi:10.1104/pp.105.3.955

Cited by 28 publications

(28 citation statements)

References 29 publications

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“…In addition to these transporters, envelope membranes contain a series of ion channels, pumps, permeases, pore proteins, and other substances (Fischer et al, 1994;Mi et al, 1994;Heiber et al, 1995;Pohlmeyer et al, 1997) essential for the functional integration of plastids within cells. Most of these proteins have been characterized in chloroplasts; little is known about nongreen plastids.…”

Section: Other Transporters Regulate Chloroplast Metabolic Activitymentioning

confidence: 99%

The Biochemical Machinery of Plastid Envelope Membranes

et al. 1998

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Plastids are semiautonomous organelles with a wide structural and functional diversity and unique biochemical pathways. As such, they are able to transcribe and translate the information present in their own genome but are strongly dependent on imported proteins that are encoded in the nuclear genome and translated in the cytoplasm. Plastids are present in every plant cell, with very few exceptions (such as the highly specialized male sexual cells), and their structural and functional diversity reflects their role in different cell types. According to their developmental stage, we distinguish them as juvenile (proplastids), differentiating, mature, and senescent. Meristematic cells contain proplastids, which ensure the continuity of plastids from generation to generation and are capable of considerable structural and metabolic plasticity to develop into various types of plastids that remain interconvertible. When leaves are grown in darkness, proplastids differentiate into etioplasts, which can be converted into chloroplasts under illumination.The metabolism of these various types of plastids is linked to the function of the tissue in which they are found. For instance, whereas the chief function of illuminated leaves is the assimilation of CO 2 by chloroplasts, root plastids are mainly involved in the assimilation of inorganic nitrogen. Amyloplasts, which contain large starch grains, behave as storage reservoirs in stems, roots, and tubers. Chromoplasts synthesize large amounts of carotenoids and are present in petals, fruits, and even roots. The interconversions between these different plastids are accompanied by dramatic changes, including the development or regression of internal membrane systems (e.g. thylakoids and prolamellar bodies) and the acquisition of specific enzymatic equipment reflecting specialized metabolism. However, at all stages of these transformations, the two limiting envelope membranes remain apparently unchanged.Located at the interface between plastids and the surrounding cytosol, the envelope is a key structure for the integration of plastid metabolism within the cell. Because plastids are semiautonomous organelles, a tight coordination between plastidial development and cell differentiation is required. Envelope membranes are an essential checkpoint between the expression of plastidial and nuclear genomes, for example, as the site for the specific recognition and transport of the precursor plastid proteins synthesized on cytosolic ribosomes. Plastid membranes contain an astonishing variety of specific lipids, including polar lipids (e.g. galactolipids, phospholipids, and SLs), pigments (e.g. carotenoids and chlorophylls), and prenylquinones (e.g. plastoquinone and tocopherols). This diversity requires complex metabolic pathways that are closely associated with envelope membranes.A unique biochemical machinery (Fig. 1) is present in envelope membranes and reflects the stage of development of the plastid and the specific metabolic requirements of the various tissues. PLASTID ENVELOPE ME...

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Section: Other Transporters Regulate Chloroplast Metabolic Activitymentioning

confidence: 99%

The Biochemical Machinery of Plastid Envelope Membranes

et al. 1998

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“…Several ion channels of the inner as well as the outer envelope membrane have been identified by reconstitution of the channels in giant liposomes or in planar lipid bilayers (5)(6)(7)(8). Direct electrophysiological measurements on isolated chloroplasts have also revealed a number of envelope ion channels (9,10).…”

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confidence: 99%

A 50-Picosiemens Anion Channel of the Chloroplast Envelope Is Involved in Chloroplast Protein Import

Wijngaard

Vredenberg

1997

Journal of Biological Chemistry

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Single channel recordings were used to investigate the changes on the pea chloroplast envelope during protein import. In the inside-out patch configuration a 50-picosiemens (pS) anion channel of the chloroplast envelope membrane was identified. The open time probability of the channel was decreased by the addition of the wild type precursor protein of ferredoxin (wt-prefd) to the pipette-filling solution in the presence of 0.5 mM ATP. In the absence of ATP or in the presence of 50 M ATP, wt-prefd did not affect the open time probability of the channel. A deletion mutant of prefd, ⌬6 -14-prefd, which is inactive in in vitro import, was also unable to affect the open time probability of the 50-pS anion channel. In the presence of 100 M ATP, wt-prefd decreased the open time probability of the channel to a lesser extent, as did the transit peptide alone. It is concluded that the 50-pS anion channel could be part of the protein import machinery of the inner membrane. In addition the precursor protein under import conditions induced burst-like increases of the envelope conductivity. The implication of both responses for the chloroplast protein import process are discussed.A large part of the chloroplast proteins is nuclear encoded. These proteins are synthesized in the cytosol and have to be imported into the chloroplast. Nuclear encoded chloroplast proteins are therefore synthesized with an N-terminal extension called the transit sequence. The transit sequence is both necessary and sufficient to target a protein to the chloroplast (1). Several components of the chloroplast import machinery have been identified (2, 3). A role for chloroplast envelope channels has been proposed in protein import (2-4). However, little is known about the function of channels in the import process and the response of the envelope during protein import. An increase in envelope conductivity during protein import is shown to occur in Peperomia metallica chloroplasts (4). This increase was identified using electrophysiological measurements in the whole chloroplast configuration. It was suggested that the opening of protein translocation channels is the cause of the observed response of the envelope.Ion channels in the chloroplast envelope are also thought to be involved in osmoregulation during photosynthesis. Several ion channels of the inner as well as the outer envelope membrane have been identified by reconstitution of the channels in giant liposomes or in planar lipid bilayers (5-8). Direct electrophysiological measurements on isolated chloroplasts have also revealed a number of envelope ion channels (9, 10).In this report the use of single channel recordings in investigating the role of chloroplast envelope components in protein import is described for the first time. The involvement of a chloroplast envelope anion channel in protein import is identified. A hitherto unidentified electrical response of the chloroplast envelope associated with protein import is also described. The implications of both responses for the chloroplast protei...

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“…A GenBank search indicates that all proteins that contain sequences homologous to the channel pore peptide are K+ channels (7). (iii) Biochemical analysis (19) demonstrates that when detergentsolubilized chloroplast IE protein is fractionated on a sucrose density gradient, the immunoreactive polypeptide and K+-channel activity comigrate.…”

Section: Resultsmentioning

confidence: 99%

“…Purified sarcolemmal vesicles recovered from the 8%/33% sucrose interface were washed and stored (at 5 mg of protein per ml) in 160 mM NaCl/20 mM Tris-HCl, pH 7.4. Chloroplast envelope vesicles were prepared from spinach (Spinacea oleracea L.) as described (19,20) by exposure of intact chloroplasts (isolated from 5-10 kg of spinach leaves using Percoll step gradients) to freeze/thaw cycles in hyperosmotic medium.…”

Section: Methodsmentioning

confidence: 99%

Development of a K(+)-channel probe and its use for identification of an intracellular plant membrane K+ channel.

Mi¹,

Berkowitz²

1995

Proc. Natl. Acad. Sci. U.S.A.

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Polyclonal antibodies were generated against a 9-amino acid, synthetic peptide corresponding to the selectivity filter in the pore region of K+-channel proteins. The sequence of amino acids in the ion-conducting pore region of K+ channels is the only highly conserved region of members of this protein family. The objectives of the present work were (i) to determine whether the anti-channel pore peptide antibody was immunoreactive with known K+-channel proteins and (ii) to demonstrate the usefulness of the antibody by employing it to identify a newly discovered K+-channel protein. Anti-channel pore peptide was immunoreactive with various K+-channel subtypes native to a number of different species. Immunoblot analysis demonstrated affinity of the antibody for the drkl, maxi-K, and KAT1 K+-channel proteins. Studies also suggested that the anti-channel pore peptide antibody did not immunoreact with membrane proteins other than K+ channels. The anti-channel pore peptide antibody was used to establish the identity of a 62-kDa chloroplast inner envelope polypeptide as a putative component of a K+-channel protein. It was concluded that an antibody generated against the conserved pore region/selectivity filter of K+ channels has broad but selective affinity for this class of proteins. This K+-channel probe may be a useful tool for identification of K+-channel proteins in native membranes.K+ channels are a diverse and ubiquitous family of proteins. Over the past few years, homology screening of cDNA libraries and expression cloning (in Xenopus oocytes or yeast null mutants) have led to identification of nucleotide sequences encoding numerous newly discovered K+ channels. However, virtually everything we know about the structure of these proteins is deduced from analysis of the corresponding nucleotide sequences. Few molecular analyses have been undertaken on K+-channel proteins expressed in native membranes. This is due to the lack of useful probes with strong and specific affinity for a broad range of K+-channel proteins. Radiolabeled neurotoxins such as charybdotoxin and dendrotoxin have aided the study of a few K+-channel proteins (1, 2). However, these toxins do not bind to all K+-channel types and have been found to have no affinity for plant K+ channels (3). Here, we describe an alternative approach to identification of newly discovered members of this protein family. This research focuses on the development and testing of a "generic" K+-channel antibody. The strategy we followed was to raise polyclonal antibodies to a peptide corresponding to a highly conserved region of K+ channels.The K+-conducting portion of native K+ channels is thought to be a multimer of four identical or similar polypeptides (4). Recent reviews group known K+-channel polypeptides into four subfamilies (4,5). Deduced sequences of K+ channels belonging to the four major groupings share some structural homology. There is, however, virtually no region of the The publication costs of this article were defrayed in part by page charge paymen...

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Characterization of a Chloroplast Inner Envelope K+ Channel

Cited by 28 publications

References 29 publications

The Biochemical Machinery of Plastid Envelope Membranes

The Biochemical Machinery of Plastid Envelope Membranes

A 50-Picosiemens Anion Channel of the Chloroplast Envelope Is Involved in Chloroplast Protein Import

Development of a K(+)-channel probe and its use for identification of an intracellular plant membrane K+ channel.

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