636
Chlamydomonas
structure. Numerous mutants affected in
psaA
trans-splicing have been isolated and
can be grouped into three classes. In the
Frst are mutants unable to splice exons 1
and 2, the second comprises mutants un-
able to splice exons 2 and 3, and the third
consists of mutants unable to perform ei-
ther trans-splicing reaction. At least 14
nuclear loci are involved in this complex
maturation pathway. Whether the unusual
split structure of the
psaA
gene reflects
an ancient gene structure or whether it
was created through division of an ances-
tral
psaA
gene by DNA rearrangements
r
em
a
in
sanop
enqu
e
s
t
ion
.Th
enu
c
l
e
a
r
genes of several of the factors involved in
trans-splicing have been cloned and char-
acterized. Some of them resemble known
proteins involved in RNA metabolism such
a pseudouridine synthases, although the
enzymatic activity is not required for trans-
splicing. Whether these proteins have a
dual role in chloroplast RNA maturation
is not yet clear. Some of these factors are
part of high molecular weight RNA protein
complexes that are reminiscent of snRNPs
in eukaryotic cells, although no signiFcant
sequence identity is apparent between the
subunits of these complexes.
Mutants affected in chloroplast trans-
lation have also been examined. These
mutants were Frst identiFed on the basis
of pulse labeling of cells in the pres-
ence of an inhibitor of cytoplasmic protein
synthesis. Under these conditions, only
polypeptides translated in the chloroplast
compartment are labeled. ±rom the label-
ing patterns, it is, however, difFcult to
distinguish mutants truly affected in trans-
lation from those having increased protein
turnover. By using a strategy resembling
the one based on the use of chimeric genes
outlined earlier for the nuclear mutant
deFcient in
psbD
RNA accumulation, it
has recently been shown that some of
the mutants are indeed affected at the
level of initiation of translation. Several of
the trans-acting factors have been recently
identiFed (Table 2).
4.1.3
Metal Ion Control of Photosynthetic
Gene Expression
The
copper-containing
protein
plasto-
cyanin catalyzes electron transfer between
the cytochrome
b
6
f
complex and photo-
system I (±ig. 5). Growth of terrestrial
plants in copper-deFcient habitats leads to
symptoms of copper deFciency. However,
Chlamydomonas
, like other aquatic algae
and cyanobacteria, remains photosynthet-
ically competent under copper-deFcient
growth conditions. This is because, in
theabsenceo
fcopper
,thea
lgaexpresses
the heme-containing cytochrome
c
6
,an
alternate electron carrier to plastocyanin.
Although plastocyanin is still expressed
under conditions of copper deFciency, the
apoprotein is highly unstable and is rapidly
degraded. Expression of cytochrome
c
6
proceeds only in the absence of copper
and is regulated at the transcriptional level.
Thecupricion–responsivepromoterofthe
cytochrome
c
6
gene displays unique metal
speciFcity and high sensitivity; in addition,
it governs reciprocal control of synthe-
sis of two proteins, cytochrome
c
6
and
plastocyanin, which are distinct although
functionally equivalent. On the basis of
the observation that the amount of cop-
per ions needed to repress cytochrome
c
6
transcription matches the amount of
plastocyanin in
Chlamydomonas
cells, it
has been proposed that repression of tran-
scription requires a copper-binding factor
that is titrated by copper only after plasto-
cyanin has accumulated to a level needed
for photosynthesis. Therefore, the con-
trol of cytochrome
c
6
expression results
mainly from a direct cellular sensing of
available copper rather than indirectly as
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