Journal of Experimental Botany, Vol. 55, No. 394, pp. 11±25, January 2004
DOI: 10.1093/jxb/erh011 Advanced Access publication 28 November, 2003
FOCUS PAPER
Global aspects of C/N interactions determining
plant±environment interactions
John A. Raven1,2,*, Linda L. Handley2,3,² and Mitchell Andrews4
1
Division of Environmental and Applied Biology, School of Life Sciences, University of Dundee, Dundee DD1
4HN, UK
2
Columbia University Biosphere 2 Center, 32540 S. Biosphere Road, PO Box 689, Oracle, AZ 85623, USA
3
Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, UK
4
School of Sciences, University of Sunderland, Sunderland SR1 3SD, UK
Received 2 May 2003; Accepted 11 August 2003
Abstract
The atomic C:N ratio in photolithotrophs is a function
of their content of nucleic acids, proteins, lipids, polysaccharides, and other organic materials, and varies
from about 5 in some protein-rich microalgae to much
higher values in macroalgae and in higher plants with
relatively more structural and energy storage materials. These differences in C:N ratios among organisms
means that there is more N assimilation by photosynthetic organisms in the oceans than on land despite
the near equality of global photosynthetic C assimilation rates in the two environments. Aquatic organisms
obtain inorganic C and inorganic N from the surrounding water. Terrestrial photolithotrophs obtain inorganic C, dinitrogen (by diazotrophy) and some
combined N from the atmosphere, with the remaining
combined N coming from the soil. The nitrogen cost
of growth (biomass production rate per unit plant N)
varies with the C:N ratio and speci®c growth rate of
the organism. The C:N ratio of plants can be increased
with no, or minimal, decrease in growth rate by
switching from N-containing to N-free solutes
involved in, for example, UV-B screening or by reducing the content of particular proteins. The water cost
of growth (water lost per unit biomass gain) in terrestrial plants is a function of N supply and of C supply;
water cost is lower with higher N and C availability.
Water supply is also important in determining denitri®cation rates on land and on N (and C) ¯uxes from terrestrial to aquatic systems.
Key words: Agriculture, algae, astrochemistry, carbon,
chemical defence, compatible solutes, ecology, evolution,
free radical scavenging, nitrogen, UV-B screening.
Introduction
Carbon and nitrogen are major components of all living
organisms, and their metabolism is intimately linked. In
this paper a number of questions about the interactions of
carbon and nitrogen in plant metabolism are addressed. (1)
What features of carbon and nitrogen relate to their
biological roles? (2) What are the major carbon- and
nitrogen-containing components of plants and how do they
vary quantitatively among plants? (3) What possibilities
are there for altering the content of these components
without compromising plant growth and, for crop plants,
the quality and yield of the harvested product?
These questions are considered in a wider context. In
exploring the evolutionary background to the roles of
carbon and nitrogen in organisms and its implications for
their functioning, the genesis of carbon and nitrogen in
stars and their partitioning to our solar system and,
especially, to the Earth's surface is reviewed ®rst. The
chemical properties of carbon and nitrogen in the context
of what is required of life `as we know it' then follows. The
quantitative requirements for carbon and nitrogen in
photosynthetic organisms of different life forms are then
considered, and special attention is given to the extent to
which the carbon:nitrogen ratio can be varied, with
emphasis on economizing on the use of nitrogen and
* Permanent address and to whom correspondence should be sent: Division of Environmental and Applied Biology, School of Life Sciences, University of
Dundee, Dundee DD1 4HN, UK. Fax: +44 (0)1382 344275. E-mail: [email protected]
²
Permanent address: Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, UK.
Journal of Experimental Botany, Vol. 55, No. 394, ã Society for Experimental Biology 2004; all rights reserved
12 Raven et al.
what effects such restriction of nitrogen content might
have for growth under natural conditions. Finally, the
requirements for carbon and nitrogen are related to the
habitats of the organisms, and how the acquisition and
processing of carbon and nitrogen impact on the requirements for other resources.
The origins and cosmic distribution of carbon
and nitrogen and their compounds
The `Big Bang' only produced elements of low atomic
number, mainly H and He. Nucleosynthesis of elements
larger than this, including carbon and nitrogen, occurs in
stars (Williams and FrauÂsto da Silva, 1996, 2003; Henning
and Salama, 1998; Beers, 2003). Thus, the availability of
carbon and nitrogen and of planets made from these and
higher atomic number elements, required one cycle of now
extinct stars which generated these elements. Nitrogen is
the ®fth most abundant element in the universe, and carbon
is the sixth.
Some of the carbon and nitrogen in the universe is in
stars and planets; the rest is in interstellar space.
Spectroscopic studies on interstellar molecules show a
large range of compounds containing carbon and/or
nitrogen (Henning and Salama, 1998; Thaddeus et al.,
1998; Ehrenfreund and Charnley, 2000).
Much of the interstellar organic carbon is in polyaromatic hydrocarbons and their relatives the buckminsterfullerenes. However, there are also many other molecules
such as CO, CO2, CH4, CH2O, and CH2OHCOOH.
Nitrogen occurs as N2, N2O and NH3. Interestingly for
the subject of this paper there are also compounds
containing both C and N. Examples are HCN and the
homologous series of cyanopolyines HC1+2nN where n is 0
(HCN), 1 (HC3N), 2 (HC5N), 3 (HC7N), 4 (HC9N), and 5
(HC11N). HC11N is relatively abundant and has the
formula H-C º C-C º C-C º C-C º C-C º C-C º N. Of
more biological relevance are compounds such as CH3NH2
and, much less certainly, CH2(NH2)COOH. These data
show that, even in interstellar space, photochemistry
produces molecules containing both carbon and nitrogen.
Coming down to Earth, it is found that this planet is
depleted in carbon and nitrogen relative to many heavier
elements when compared to cosmic abundance; even
phosphorus is more abundant on Earth than are carbon and
nitrogen. Attempts to simulate in the laboratory the
mechanisms of prebiotic productions of carbon and
nitrogen compounds go back 50 years (Miller, 1953).
These experiments showed that organic molecules, including glycine and alanine, could be produced in relatively
high yield when an electric discharge energized molecules
in a mixture of gases which was thought to be representative of the prebiotic atmosphere of the Earth. Later
experiments involved UV as the energy source, and, when
HCN was a component of the gas phase, the production of
adenine. However, the gas mixture used by Miller (1953),
i.e. CH4, CO2, NH3, and H2O, was more reducing than
what is now believed to have existed in prebiotic times on
Earth ~4 Ga ago. Repetition of Miller's experiments with
these less reducing atmospheres gave much smaller yields
of organic compounds. Current thinking about the origin of
carbon- and nitrogen-containing compounds on Earth
emphasizes inputs from comets and syntheses by chemolithotrophic processes, for example, at hydrothermal vents.
To summarize, the cosmic abundance of carbon and
nitrogen is high, and, despite the Earth being depleted in
these elements relative to their cosmic abundance, carbon
and nitrogen are relatively abundant in the Earth's
atmosphere, hydrosphere and crust.
The availability of carbon and nitrogen to
photosynthetic organisms over the last 3.5 Ga
The relative availability of carbon dioxide (more generally, inorganic carbon) for photosynthesis and of combined
nitrogen over the last 3.5 Ga in which photosynthetic O2evolvers may have existed is discussed by Falkowski and
Raven (1997), Falkowski (1997), Falkowski et al. (1998),
Raven and Yin (1998), Stewart and Schmidt (1999),
Navarro-Gonzalez et al. (2001), and Anbar and Knoll
(2002); for an overview of biogeochemistry see Berner and
Berner (1996). Carbon dioxide availability has decreased
over the last 3.5 Ga, although the fall has not been
monotonic and there have been a number of ¯uctuations,
with low atmospheric carbon dioxide in the late Devonian±
Permian and during glacial episodes in the Quaternary
(Falkowski and Raven, 1997).
Combined nitrogen (in this case as NO and NO2)
availability from electric storms is thought to have
diminished during the period in the Archean in which the
atmosphere became oxidized and carbon dioxide was
decreasing (Navarro-Gonzalez et al., 2001). Anbar and
Knoll (2002) suggested that a Palaeo- and MesoProterozoic sulphidic ocean could have limited molybdenum availability for biological nitrogen ®xation, and cite
15 14
N/ N natural abundance values to support restricted
biological diazotrophy. However, the possibility of the
activity of `alternative' nitrogenases using vanadium or
only iron, and with 15N/14N discrimination values greater
than those normally cited (Rowell et al., 1998), should not
be ignored.
There is a widespread view that primary productivity is
restricted by the availability of combined nitrogen
ÿ
ÿ
NH‡
4 ; NO3 ; NO2 , and organic N) in many terrestrial
and aquatic habitats today (Falkowski, 1997, 2000;
Falkowski and Raven, 1997; Falkowski et al., 1998;
Field et al., 1998). Nitrogen and carbon co-limitation can
occur in terrestrial C3 plants, with increases in atmospheric
carbon dioxide above the present 370 mmol mol±1 yielding
increased primary productivity, provided nitrogen (or
Global aspects of C/N interactions 13
other resources, including water and light) is not in too
limiting supply (Andrews et al., 2001). Geochemically,
phosphorus is the element which is expected ultimately to
limit primary productivity, and such limitation now occurs
in some areas (Red®eld, 1958; Beadle, 1966; Francis and
Read, 1994; Wu et al., 2000). Furthermore, there is
evidence that phosphate availability limits diazotrophy by
the cyanobacterium Trichodesmium in the south-west
North Atlantic (SanÄudo-Wilhelmy et al., 2001).
However, elsewhere in the ocean, iron limitation of
diazotrophy is likely (Falkowski, 1997, 2000; Falkowski
and Raven, 1997; Falkowski et al., 1998; Field et al.,
1998). In some parts of the ocean, where combined
nitrogen and phosphorus are relatively abundant yet
primary productivity is low, it seems that iron is limiting
the use of combined nitrogen in algal photosynthesis. Fe
de®ciency does not, however, decrease the nitrogen
contents relative to the carbon contents of phytoplankton
growing by N2-®xation (Berman-Frank et al., 2001) or by
use of combined nitrogen (Green et al., 1991; La Roche
et al., 1993; Flynn and Hipkin, 1999; De La Rocha et al.,
2000; Milligan and Harrison, 2000; Davey and Geider,
2001).
The evidence that has just been discussed shows that
nitrogen ®xation is a process which is relatively restricted
in extent, and that this results in frequent nitrogen
limitation of growth of photosynthetic organisms in land
and in the sea. This can be illustrated by comparing global
carbon and nitrogen assimilation in net primary productivity with the assimilation of inorganic carbon and of
atmospheric N2 (Table 1). Even taking into account inputs
of `new' combined nitrogen from processes other than
biological N2 ®xation, for example, lightning, `arti®cial'
fertilizers, and NOx resulting from high-temperature
combustion of fossil fuels (Table 1), it can be seen that
the nitrogen requirements for photosynthetic primary
production are largely met by recycled combined nitrogen
rather than N2 ®xation. Combined nitrogen is removed by
denitri®cation which generates N2, which can only be
reconverted biologically to combined nitrogen by N2
®xation, and N2O, which is a form of combined nitrogen
that is not generally available as a nitrogen source for
primary producers. The loss of combined nitrogen by
denitri®cation is generally not directly controlled by
primary producers, an exception being the generation of
anoxic waterlogged ecosystems in areas with high precipitation relative to evaporation. Loss of combined
nitrogen by denitri®cation, and the cost in other resources
of biological N2 ®xation discussed in the preceding
paragraph and by Vitousek et al. (2000), restricts the size
of the combined nitrogen pool in many habitats, leading to
a combined nitrogen famine despite the apparent potential
for a nitrogen feast using N2.
Overall, it appears that nitrogen is globally `more
limiting' than is carbon for photosynthetic organisms, and
that economizing on nitrogen use may be more signi®cant
in natural selection than is economizing in carbon use. The
amounts of carbon and nitrogen that are used in biology
and how the use of the elements can be substituted by other
elements will be discussed next.
The roles of carbon and nitrogen in living
organisms: background
The properties of certain elements which have been
capitalized upon by life on Earth are reviewed by
Williams and FraÂusto da Silva (1996, 2003). Carbon is
unique in forming stable chains and rings of the element;
this is the basis of organic chemistry, and nitrogen
participates very strongly in both aliphatic and aromatic
organic chemistry, and is also important in hydrogen
bonding and in forming ligands with metals. The diversity
Table 1. Global biological production of organic C and of combined N, biological assimilation of combined N, and inputs of
combined N, in Tmol of C or N per year
a
Net primary inorganic C assimilation by photosynthetic organisms
Net primary N2 and combined N assimilation by photosynthetic organisms
Combined N input from lightning
Combined N input from combustion
Combined N input as fertilizer
Combined N input as biological N2 ®xation
a
Oceans
Land (and fresh water)
4040
577b
0.36d
0e
0f
2.5g
4700
209c
0.36d
1.4e
3.6 f
10g
From Field et al. (1998).
From C assimilation data of Field et al. (1998) and a C:N ratio of 7 (Raven et al., 1992a). Chemolithotrophic inorganic C assimilation globally each
year is of the order of 1% of the photosynthetic assimilation (Raven, 1996; Wolstencroft and Raven, 2002).
c
From C assimilation data of Field et al. (1998) and a C:N ratio of 15 for herbs and 50 for trees (Raven et al., 1992a).
d
From Lynch and Hobbie (1998) assuming equal inputs to land and sea.
e
From Lynch and Hobbie (1998); most combustion of fossil fuels occurs on land but signi®cant deposition of NOx occurs into the sea.
f
From Lynch and Hobbie (1998); all fertilizer N input is to land, but some fertilizer N reaches the sea via rivers. Other estimates of fertilizer N are
higher, for example, 5.7 Tmol N from http.//www.fertilizer.org
g
From Lynch and Hobbie (1998). The N2 ®xation on land is about 6.4 Tmol N from agricultural land and 3.6 Tmol N from forest and non-agricultural
land.
b
14 Raven et al.
of organic chemistry, with elements such as oxygen and
nitrogen, gives further possibilities for hydrogen bonding.
For biology, organic chemistry has the advantage that
most reactions do not occur at a signi®cant rate at
temperatures permitting the existence of liquid water
within the range of pressures found in habitats on Earth.
This means that catalysis is essential for organic biochemistry, allowing regulation of the rate and products of
reactions (Williams and FraÂusto da Silva, 1996).
As a background to the following discussion of the
qualitative and quantitative roles of carbon and nitrogen in
photosynthetic organisms, it was noted that the atomic C:N
ratio varies from ~ ®ve in microalgae and cyanobacteria to
more than 100 in woody plants (Sterner and Elser, 2002).
Nitrogen-containing macromolecules: DNA, RNA
and protein
The core of molecular biology involves the organic
chemistry of nitrogen. Fifty years ago Watson and Crick
(1953) presented the structure of the double helix of DNA,
with a crucial role for hydrogen bonds involving nitrogen
as well as oxygen in forming the double helix and hence in
replication, transcription and repair. X-ray crystallographic studies of proteins showed the role of nitrogenrelated hydrogen bonds and electrostatic interactions
bound in the alpha helix and in the beta pleated sheet.
Rather later, studies of the structure of rRNA, tRNA and
mRNA also emphasized the role of hydrogen bonds
involving nitrogen in the bases in maintaining the
secondary structure and in transcription and translation.
To these qualitative needs for nitrogen and carbon in the
components of the central dogma there is, in terms of
overall requirement for carbon and nitrogen by organisms,
the question of how many kinds of genes and hence of gene
products occur in an organism, how large the genes and
hence the gene products are, and how much of each gene
product is produced under the growth conditions that are
being considered. Determinants of these variables are
known in general terms (Maynard-Smith and SzathmaÂry,
1995), but it is less easy to be speci®c. Even the
determinants of the number of protein amino-acids (21,
including selenocysteine) are still a matter for debate
(Maynard-Smith and SzathmaÂry, 1995). Only perhaps four
or ®ve amino-acid residues can be directly involved in the
active centre of an enzyme, and some of the rest of the
amino-acid residues (of the order of 100) have regulatory
functions, for example, interactions with regulatory
ligands, and controlling and facilitating access of substrate(s) and removal of product(s) (Maynard-Smith and
SzathmaÂry, 1995; Seligmann, 2003). As to genome size
and gene number, the smallest fully sequenced genome of
a photosynthetic O2-evolver is that of the cyanobacterium
Prochlorococcus marinus, of which the MIT strain has a
genome size of 1.7 Mbp with 1500 genes (Hess et al.,
2001). Arabidopsis thaliana has a nuclear genome of 125
Mbp with 25 500 genes (The Arabidopsis Genome
Initiative, 2000). Arabidopsis thaliana was chosen for
sequencing because, inter alia, of its small genome size,
and most higher plant genomes are much larger even if
they do not code for more genes. Genomic obesity, for
example via retrotransposons, involves additional DNA in
the nucleus which is not involved in coding for genes that
are required for plant function.
Turning to the quantitative requirements for RNA for
plant function, most of the RNA in a cell is rRNA. In
heterotrophs (non-autotrophic bacteria, fungi, metazoa)
the rRNA per cell in a given organism is a linear function
of the relative growth rate (Sterner and Elser, 2002).
However, there is a baseline rRNA content that is needed
for protein turnover even without growth. This requirement is considered in the context of mature leaves of
higher plants, where the turnover of the D1 (psbA gene
product) protein of photosystem II is a major component of
protein turnover (Raven et al., 2002a).
For growing cells of photosynthetic O2-evolvers, for
example, unicellular algae, and the meristems of higher
plants, there are few data on the relationship of cellular
rRNA content to growth rate. There is certainly not the
monotonic, linear, increase in rRNA per cell with relative
growth rate that is found in heterotrophs (Laws et al., 1983;
Thomas and Carr, 1985; Binder and Liu, 1998; Binder,
2000). Using light as the limiting resource, Laws et al.
(1983) and Thomas and Carr (1985) found an RNA content
which was independent of growth rate for a number of
eukaryotic microalgae. Binder and Liu (1998) and Worden
and Binder (2003) (cf. Lepp and Schmidt, 1998; Binder,
2000) measured rRNA content in two strains of marine
Synechococcus and in a strain of the marine
Prochlorococcus, and found a linear relationship between
rRNA content and growth rate over a restricted, intermediate, range of growth rates for these three strains of
cyanobacteria. At lower growth rates the rRNA content
was invariant with changes in growth rate, while at the
highest growth rates the rRNA content decreased with
increasing growth rate. For the relatively small range of
O2-evolving organisms examined, it appears that the
speci®c reaction rate of rRNA, rather than rRNA content,
increases with growth rate, so that cells are over-provided
with rRNA at low growth rates.
As for the protein content, it is clear that the speci®c
reaction rate of enzymic, and other catalytic (e.g. lightharvesting pigment±protein complex) reactions, in photosynthetic organisms generally increases with growth rate,
so that rapidly growing organisms of a given genotype
have a lower protein cost of growth (biomass increase per
unit protein in existing biomass per unit time: Raven,
1984a, b; Berends and Aerts, 1987; Agren and Bosatta,
1996; Andrews et al., 1999). This is despite, and sometimes because of, acclimatory changes in the content of
Global aspects of C/N interactions 15
speci®c proteins. Thus, acclimatory increases in the
content of light-harvesting pigment±protein complexes
with growth at lower photon ¯ux densities decreases the
speci®c reaction rate (photons absorbed per unit chromophore per unit time) because the increase in chromophores
does not offset the decreased photon ¯ux density (Raven,
1984a, b). Furthermore, if the increased chromophore
content increases the `package effect' (i.e. self-shading),
then the rate of absorption of photons per unit chromophore is further decreased. The capacity to absorb a greater
fraction of incident photons is obtained at the cost of
additional investment of nitrogen (Raven, 1984a, b).
Another aspect of variation in proteins related to light
harvesting is phylogenetic rather than acclimatory. Raven
(1984a, b) notes the larger protein content per unit
chromophore for the phycobilin light-harvesting pigment±
protein complexes of most cyanobacteria, all red and
glaucophyte algae, and cryptophyte algae. By contrast, the
protein per chromophore is lower in the chlorophyll- and
carotenoid-containing pigment±protein complexes of
some cyanobacteria and all cryptophytes in addition to
their phycobilins, and in all other O2-evolvers (Raven,
1984a, b). In organisms adapted or acclimated to low
photon ¯ux densities, the differences in protein allocation
per unit chromophore can make a signi®cant (up to 10%)
difference in overall protein content for an organism with a
phycobilin-based rather than a chlorophyll-based lightharvesting apparatus in an otherwise similar organism
(Raven, 1984a, b; MacIntyre et al., 2002; Ting et al., 2002;
contrast Sterner and Elser, 2002).
Better-known phylogenetic variations in protein allocation in photosynthetic organisms concern the different
quantities of Rubisco protein needed to sustain a given rate
of CO2 ®xation per unit leaf area in C3 plants and C4 plants
in air. The C4 plants need, and have, less Rubisco per unit
area of leaf, and attain the same or higher CO2 ®xation
rates per unit leaf area in air levels of CO2 because the CO2
concentration, and CO2/O2 ratio, at the site of Rubisco
activity is higher in C4 than in C3 plants. Even when the
extra protein needed in the C3+C1 carboxylation, and C4±
C1 decarboxylation cycles is accounted for, the C4 plants
still need less leaf N to attain a given rate of CO2 ®xation,
with the decreased content of enzymes of the photorespiratory carbon oxidation cycle as a contributory factor.
In the present context of C/N interactions, it is of interest
that transgenic rice (Orzya) and tobacco (Nicotiana) with
decreased content of Rubisco show altered N allocation in
the plants (Makino et al., 2000a, b; Matt et al., 2002; see
also Parry et al., 2003). Makino et al. (2000a, b) found that
the lower leaf N content when non-transformed rice plants
were grown in high, rather than ambient, CO2 concentrations did not show optimal N allocation, in that Rubisco
did not decrease relative to the decrease in total leaf N.
Optimal allocation requires that Rubisco content should
fall in C3 plants grown at high CO2, since the same
carboxylation rate can be sustained at high CO2 with a
lower Rubisco content (assuming equal activation states
under the two growth conditions). Rubisco-antisense rice
with 65% of wild-type Rubisco had 20% lower photosynthesis in normal atmospheric CO2 levels, but 5±15% higher
rates at saturated CO2. However, while the transgenic
plants had a higher N use ef®ciency for growth than did
controls after the seedling stage, in terms of overall
biomass production this was offset by slower growth in the
early stages of growth.
Matt et al. (2002) used Rubisco antisense technology to
decrease Rubisco expression in tobacco. The decreased
photosynthetic carbon assimilation caused an inhibition of
nitrogen assimilation, and altered primary and secondary
(chlorogenic acid; nicotine) metabolite levels for both
nitrogen-containing and nitrogen-free compounds.
Raven et al. (2002b) discuss other work showing that
down-regulation of a number of photosynthetic enzymes
other than Rubisco by 25% (or, in some cases, much more)
does not cause a decrease in the rates of photosynthesis and
growth under the conditions tested. However, such downregulations do not always lead to increased biomass
production rates because compensatory increases in other
proteins occur, for example, an increase of Rubisco when
Rubisco activase is down-regulated. Furthermore, there is
always the possibility that growth conditions which do
produce a phenotype of inhibited photosynthesis and
growth exist, but have not yet been tested. However, in
some cases there are large effects of small decreases in
activity of certain photosynthetic enzymes other than
Rubisco. The work of Henkes et al. (2001) shows for
antisense Nicotiana transformants that a small decrease in
plastid transketolase, which is involved in the oxidative as
well as the reductive pentose phosphate pathway, causes
not only decreases in the rate of photosynthesis over a wide
range of photon ¯ux densities but also decreased synthesis
of phenylpropanoids.
Raven et al. (2002c) also consider the possibility of
economizing on nitrogen by genetic manipulation of
rRNA. They conclude that there is an apparent overprovision of rRNA in mature leaves relative to what is
apparently needed for the turnover of D1 and of other
proteins.
More radical attempts to modify plant performance by
manipulating protein content is the expression of genes
from other organisms in a plant. One such procedure is the
expression of the bifunctional cyanobacterial enzyme
fructose-1,6-bisphosphate-1-phosphatase/sedoheptulose1,7-bisphosphate-1-phosphatase in higher plant plastids
which already possess separate enzymes with these two
phosphatase activities (Miyagawa et al., 2001). This
procedure increases the growth and photosynthetic rates
of Nicotiana at least under optimal conditions, although
effects on expression of the native phosphatases and on the
rate of biomass increase per unit plant nitrogen is not clear.
16 Raven et al.
Another procedure involved the expression of algal
Rubiscos with a higher CO2/O2 selectivity (e.g. from red
algae or diatoms) in higher plants. Complete replacement
of the native Rubisco with such algal enzymes could
increase the photosynthetic rate per unit nitrogen over the
entire light and CO2 range (Whitney and Andrews, 2001).
However, while the transcripts are produced in the
transformed plants, the translation products are not properly assembled in the stroma and do not function (Whitney
et al., 2001). By contrast, the gene for Form 1 Rubisco
from an alpha-proteobacterium which was transferred to
the plastid genome of Nicotiana was expressed and the
product assembled into a functional enzyme which could
support photosynthesis and growth. However, the bacterial
Rubisco has a very low selectivity for CO2 over O2, and
photosynthesis and growth of Nicotiana using this Rubisco
requires high CO2 concentrations (Whitney and Andrews,
2001).
Other roles for proteins include functions in the
cytoskeleton, including molecular motors. There does not
seem to be much scope for reducing the cellular content of
these protein components. Membrane lipids contain nitrogen in the polar head-group in the case of some, mainly
non-thylakoid, constituents.
Extracellular structural materials
Extracellular structural materials in photosynthetic organisms include some proteins, as well as other nitrogencontaining polymers, including peptidoglycan in cyanobacteria, and chitin in some diatoms and (in symbiosis) the
cell walls of fungal associates in lichens and mycorrhizas.
However, in general, photosynthetic organisms have
nitrogen-free carbohydrate polymers as their turgor (tension)-resisting components and invariably have (nitrogenfree) lignin as the component which resists compression
(Raven, 1993, 1995b, 2000).
Macromolecular and other insoluble compounds
storing energy and organic carbon
Storage polymers related to energy and organic carbon
storage include polysaccharides and triglycerides. Storage
proteins are found in many seeds, and the aspartate±
arginine polypeptide cyanophycin in cyanobacteria.
Attempts to ®nd speci®c vegetative storage proteins in
several plants have failed; there seem to be no speci®c
proteins used during vegetative dormancy.
Substitution of other elements for carbon and
nitrogen in macromolecules
Extracellular structural constituents, and intracellular
storage polysaccharides, differ from nucleic acids and
proteins in that elements other than carbon (and nitrogen)
can, in principle, be substituted for the carbon (and
nitrogen)-containing component.
One example is energy storage as polysaccharides or
triglycerides. In ATP units, each (CH2O) unit in a
polysaccharide can yield up to 6 ATP, depending on
assumptions made about stoichiometries of protons in
mitochondrial electron transport and ADP phosphorylation. Energy (but not organic carbon) can also be stored as
polyphosphate, where all but one of the phosphate units in
a polyphosphate needs 1 ATP converted to 1 ADP to add
that unit and can subsequently be used to phosphorylate 1
ADP to form 1 ATP. While such energy storage
economizes on carbon it is very expensive in terms of
phosphate, and polyphosphates usually function as osmotically (relatively) inert stores for phosphate. Even when
they may serve as energy stores (Lambert et al., 2002)
under conditions of ample phosphate supply the maximum
quantity that can be stored in a cell can only energize a
growing cell for minutes rather than the hours or days in
the case of polysaccharides.
For extracellular structural components there is the
possibility of substituting the rigid organic components
(mainly lignin) with inorganic components such as
polymeric silica or crystalline calcium carbonate. Lignin
resists mechanical forces of compression caused by gravity
in above-ground parts of vascular plants which are too
large to be supported by a hydrostatic skeleton, and by
tension of the water column in the walls of conducting
conduits of xylem (Raven, 1993, 1995b). Higher (vascular)
plants with high contents of polymeric silica include
Equisetum, lycopods, members of the Urticales and
Cucurbitales, members of the Eriocaulaceae, some sedges,
and all grasses (Raven, 1983, 2001b; Epstein, 2001). In
some cases (Equisitum, lycopods) the high silica content is
correlated with a low lignin content (Raven, 1983).
However, in Cucurbita fruits the Hard Rind (Hr) gene
controls lignin deposition, and, in parallel, silica deposition
(Piperno et al., 2002). Accordingly, the replacement of
lignin as a structural component of vascular plants with
silica is not a universal aspect of silica deposition in these
plants (Bonilla, 2001; Datnoff et al., 2001). However,
silica in higher plants clearly has an important role in
limiting grazing and parasitism (Epstein, 2001; Kim et al.,
2002).
Some work has been performed on the mechanical
properties of higher plant silica in situ (Speck et al., 1998),
but more is needed. Recent work demonstrated, quantitatively, the mechanical properties of silica in situ in diatoms
(Hamm et al., 2003). The silici®ed cell walls (frustules)
are, as is universal for silici®ed structures, not extensible,
so they cannot show plastic extension during growth, or
elastic changes with variations in external osmolarity, in
the manner of typical plant or algal cells. The increase in
volume of growing cells occurs by a decreasing overlap of
Global aspects of C/N interactions 17
the two halves of the wall (valve). Hamm et al. (2003)
showed that the mechanical properties of silici®ed cell
walls of diatoms were able to account for the inability of
certain crustacean grazers to crush the cells.
By contrast to silica, calcium carbonate seems to have a
much less important mechanical role in algae and plants,
and does not substitute signi®cantly for structural polysaccharides. However, it can have anti-herbivore functions
(Meyer and Paul, 1995).
Carbon and nitrogen in low relative molecular
mass (Mr) organic compounds
While macromolecules account for most of the carbon and
nitrogen in algae and plants (Gnaiger and Bitterlich, 1984;
Handley et al., 1989; LourencËo et al., 1998; Falkowski,
2000; Raven, 2001a; Geider and La Roche, 2002) there are
signi®cant quantities of lower Mr, soluble compounds
which have important roles and can account for quite a
large fraction of total cellular carbon and/or nitrogen, for
example, in compatible solutes in organisms growing in
habitats of very high osmolarity.
The functions of low Mr organic compounds
In addition to low Mr organic compounds which are parts
of energy transformation pathways, or of biosynthetic
pathways leading to macromolecules, the functions of low
Mr compounds can be categorized as follows (Table 2). (1)
Storage of energy/reduced carbon and nitrogen. (2)
Accumulation of organic salts related to pH regulation
(Raven and Smith, 1976; Raven, 1985; Andrews, 1986;
Andrews et al., 1995a, b; Yin and Raven, 1997). (3)
Compatible solutes and osmoregulation (Raven, 1985;
Oren, 1999). (4) UV-B screening and/or visual signalling
to animals (Gronquist et al., 2001). (5) Restriction of
grazing or parasitism, and establishment of mutualisms
(Harborne, 1993). (6) Free radical scavenging (Smirnoff
and Cumbes, 1988; Sunda et al., 2002). (7) The organic C
which is cycled in the carboxylation±decarboxylation
cycle of Crassulacean Acid Metabolism, involving much
more carbon than in the case of C4 photosynthesis (Raven
and Spicer, 1996).
Nitrogen-containing and nitrogen-free low Mr organic
compounds
Some nitrogen-containing solutes ful®l more than one of
these functions, for example, proline, which is a protein
amino acid, can also act as a compatible solute, and
scavenges free radicals (hydroxyl radicals) (Smirnoff and
Cumbes, 1988). Citrulline is not a protein amino acid, but
acts as a nitrogen storage and long-distance transport
compound and as a hydroxyl radical scavenger (Akashi
et al., 2001). Glycinebetaine is a compatible solute with
very little capacity to scavenge hydroxyl radicals
(Smirnoff and Cumbes, 1988). Mycosporine-like amino
acids (MAAs) are important UV-B-screening compounds
in many aquatic organisms, with high trophic levels
deriving their MAAs from the primary producers (e.g.
dino¯agellates, red algae) which synthesize them (Shick
and Dunlap, 2002). The MAAs have little capacity to
scavenge hydroxyl radicals, unlike the parent, nitrogenfree, gadusols (Shick and Dunlap, 2002). The MAAs are
often present at high concentrations in compartments with
a high concentration and diversity of proteins, and so must
be compatible solutes. Nitrogen-containing compounds
also function as relatively grazer-speci®c anti-grazer
agents in higher plants (alkaloids) and as ¯ower, fruit
and seed pigments in the Chenopodiales (Centrospermae)
(as betalains) (Harborne, 1993).
With the obvious exception of nitrogen storage, these
functions can also be performed by nitrogen-free compounds (Table 2). Compatible solutes lacking nitrogen
include sugar alcohols, non-reducing sugars, and the
sulphur-containing dimethylsulphoniopropionate (DMSP).
These compounds also act as effective free radical scavengers and, when stored in vacuoles where they can be
replaced by inorganic solutes, the sugar derivatives can act
as organic carbon and energy stores. DMSP is broken down
using a lyase enzyme (with replacement by de novo
synthesis), resulting in acrylate and dimethylsulphide
(DMS), both of which are very effective hydroxyl radical
scavengers (Sunda et al., 2002). Oxidation products of
DMS, ®rst dimethylsulphoxide (DMSO) and then methanesulphinic acid, are also very effective hydroxyl radical
scavengers (Sunda et al., 2002).
Ascorbate is another sugar derivative that is an effective
hydroxyl radical scavenger, as well as being involved as an
enzyme substrate in peroxidases. Ascorbate is often
present at high enough concentrations in plastid stroma
to qualify as a compatible solute. The sugar derivatives and
DMSP (and derivatives) are not UV-B absorbers, but there
are a range of nitrogen-free compounds which act as UV-B
absorbers. Examples are phlorotannins in the brown algae
(which lack MAAs) (Schoenwalder, 2002), coumarins in
some green marine macroalgae which have very low
contents of MAAs (Gomez et al., 1998; Perez-Rodriguez
et al., 1998, 2001) and various soluble phenylpropanoids
(mainly ¯avonoids) in higher plants.
Chemical categories which contain UV-B-absorbing
nitrogen-free compounds can also function in optical
signalling to animals (anthocyanins in ¯owers, fruits and
seeds) and as grazing deterrents (tannins in higher plants;
phlorotannins in brown algae). Terpenoids are also
involved as feeding deterrents in algae and higher plants
(Paul et al., 1993; Meyer and Paul, 1995).
The two preceding paragraphs show that, with the
exception of storage of nitrogen as low Mr organic nitrogen
compounds, essentially all of the functions considered can
be performed by nitrogen-containing or by nitrogen-free
low Mr organic compounds. The functions are storage,
18 Raven et al.
Table 2. Low molecular mass organic compounds in photosynthetic organisms and their functions, emphasizing the occurrence of
nitrogen-containing and nitrogen-free compounds
Function
Nitrogen-free examples
Nitrogen-containing examples
Reference
Carbon and energy storage
Nitrogen storage
Organic anion salts resulting
from acid-base regulation
Compatible solutes,
osmoregulation
Sucrose, polyols
±
Malate, oxalate
±
Amino acids, amides
±
Polyols, glycerol glucosides
and galactosides, disaccharides,
dimethylsulphonio-propionate
Phlorotannins, ¯avonoids
Citrulline, glycinebetaine,
proline
Raven
Raven
Raven
Raven
Raven
UV-B screening and/or visual
signalling to animals
Restriction of grazing and
parasitism
Scavenging of free radicals
Organic C cycling in
carboxylation/decarboxylation
cycle in CAM and C4
photosynthesis
Betalains, mycosporine-like
amino acids
Tannins, terpenoids
Alkaloids
Ascorbate, linear polyols,
phlorotannins, tannins
Citrulline, glutathione, proline
Malate (CAM and C4); starch,
sugars (CAM only); pyruvate,
phosphoenol-pyruvate (CAM
and C4)
Aspartate (C4 only) alanine
(C4 only)
compatible solute, UV-B screening and optical signalling,
defence against grazers and parasites, and free radical
scavenging (Table 2).
Taxonomic distribution of nitrogen-containing and
nitrogen-free low Mr organic compounds
There are considerable differences among taxa with
respect to the use of nitrogen-free and nitrogen-containing
low Mr organic compounds. Marine red algae use MAAs as
their UV-B screening compounds, but otherwise use
nitrogen-free low Mr organic compounds (Karsten et al.,
1994; Bischoff et al., 2000). However, marine red algae
also use phycobilins in their light-harvesting complexes;
these are more nitrogen-costly than chlorophyll-based
light-harvesting complexes. Furthermore, perhaps a third
of marine macroalgae rely on diffusive CO2 entry rather
than a carbon concentrating mechanism (Raven et al.,
2002b, c). Diffusive CO2 entry could involve a greater
nitrogen investment in Rubisco plus photorespiratory
enzymes than would be involved in a carbon-concentrating
mechanism. The red algae have a very low content of
nitrogen-containing carbohydrates, or of proteins, in their
cell walls. The brown algae are even less involved in
nitrogen-containing low Mr organic solutes, since they use
phlorotannins rather than MAAs as their UV-B screens.
Green algae may have low concentrations of MAAs, and
can use proline as a compatible solute, but also operate a
relatively low-nitrogen economy for low Mr solutes.
Neither brown nor red algae use the nitrogen-costly
phycobilins in light-harvesting, and generally have the
(probably) nitrogen-thrifty carbon concentrating mechanisms rather than diffusive CO2 entry (Raven et al.,
2002b c).
(1984a)
(1984a)
and Smith (1976);
(1985); Yin and Raven (1997)
(1985); Oren (1999)
Harborne (1993); Gronquist et al.
(2001); Schoenwalder (2002);
Shick and Dunlap (2002)
Harborne (1993); Schoenwalder
(2002)
Smirnoff and Cumbes (1988);
Akashi et al. (2001);
Sunda et al. (2002)
Raven and Spicer (1996)
For diatoms the compatible solutes of marine representatives include nitrogenous compounds such as glycinebetaine and homarine, as well as DMSP (Keller et al.,
1999a, b). Other clades of algae generally have some
nitrogenous compounds (e.g. glycinebetaine) and nitrogenfree compatible solutes such as polyols, glucosyl glycerides or galactosyl glycerides, or DMSP (Keller et al.,
1999a, b). Diatoms (even in high osmolarity environments) also lack high concentrations of soluble UV-B
absorbing compounds (Beardall and Raven, 2004;
Tartarotti and Sommaruga, 2002).
Higher plants lack nitrogen-containing UV-B screening
compounds (other than betalains), but some have nitrogencontaining compatible solutes such as proline, glycinebetaine and citrulline.
Quantitative aspects of low Mr organic compounds
Discussion thus far of the occurrence of nitrogen-containing and nitrogen-free low Mr organic solute has been
qualitative, insofar as the concentration of each of the often
multifunctional solutes in the cells is concerned. Before
considering speci®c cases of the quantity of the solutes in
organisms, some generalities can be discussed. The largest
relative and absolute range of concentrations of low Mr
organic solutes relates to the genotypic and phenotypic
variation in total low Mr organic solutes as a function of the
Yp component of external Yw. The water depth component
of Yw, i.e. 1 MPa per 100 m, does not require balancing
intracellular osmolytes.
Raven (1982, 1995a, 2003) shows that the low intracellular osmolarity of effectively wall-less freshwater cells
can be related to the continuous energy expenditure for
active water ef¯ux. `Effectively wall-less' means cells
with exposed plasmalemma, i.e. all ¯agellates regardless
Global aspects of C/N interactions 19
of whether the cell body has a `wall' or not, and amoeboid
cells (Raven, 1982). Raven (1982, 1995a, 2003) points out
that the energy cost of active water ef¯ux is, for a given
hydraulic conductivity of the plasmalemma, directly
proportional to the square of difference in water potential
between the cytosol and the medium. Clearly, minimizing
the energy cost of volume regulation by active water ef¯ux
need not be the prime metabolic priority under all growth
conditions, but low osmolarity (as low as 40 osmol m±3)
does characterize many freshwater effectively wall-less
cells (Raven, 1982). Low intracellular osmolarity means
that no compatible solutes can be accommodated, and
there are signi®cant restrictions on the concentration of
soluble UV-B absorbers, free radical scavengers, and antibiophage compounds. These constraints on the quantity
per unit cell volume of compounds which meet abiotic and
biotic challenges have not thus far been characterized as
decreasing inclusive ®tness relative to walled cells of
higher osmolarity in the same habitat. Furthermore, there
do not seem to be data on the fraction of total cell carbon
and nitrogen in these low-osmolarity cells which is in low
Mr compounds rather than macromolecules. The prediction
for low osmolarity cells is that a smaller fraction of cell
carbon and nitrogen is in low Mr solutes. Inter alia, this
would mean that close predictions of cell protein could be
obtained from cell nitrogen and a factor derived from the
protein:nitrogen mass ratio for phototroph protein (corrected for other nitrogen-containing macromolecules, i.e.
RNA and DNA) (Handley et al., 1989; LourencËo et al.,
1998). By contrast, a less close prediction would occur for
cells with a higher intracellular osmolarity and hence
potentially greater concentrations of low Mr storage
nitrogen compounds, and nitrogen-containing UV-B
screens, free radical scavengers and anti-biophage compounds.
Even when nitrogen in macromolecular nitrogen compounds, and in major categories of low Mr nitrogenous
compounds, is accounted for, there can be a signi®cant
shortfall of nitrogen in measured components of a cell
relative to the total nitrogen in the cell (Keller et al., 1999a,
b; Geider and La Roche, 2002).
At the other extreme are photosynthetic cells in very
high osmolarity environments (e.g. the Dead Sea; the Great
Salt Lake). The only phototrophs able to grow in these
habitats are Dunaliella species; these green ¯agellates are
iso-osmotic with the medium, so at least they do not
maintain a higher internal than external osmolarity.
Nevertheless, these cells have to maintain compatible
solute at a concentration suf®cient to equal, with other
intracellular solutes, the osmolarity outside, involving a
concentration of compatible solute of up to 5 kmol m±3.
The compatible solute in Dunaliella is glycerol, which,
with only three carbon atoms, is the cheapest (in energy
and carbon terms) compatible solute to produce; it is still
not clear how leakage of glycerol to the medium is
maintained at the low level which is observed (Raven,
1984b). Even using this low-carbon (and no nitrogen)
compatible solute, the concentration of carbon in glycerol
in Dunaliella comprises 60% of the total cell carbon. Other
things being equal, the C:N ratio of a freshwater
Chlamydomonas (with no compatible solute) should only
be half that of its close relative Dunaliella growing in very
high osmolarity environments. None of the organisms able
to grow at the highest osmolarities tolerated by photolithotrophs has a nitrogen-containing compatible solute. In
addition to any shortage of nitrogen in hypersaline waters, there is also the question of the higher
energy cost of producing nitrogenous compatible solutes,
none of which have fewer than ®ve carbon atoms and one
nitrogen atom per molecule, especially if nitrate is the
nitrogen source (Raven, 1985).
Extracellular low Mr carbon and nitrogen
compounds
Thus far only the materials retained within the alga or plant
have been considered. Organic materials lost by the alga or
plant must also have been processed through the resource
acquisition and metabolism pathways, and have energetic
and catalytic requirements for their synthesis, in addition
to those of the organic carbon and nitrogen that are
retained by the organism. While both nitrogen-free and
nitrogen-containing organic compounds are lost from
algae and plants, it is not clear if the ratio in which the
elements are lost is less than, equal to, or greater than the
ratio in the organism. Some genotypically and/or phenotypically determined losses of organic compounds from
algae and plants are now considered.
(1) Secretion of organic anions and protons in relation to
acquisition of nutrients by roots of higher plants. Secretion
of malate or citrate with protons facilitates the release of
phosphate from solid mineral forms (apatite; ferric iron
complexes) (Dakora and Phillips, 2002; Lambers et al.,
2002; Roelefs et al., 2001; Ryan et al., 2001; Penaloza
et al., 2002; Sas et al., 2002). The most extreme example
of this is in plants with cluster roots (Skene, 2003), where
up to a quarter of the carbon assimilated in net primary
productivity can be secreted as citrate (Dinkelaker et al.,
1989). Here there is a clear nutritional role in the excretion
of organic anions.
(2) Secretion of malate anion by root tips of A1-tolerant
genotypes of Triticum aestivum has a role in decreasing A1
toxicity by chelating toxic A1 species (Ryan et al., 2001).
(3) While the role is unknown, Mahmood et al. (2002)
showed that sugars were excreted by roots of ammoniumgrown Leptochloa fusca to a much greater extent than by
roots of nitrate-grown plants. This difference could, if
generalizable, help to explain the lower than expected
energetic and growth rate advantages of growth on
ammonium rather than on nitrate (Britto et al., 2001).
20 Raven et al.
(4) Organic matter losses from shoots include gaseous
losses of methanol and of terpenoids (Monson and
Holland, 2001; Sharkey and Yeh, 2001; Rosenstiel et al.,
2003). These losses can be signi®cant fractions of net
primary productivity, with wide genotypic variability.
Older ideas that such losses economize on water loss in
transpiration by diverting energy dissipation as the latent
heat of evaporation of organic materials rather than water
are negated by considerations of the large number
(hundreds) of water molecules lost in transpiration to ®x
the carbon needed to produce even a one-carbon volatile
organic compound, and the lower latent heat of evaporation per molecule of methanol, and of terpenoids, than of
water.
A role for volatile and in¯ammable organic material can
be seen in ®re-tolerant plants in ®re-prone habitats.
Production of an explosive mixture of organic volatiles
and air can promote the spread of ®re initiated by lightning,
destroying ®re-intolerant competitors of the ®re-tolerant,
®re-promoting plants (Gill et al., 1981).
(5) Losses of non-volatile organic solutes from shoots
include organic anions with and without protons, e.g. malic
acid from shoot glands of Cicer (Raven, 1985). Restriction
of losses of inorganic and organic nutrients from shoots
may be a function of the thick, waxy cuticle of
schlerophyllous/xeromorphic plants (Beadle, 1966;
Wright and Westoby, 2003; cf. Lamont et al., 2002).
In addition to these losses of nitrogen-free organic
matter, there are also losses of nitrogen-containing organic
compounds. As well as amino acid losses from roots with
no obvious function, there are also losses of other organic
nitrogen compounds which may act as siderophores in
cyanobacteria and grasses. Losses of volatile organic
nitrogen from land plant shoots include the indoles
involved in attracting carrion-eating pollinating insects to
¯owers and in¯orescences, especially the thermogenic
in¯uorescences of aroids (Kite, 1995; Thien et al., 2002).
Losses of inorganic carbon and nitrogen
compounds
Major contributions to the overall carbon, but not generally
the nitrogen, balance of plants and algae are losses of
inorganic compounds. Respiratory and photorespiratory
(C3 plants) losses of carbon dioxide account for half or
more of gross ®xation of carbon dioxide in photosynthesis.
Net loss of gaseous ammonia from plants to low-ammonia
atmospheres are a small component of the overall nitrogen
¯ux in plant metabolism. Raven et al. (1992a) cite data
suggesting that global gaseous ammonia losses from
vegetation (natural and managed) do not exceed 2.3
Tmol N per year, i.e. only about 1% of the total primary
assimilation of nitrogen by land plants of 209 Tmol N per
year (Table 1). In high-ammonia atmospheres (e.g. near
seabird colonies, or intensive pig or poultry production
units) there can be net uptake of ammonia by plant shoots
(Raven et al., 1992a, 1993; Hill et al., 2001). Global
gaseous losses of nitrogen from vegetation as nitric oxide
are about two orders of magnitude lower than losses as
ammonia (Wildt et al., 1997).
Interactions of nitrogen nutrition with water
relations and carbon dioxide supply
For terrestrial plants, water availability can be a signi®cant
constraint on growth. CO2 ®xation and growth necessarily
involves very signi®cant water loss in transpiration. For
vascular plants the magnitude of water loss per unit dry
matter increases from CAM (50±100 g g±1) to C4 (200±300
g g±1) and to C3 (400±500 g g±1). The extent of water loss
per unit dry matter gain can, in some plants, vary with the
form and concentration of nitrogen sources. Most of the
experimental observations and quantitative modelling has
involved C3 plants. Here there is some mis-match between
theoretical predictions and experimental observations.
Theoretical considerations suggest that the order of
increasing water cost of dry matter gain should be
ammonium < nitrate < dinitrogen (Raven, 1985).
However, measurements of water loss per unit dry matter
gain rarely show lower values for ammonium than for
nitrate as the nitrogen source (Raven et al., 1992b, 1993;
Yin and Raven, 1998). For a number of dicotyledonous
crops the water lost per unit dry matter gain is greater for
plants grown on ammonium than on nitrate; however, at
least part of the greater water cost for growth on
ammonium could be related to slower growth on the
reduced than on the oxidized nitrogen source (Raven et al.,
1992a, b, 1993; Yin and Raven, 1998). For dinitrogen
®xation, fewer data are available; while the available
information suggests that the water cost of dry matter
accumulation is greater for diazotrophically growing
Phaseolus vulgaris than when this plant is growing on
combined nitrogen (Allen et al., 1988). Data on the d13C of
plant organic matter, as an indicator of water loss in
transpiration per unit carbon gain in gross photosynthesis,
suggests that there is also a higher water cost in
diazotrophic plants of Casuarina equisetifolia than in
otherwise similar plants growing on combined nitrogen
(Martinez-Carrasco et al., 1998)
The comparisons of theoretical and observed values
considered all involve laboratory studies, and mainly
concern different forms of nitrogen. Field observations are
also available (Patterson et al., 1997) which indicate the
occurrence, genotypic and phenotypic, of trade-offs
between water costs of growth and nitrogen costs of
growth.
The hydrological cycle has an important role in
determining nitrogen supply to photosynthetic organisms.
Waterlogging of soils leads to nitrate loss by denitri®cation, while through-¯ow removes nitrate. Non-water-
Global aspects of C/N interactions 21
logged soils promote nitri®cation of mineralized organic
matter. The hydrological cycle also removes organic and
inorganic carbon and nitrogen from terrestrial habitats to
inland waters and, ultimately, marine habitats (Berner and
Berner, 1996; Falkowski and Raven, 1997; Raven and
Falkowski, 1999). The atmosphere also provides a conduit,
albeit minor, for combined nitrogen exchange between
terrestrial and aquatic habitats (Raven et al., 1992a, 1993).
In the ocean, the thermohaline circulation is important in
determining the supply of nitrate to surface waters from
deep waters where nitrate is generated from sedimented
organic nitrogen by mineralization and nitri®cation
(Falkowski and Raven, 1997).
An unequivocal aspect of global environmental change
is increased atmospheric CO2 concentration. Particularly
in C3 plants, increasing CO2 concentrations for growth can
increase the plant C/N ratio, in part due to the decreased
expression of Rubisco as increased CO2 and CO2/O2
increase CO2 ®xation per unit Rubisco (Loladze, 2002;
Sterner and Elser, 2002). The reduction in nitrogenous
compounds relative to nitrogen-free compounds in plants,
and in the dilution of other nutrients needed by heterotrophs, has implications for natural food webs as well as
for agriculture and horticulture (Loladze, 2002; Sterner
and Elser, 2002). However, not all plants show the
`expected' acclimation to elevated CO2, regardless of the
nitrogen availability. An example of such a plant is
Helianthus annuus (Zerihun and Bassirad, 2000).
Conclusions
The chemical properties of carbon and nitrogen can, to a
signi®cant extent, be used to rationalize their roles in
organisms, including plants. The ratio of carbon to
nitrogen in photosynthetic organisms varies greatly, from
about ®ve in microalgae, through higher values in
macroalgae and herbaceous vascular plants to very high
values (>100) in woody plants. The possibilities for
economizing on nitrogen use in crop plants of a given
life form are relatively limited. Among macromolecules it
may be possible to manipulate the content of major
proteins (e.g. Rubisco) provided that this does not reduce a
plant's capacity to cope, in terms of yield and quality, with
the full range of conditions encountered in its agricultural
or horticultural environments. More adventurous possibilities include replacement, or supplementation, of native
enzymes with homologues with different kinetic properties
from other organisms. Examples include (possibly) a Form
1D Rubisco from red algae, or other algae which obtained
their plastids by secondary endosymbiosis involving red
algae, and (more certainly) cyanobacterial fructose-1,6-/
sedoheptulose-1,7-bisphosphatase. There is also the possibility of economizing on nitrogen, by replacing low
molecular weight solutes containing nitrogen with compounds lacking nitrogen acting as compatible solutes, UV-
B screens and signalling compounds. This possibility, via
genetic transformation, could be especially applicable to
members of the Chenopodiaceae with betalain pigments
and glycinebetaine as the compatible solute.
Acknowledgements
JAR gratefully acknowledges funding from AFRC, BBSRC, NERC,
SEERAD, and SERC for work involving the study of carbon±
nitrogen interactions in a number of photosynthetic organisms. The
Scottish Crop Research Institute is grant-aided by the Scottish
Environment and Rural Affairs Department. We are grateful to two
anonymous referees whose comments have signi®cantly improved
this paper.
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