Characterization of transcriptionally active DNA-protein complexes from chloroplasts and etioplasts of mustard (Sinapis alba L.)

Authors


Correspondence to G. Link, Institut für Biologie II, Universität Freiburg, Schänzlestraße 1, D-7800 Freiburg, Federal Republic of Germany

Abstract

DNA-protein complexes that are capable of RNA synthesis in vitro (transcriptionally active chromosomes) were isolated from both chloroplasts and etioplasts of mustard (Sinapis alba L.) seedlings. Analyses of the polypeptide pattern of these complexes indicate that they comprise a specific subset of plastid proteins, distinct from the overall pattern of either the soluble or membrane-bound plastid proteins. DNA-protein complexes from the two plastid types have polypeptides in common. However, at least several polypeptides are highly enriched in either the chloroplast or the etioplast DNA-protein complex.

The EcoRI restriction endonuclease fragments of the DNA associated with the complexes from either plastid type are the same. They are identical with the fragments obtained from highly purified chloroplast DNA.

The transcriptional activity of the chloroplast complex is more than ten times higher than the activity of the etioplast complex. However, the complexes from either plastid type are capable of transcribing DNA regions containing genes for both the plastid rRNAs and for plastid proteins. In vitro transcripts were found to hybridize not only to DNA regions for mature in vivo RNA but also to adjacent regions, indicating synthesis of precursor RNA sequences by the transcriptionally active chromosomes.

Abbreviations
TAC

transcriptionally active chromosome

ptDNA

plastid DNA

SDS

sodium dodecyl sulfate

PAGE

polyacrylamide gel electrophoresis

Enzymes
 

RNA polymerase or nucleosidetriphosphate: RNA nucleotidyltransferase (EC 2.7.7.6)

 

T4 polynucleotide kinase or ATP: 5′-dephosphopolynucleotide 5′-phosphotransferase (EC 2.7.1.78)

 

restriction endonucleases BamHI (EC 3.1.23.6), EcoRI (EC 3.1.23.13), and PstI (EC 3.1.23.31)

 

ribulosebisphosphate carboxylase or 3-phospho-d-glycerate carboxylase (EC 4.1.1.39)

 

DNase I (EC 3.1.21.1)

 

proteinase K (EC 3.4.21.14)

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