Plant Molecular Biology 34: 181–189, 1997.
1997 Kluwer Academic Publishers. Printed in Belgium.
Transcription factor phosphorylation by a protein kinase associated with
chloroplast RNA polymerase from mustard (Sinapis alba)
Sacha Baginsky, Kai Tiller and Gerhard Link
University of Bochum, Plant Cell Physiology and Molecular Biology, Universit
atsstrasse 150, 44780 Bochum,
author for correspondence)
Received 28 August 1996; accepted in revised form 10 February 1997
Key words: DNA-dependent RNA polymerase, chloroplast gene expression, phosphorylation cascade, protein
kinase, sigma-like transcription factors
The chloroplast transcription machinery involves multiple components with both catalytic and regulatory func-
tions. Here we describe a serine-speciﬁc protein kinase activity that is associated with the major chloroplast RNA
polymerase and phosphorylates sigma-like transcription factors in vitro. The kinase activity can be assigned to a
54 kDa polypeptide of partially puriﬁed RNA polymerase (KPC, kinase polymerase complex). This polypeptide
is also present in a smaller complex that contains several putative polymerase subunits and reveals kinase activity
but lacks transcription activity (KC, kinase complex). Although the 54 kDa component could not be chromato-
graphically separated from the rest of this complex without loss of activity, it retained residual kinase activity
in an electrophoretic blot assay. The polymerase-associated kinase is itself affected by in vitro phosphorylation
and dephosphorylation, which raises the possibility that it is part of a signalling cascade that controls chloroplast
transcription in vivo by factor phosphorylation.
Chloroplasts, like other plastids,are semi-autonomous
DNA-containing cell organelles speciﬁc to plants.
Dark-grown tissue may develop etioplasts which upon
exposure to light become photosynthetically active
chloroplasts. Plastid geneexpression,duringdevel-
opment and function, is controlled at levels ranging
from translation and protein stability to RNA matura-
tion and transcription [20, 21, 23, 35, 43, 48, 56].
The chloroplast transcription apparatus, which has
been intensely studied by combined biochemical and
molecular-genetic approaches (for reviews, see [6, 25,
32, 33, 39]), has turned out to be much more complex
than anticipated. One aspect that has recently received
increasing attention is the presence of multiple forms
of organellar RNA polymerases [2, 31, 37, 40, 55,
57]. Likewise, multiple speciﬁcity factors have been
reported (for reviews, see [6, 33]), including several
that resemble bacterial sigma factors [7, 30, 34, 50,
For instance, three sigma-like factors (SLFs) were
found to be present in both chloroplasts and etioplasts
from mustard (Sinapisalba) [51, 52]. The correspond-
ing factors from the two plastid types differ, however,
in their properties, which are related to the phos-
phorylation state of these proteins. For two of these
, pretreatment in vitro with
animal protein kinase and phosphataseled to intercon-
version of ‘chloroplast’ into ‘etioplast’ characteristics
and vice versa .
Chloroplasts contain a number of protein kinases
known to phosphorylate both membrane-bound and
soluble organellarproteins, includingseveralcompon-
ents of the photosynthetic apparatus [4, 19, 41, 42].
Here we show that a chloroplast transcription factor
can be a target for an organellarprotein kinase activity
that is associated with the transcriptionapparatus. This
kinase could be a key element in the control of plastid
gene transcription in vivo.
GR: 201001907, Pips nr. 135297 BIO2KAP
plan3659.tex; 7/05/1997; 12:59; v.7; p.1