1021-4437/01/4804- $25.00 © 2001
Russian Journal of Plant Physiology, Vol. 48, No. 4, 2001, pp. 459–463. Translated from Fiziologiya Rastenii, Vol. 48, No. 4, 2001, pp. 536–541.
Original Russian Text Copyright © 2001 by Rosov, Shleev, Petrova, Tsyganov, Borisov, Topunov, Tikhonovich.
Nitrogen metabolism is one of the key problems of
plant-cell biology. Plants are constantly in short nitro-
gen supply and consume it rather frugally. Most
legumes (Fabaceae) are capable of ﬁxing atmospheric
nitrogen in the symbiosis with nodular bacteria (Rhizo-
This symbiosis is very susceptible to various stress
factors that drastically reduce the rate of nitrogen ﬁxa-
tion. The sequence of nodule responses to any stressful
factor can be described in the framework of the classi-
cal stress concept . Studying the adaptive changes
that occur in response to stress can help in understand-
ing the mechanisms controlling the normal process of
The effects of nitrate stress on the nitrogen-ﬁxing
symbiosis are under active investigation. Exogenous
nitrate is well known to suppress the nodule formation,
their development, and nitrogen-ﬁxing capacity [2–7].
Long-term legume exposure to high nitrate concentra-
tions results in nodule senescence . This treatment
increases the content of endogenous nitrogen, and,
within several hours, the metabolism switches from the
ﬁxation of atmospheric nitrogen to nitrate consump-
tion. The mechanism of this switching and its regula-
tion are yet to be fully elucidated. However, several
hypotheses have been put forward concerning the sup-
pression of the nodule nitrogen-ﬁxing capacity by high
nitrate concentrations [9, 10].
In the nodule cortex, a so-called “gas-diffusion bar-
rier”  is known to preclude the oxygen diffusion
into the central nodule zone. This barrier creates local
anaerobic conditions favorable for nitrogen ﬁxation.
The resistance of this barrier to the oxygen ﬂux is vari-
able : it can be affected by various environmental
factors, a high nitrate level in particular.
According to the ﬁrst group of hypotheses, the fol-
lowing sequence of events is supposed to take place.
Nitrate somehow increases the resistance to the oxygen
ﬂux into the nodule, and, as a result, oxygen deﬁciency
arises, resulting in a shortage of ATP production and
the limitation of nitrogenase activity . A decrease in
nitrogenase activity can also result from a disturbance
in the oxygen-transporting system within the infected
cells containing bacteroids, i.e., the modiﬁed bacterial
cells. This system includes an oxygen-carrying hemo-
protein leghemoglobin. Only its reduced form is capa-
ble of oxygen transport.
According to the second group of hypotheses,
nitrate reductase, whose activity is high under nitrate-
stress conditions , catalyzes the formation of nitrite
and nitrogen oxide (NO), which either oxidize Lb or
produce a stable complex with it. In both cases, Lb
looses its capacity to carry oxygen. At present, the sup-
pression of nitrogenase activity due to Lb inactivation
is the least studied stage.
Some evidence is available concerning a direct toxic
nitrate effect on the nitrogen-ﬁxing bacteroids .
Gene Responsible for Bacteroid Differentiation
Is Involved in Nitrate-Dependent Nodule Formation in Pea Plants
F. N. Rosov*, S. V. Shleev*, N. E. Petrova*, V. E. Tsyganov**,
A. Yu. Borisov**, A. F. Topunov*, and I. A. Tikhonovich**
*Bach Institute of Biochemistry, Russian Academy of Sciences, Leninskii pr. 33, Moscow, 117071 Russia;
fax: 7 (095) 954-2732; e-mail: firstname.lastname@example.org
** All-Russia Research Institute of Agricultural Microbiology, Russian Academy of Agricultural Sciences,
sh. Podbel’skogo 3, Pushkin, St. Petersburg, 189680Russia
Received June 5, 2000
—The effects of exogenous nitrate on the number of developing nodules and their leghemoglobin con-
tent in the original pea (
L.) line and its symbiotic mutants were studied. Mutation in the
gene conferred the tolerance to nitrate in the corresponding pea line and manifested itself as the number of nod-
ules independent of the nitrate concentration. Thus, the
gene was identiﬁed as the only known symbiotic
gene involved in both the differentiation of symbiotic compartments and the nitrate-dependent process of nod-
ule formation. The presence of leghemoglobin in double mutants (
) indicates the possibility of
the complementary contribution of these genes in the control of leghemoglobin synthesis.
Key words: Pisum sativum - nodule - symbiotic genes - mutants - nitrate - leghemoglobin
: Lb—leghemoglobin; PMSF—phenylmethylsulfo-