Biology Reference
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the view based on
patS
expression in proheterocysts and mature heterocysts. When
patS
promoter
was fused with genetically engineered
gfp
[that has the ability to move from cytoplasm into the
periplasm of proheterocysts and heterocysts due to the presence of twin-arginine signal sequence
(ss)] as the reporter gene and introduced into
Anabaena
sp. PCC 7120, fl uorescence could be detected
in the periplasm of not only the proheterocysts and heterocysts but also in the periplasm of adjacent
vegetative cells. According to these workers, periplasm continuity can form a suitable conduit for
the transport of substances into heterocysts and vegetative cells and vice-versa (Mariscal
et al
., 2007).
On the other hand, Zhang
et al
. (2008) put forward the view that there exists a periplasmic barrier
for GFP to be transported from cell-to-cell in
Anabaena
sp. strain PCC 7120. They chose three genes,
two of them (
hepA
expressed around 5-10 h and
patB
expressed around 18 h) exclusively expressed
in proheterocysts and heterocysts and the third gene
rbcL
expressed in vegetative cells (but turned
off in heterocysts at late maturation phase) and the respective plasmids with gene constructs
P
hepA
-
ssgfp
, P
patB
-
ssgfp
and P
rbcL
-
ssgfp
were introduced separately into wild-type
Anabaena
sp. strain PCC 7120.
As controls similar gene constructs without
ss
for
gfp
have been used. Through techniques such as
FRAP (fl uorescence recovery after photobleaching) and FLIP (fl uorescence loss in photobleaching),
the green fl uorescence has been observed only in proheterocysts or heterocysts and no intercellular
transfer through periplasm could take place. Likewise, in vegetative cells green fl uorescence protein
could not move into proheterocysts or heterocysts despite possessing signal sequence (Zhang
et al.
,
2008). To understand the mechanism of intercellular molecular exchange, Mullineaux
et al.
(2008)
employed non-fl uorescent acetoxymethylester derivative, calcein that reacts with endogenous
esterases to form a fl uorescent hydrophilic product. Once taken up by the cells if calcein is not
exchanged in between the cells, the fl uorescence of calcein should remain constant but if there is a
rapid transfer of calcein from one cell to the other the decrease and increase of fl uorescence could
be observed. For determining the rate constants of calcein exchange, they employed
Anabaena
sp.
strain PCC 7120 cells grown in nitrate medium and after nitrogen step-down. Visualization of
calcein fl uorescence by FRAP followed by their imaging through a confocal microscope enabled
them to suggest that (i) there exists a rapid exchange of calcein in between vegetative cells; (ii) the
fl uorescence in vegetative cells adjacent to heterocysts remained fairly constant due to the presence
of polar nodules; (iii) there is no calcein fl uorescence in the periplasm of the cells; (iv) the exchange
of calcein only takes place in between the vegetative cells of heterocystous cyanobacteria but not in
between vegetative cells of non-heterocystous cyanobacteria like
Oscillatoria
; (v)
A
.
variabilis
mutant
lacking cyanophycin synthase does not form polar nodules in its heterocysts and calcein exchange
in between vegetative cells and heterocysts has been found to be much faster than in wild-type; (vi)
they suggested that the likely candidate suitable for forming pore-like structures at cell junctions
occupied by microplasmodesmata is FraG (or SepJ the product of gene
alr2338
), the channel-forming
protein; (vii) mutants of FraG showed negligible exchange of calcein fl uorescence in between the
vegetative cells; and (viii) FraG constitutes part of molecular machinery required for intercellular
molecular exchange (Mullineaux
et al
., 2008). Flores
et al
. (2007) characterized a
sepJ
insertion mutant
that exhibited extensive fragmentation without heterocyst development and nitrogen fi xation even
under anoxic conditions. The SepJ protein is thus shown to be necessary for fi lament integrity and its
localization at the intercellular septa has been confi rmed by the expression of
gfp
as a reporter gene in
the wild-type. Summing up cell-cell communication in fi lamentous cyanobacteria, Haselkorn (2008)
cautioned that while the evidences putforward by Zhang
et al
. (2008) and Mullineaux
et al
. (2008)
appear to be convincing we should wait more for confi rmation from structural studies combined
with fl uorescence involving FraG and other subunits that might constitute the microplasmodesmata.
The other component proteins that help FraG in getting organized into microplasomdesmata have