Biology Reference
In-Depth Information
formation of further cycles of hormogonia so that the cyanobiont can differentiate heterocysts and
take up the function of nitrogen fi xation. To examine this aspect, Cohen
et al
. (1994) characterized
a transposon-induced mutant of
N
.
punctiforme
that is more (50-fold) infective than its wild type
culture. Two genes have been identifi ed that have bearing on the symbiotic potential of
N
.
punctiforme
.
These are
hrmA
(which is an unique sequence not represented in any database of gene sequences)
and
hrmU
that has similarity to the sequences of a NAD(P)-H-dependent oxidoreductases. Reporter
gene constructs of
hrmA
and
hrmU
have been induced by aqueous extract of
A
.
punctatus
. Since the
aqueous extract of
A
.
punctatus
eliminates the HIF-stimulated hormogonium formation, it is likely
that gene products of
hrmAU
operon may block hormogonium formation by producing an inhibitor
or catabolizing an activator (Meeks, 1998). The increased infectivity of the mutant is explained on the
basis that it undergoes hormogonium cycle even in presence of inhibitor/repressor of HIF (Cohen
and Meeks, 1997). Campbell
et al
. (2003) reported the presence of a repressor gene (
hrmR
) within
the
hrm
locus. Sequence analysis of
hrm
locus of
N
.
punctiforme
revealed the existence of four more
genes besides
hrmU
and
hrmA
. These are
hrmI
,
hrmR
,
hrmK
and
hrmE
with two unknown short
ORFs in between
hrmK
and
hrmE
that code unknown proteins. The gene sequences of
hrmR
and
hrmI
are transcribed in the same direction as 5' of
hrmU
. The ORFs
hrmK
and
hrmE
are transcribed
in the opposite direction from the
hrmRIUA
cluster. The four additional genes (
hrmI
,
hrmR
,
hrmK
and
hrmE
) identifi ed have similarity to genes encoding enzymes of carbon metabolism. The protein
transcripts of
hrmA
and
hrmU
genes have shown similarities to hexuronic acid metabolism of certain
heterotrophic bacteria. Thus the gene products of the four genes, i.e.
hrmE
,
hrmK
,
hrmR
and
hrmI
have 55%, 49%, 39% and 36% similarities to aldehyde reductase, gluconate kinase, transcriptional
receptors
LacI
/
GalR
family, uronate isomerase, respectively. The gene product of
hrmU
has 57%
and 55% similarities to 2-keto-3-deoxygluconate and mannonate oxidoreductase, respectively. The
product of
hrmR
gene has been shown to be a DNA-binding protein that regulates the transcription
of its own gene and a near by gene
hrmE
. The activity of the repressor was inhibited by galacturonate
or the lysate from induced
N
.
punctiforme
cells suggesting that the binding of repressor is modulated
by a sugar molecule. A model for explaining the regulation of genes in the
hrm
locus by plant signals
has been proposed by Campbell
et al
. (2003).
Mutants of
N
.
punctiforme
unable to differentiate heterocysts (
Het
-
) were tested for their ability to
establish a symbiotic association with
A
.
punctatus
in cultures. Three mutants tested were defective
in
ntcA
,
hetR
and
hetF
regions.
NtcA
controls the transcription of a number of genes in the course of
heterocyst differentiation (Herrero
et al.
, 2001; Fiedler
et al
., 2001).
HetR
is the fi rst heterocyst-specifi c
gene and is considered to be the primary activator of heterocyst differentiation and is indirectly
dependent on
ntcA
(Wolk, 2000).
HetF
is a positive activator of heterocyst differentiation that helps
in the enhanced transcription of
hetR
(Wong and Meeks, 2001). Among these, mutants defective in
hetF
and
hetR
regions infected
A.
punctatus
with similar frequency as wild type but did not support
the diazotrophic growth of the plant partner. Thus functional
hetF
and
hetR
regions along with
certain heterocyst regulatory elements are required for heterocyst differentiation and nitrogenase
expression in both free-living and symbiotic states. A mutant defective in
ntcA
region was unable
to infect
A
.
punctatus
though produced hormogonia at a low frequency. The ability for production
of hormogonia and infection of the host matched that of wild-type when mutants defective in
ntcA
were complemented with functional copies of
ntcA
. Thus it is clear that a functional
ntcA
is essential
for hormogonia formation, infection and a stable symbiosis to ensue. Chapman
et al
. (2008) reported
that muation of the
cyaC
gene that encodes the multidomain adenyl cyclase enzyme had differential
symbiotic competence in
N
.
punctiforme
. An omega neomycin phosphotransferase gene with pSCR19
was ligated to the 3'- and 5'-ends of the
cyaC
gene to obtain mutants C3212 and C1068, respectively.
