Nature’s green revolution: the remarkable
evolutionary rise of C4plants
Colin P. Osborne*and David J. Beerling
Department of Animal and Plant Sciences, University of Sheffield, Sheffield S10 2TN, UK
Plants with the C4photosynthetic pathway dominate today’s tropical savannahs and grasslands, and
account for some 30% of global terrestrial carbon fixation. Their success stems from a physiological
CO2-concentrating pump, which leads to high photosynthetic efficiency in warm climates and low
atmospheric CO2 concentrations. Remarkably, their dominance of tropical environments was
achieved in only the past 10 million years (Myr), less than 3% of the time that terrestrial plants have
existed on Earth. We critically review the proposal that declining atmospheric CO2triggered this
tropical revolution via its effects on the photosynthetic efficiency of leaves. Our synthesis of the
latest geological evidence from South Asia and North America suggests that this emphasis is
misplaced. Instead, we find important roles for regional climate change and fire in South Asia, but
no obvious environmental trigger for C4success in North America. CO2-starvation is implicated in
the origins of C4plants 25–32 Myr ago, raising the possibility that the pathway evolved under more
extreme atmospheric conditions experienced 10 times earlier. However, our geochemical analyses
provide no evidence of the C4mechanism at this time, although possible ancestral components of
the C4pathway are identified in ancient plant lineages. We suggest that future research must redress
the substantial imbalance between experimental investigations and analyses of the geological
Keywords: atmospheric CO2concentration; C4plants; plant evolution; stable carbon isotopes
Photosynthetic CO2-fixation has provided the carbon
for life on Earth for at least the past 2.7 billion years
(Gyr). More than 99% of the history of this ancient
process has been dominated by C3 photosynthesis
(figure 1), so-called because its first products are
carboxylic acids formed of three linked carbon atoms.
But between 25 and 32 million years (Myr) ago, a
revolutionary innovation evolved in tropical grasses in
the form of a solar-powered carbon dioxide pump
based on four-carbon acids (‘C4 photosynthesis’),
which boosts photosynthesis in hot conditions. It
works by pumping CO2 from the mesophyll into a
specialized ring of bundle sheath cells centred around
leaf veins, where an extremely localized version of C3
photosynthesis operates, bathed in high CO2concen-
trations (figure 1; Hatch 1971). Although, this
specialized ‘Kranz’ anatomy is the norm in the vast
majority of C4plants, it is not essential, and certain
desert species have evolved an alternative form of the
pathway in which all elements are packed into single
cells (Voznesenskaya et al. 2001). A variation on the
single-celled carbon concentrating mechanism is called
Crassulacean acid metabolism (CAM) and is more
widely adopted in drought tolerant species. It operates
by temporally separating the activities of a C4-like
pump mechanism and C3photosynthesis.
The C4 photosynthetic pathway is a major evol-
utionary success, accounting for some 20–30% of
terrestrial CO2-fixation on Earth (Lloyd & Farquhar
1994) and 30% of global agricultural grain production
(Steffen et al. 2004). Plants utilizing this pathway
dominate tropical grasslands and savannahs, and rank
among the world’s most important crops, including
sugarcane (Saccharum officinarum), maize (Zea mays)
and sorghum (Sorghum bicolor). And the C4revolution
is not confined to grasses. In one of the most striking
examples of convergent evolution in plants (Conway-
Morris 2003), a C4carbon-concentrating mechanism
has originated in more than 40 independent evolution-
ary groups (Sage 2004). Paradoxically, its multiple
origins belie a complex trait that requires the coordi-
nated expression of at least 20–30 unlinked genes to
operate efficiently (Wyrich et al. 1998; Furumoto et al.
The C4and CAM mechanisms probably arose as a
‘fix’ for an intrinsic inefficiency in Rubisco (ribulose-1,
5-bisphosphate carboxylase/oxygenase), the enzyme
catalysing CO2-fixation in every plant. This ancient
enzyme originated in ‘greenhouse’ conditions when the
Earth’s atmosphere contained CO2at up to 100 times
today’s level (Rye et al. 1995) and negligible amounts of
O2(Bekker et al. 2004). A CO2-fixing enzyme in this
atmosphere had little requirement for high CO2-
affinity, and gained no advantage from distinguishing
between CO2and O2molecules. However, the ensuing
long-term decline in atmospheric CO2subsequently
Phil. Trans. R. Soc. B (2006) 361, 173–194
Published online 28 November 2005
*Author for correspondence (firstname.lastname@example.org).
Received 20 April 2005
Accepted 18 August 2005
q 2005 The Royal Society
exposed an important problem; Rubisco fixes O2as
well as CO2, at a rate depending on the CO2: O2ratio.
In today’s low CO2: O2atmosphere, O2-fixation in C3
plants wastes captured solar energy and causes a net
loss of CO2via the photorespiration pathway.
C4plants overcome this problem using a coupled
enzyme system with a much higher affinity than
Rubisco for CO2(figure 1), first dissolving the gas by
using carbonic anhydrase (CA) to form bicarbonate
carboxylase (PEPc). The four-carbon products of this
fixation diffuse into bundle sheath cells, where CO2is
released by decarboxylase enzymes (DC, figure 1) and
reaches concentrations of 3–8 times those in C3
photosynthetic cells (reviewed by Kanai & Edwards
1999). Rubisco in C4plants, therefore, experiences a
saturatingCO2environment similar to that of C3plants
growing in ancient ‘greenhouse’ atmospheres, and
photorespiration is minimized (Osmond 1971).
Through this mechanism, C4 plants achieve a sub-
stantial photosynthetic advantage over their C3con-
temporaries with falling atmospheric CO2and at high
temperatures (figure 2; Bjo ¨rkman 1971), as Rubisco
becomes increasingly unable to distinguish O2from
CO2. However, the C4mechanism carries a major cost;
its dependence on light to energize the CO2-pump
lowers photosynthetic efficiency relative to the C3type,
especially in conditions when photorespiration is
naturally suppressed, such as high CO2 and cool
temperatures (figure 2).
The use of an energy-dependent system to alleviate
photorespiration in C4plants leads to a trade-off, with
beneficial results for photosynthetic light-use efficiency
at high temperatures and a decline in efficiency at low
temperatures (figure 2). The point at which costs
match benefits is termed the ‘crossover temperature’
and decreases with CO2, as photorespiration becomes
increasingly problematic for C3plants (figure 2). This
simple physiological contrast between photosynthetic
types was first quantified 30 years ago (Ehleringer &
Bjo ¨rkman 1977), and has been invoked subsequently as
the ‘quantum yield hypothesis’ to explain biogeogra-
phical patterns of C4ecological (Ehleringer et al. 1997)
and evolutionary success (Ehleringer et al. 1991;
Cerling et al. 1997). Along geographic temperature
gradients, the crossover point matches the mean
growing season temperature where C4grasslands are
3Þ, and then fixing it using phosphoenolpyruvate
replaced by C3types (Ehleringer et al. 1997), both from
the equator towards cooler climates and on mountains,
where the air cools with altitude (reviewed by Sage et al.
1999). In warmer climates, the C4 type has a
photosynthetic advantage, while the C3type benefits
at cooler temperatures.
C4plants have clearly evolved an effective solution
for the inherent kinetic inefficiency of Rubisco.
However, the mechanisms translating this physiological
advantage into evolutionary and ecological success
remain unclear, and are major unresolved questions in
biology. Here, we review recent advances in our
understanding of C4plant evolution, integrating recent
developments across the geological, ecological, phys-
iological and molecular sciences. We begin with the
origins of the C4photosynthetic pathway, reconciling
geological evidence with molecular data on the
evolutionary history of plant lineages. Next, we
investigate the remarkable picture built-up from
isotopic analyses of fossil remains, demonstrating
rapid global expansion of C4-dominated ecosystems,
and consider its likely cause. Finally, we present a
theoretical analysis of the possible selection pressures
for a C4-type of carbon concentrating mechanism
resulting from the unusual atmospheric composition of
the Permo-Carboniferous 300 Myr ago (Ma). We
investigate the possibility that the necessary environ-
mental and physiological prerequisites were in place for
the C4 mechanism to have evolved 300 Myr earlier
than is currently accepted, through a detailed isotopic
analysis of fossil plant remains and preliminary
physiological measurements of evolutionarily ancient
2. ORIGINS OF C4PLANTS
The evolutionary origins of C4 plants may be
elucidated using fossil evidence of Kranz anatomy
and an important difference in the stable carbon
isotope composition (d13C) of C3 and C4 plants.
Rubisco discriminates strongly against the heavy
isotope of carbon (13C), relative to its more abundant
form (12C). In contrast, the dissolution of CO2to form
bundle sheath cell
Figure 1. Simple schematic diagram of the C4photosynthetic
pathway showing compartmentalization of the different
enzyme systems involved, and the connection between
CO2-pumping by the C4cycle and CO2-fixation by the C3
cycle. Abbreviations: CA, carbonic anhydrase; HCO3,
bicarbonate; PEPc, phosphoenolpyruvate carboxylase; DC,
decarboxylase enzyme(s); Rubisco, ribulose-1,5-bisphos-
leaf temperature (°C)
quantum yield (molmol–1)
C3 550 ppm CO2
C3 380 ppm CO2
C3 190 ppm CO2
Figure 2. Modelled interaction between temperature and
CO2on the photosynthetic quantum yields (maximum light-
use efficiencies) of C3 and C4 plants. Notice that the
temperature at which C3and C4quantum yields cross over
declines with falling atmospheric CO2concentration.
174C. P.Osborne & D. J. Beerling
Nature’s green revolution
Phil. Trans. R. Soc. B (2006)
bicarbonate and its subsequent fixation by the CA–
PEPc enzyme system slightly favours the heavier
isotope (Farquhar 1983). This physiological difference
translates into a marked contrast in the d13C of C3and
C4plant tissues, which persists long after they have
been eaten and incorporated into the bones or teeth of
herbivores, or decomposed into biomarkers within
geological sediments (Cerling 1999). Analysis of d13C
in geological materials, therefore, provides an import-
ant opportunity for reconstructing changes in C4plant
abundance on evolutionary time-scales. However,
interpreting these data requires care because shifts in
the d13C of atmospheric CO2cause parallel changes in
Kranz anatomy can be identified in well-preserved
fossil leaf fragments dating to the Late Miocene,
5–12 Ma. A petrified grass from the Ricardo Formation
(12.5 Ma) of California is currently the earliest
undisputed C4 plant, and additionally characterized
by a typical C4d13C signature (Nambudiri et al. 1978).
A second early example of Kranz anatomy is reported
for a silicified grass from the Ogallala Formation
(5–7 Ma) in Kansas (Thomasson et al. 1986). Several
older C4grass species are claimed from the Fort Ternan
locality in Kenya (14 Ma) on the basis of cuticle
morphology (Dugas & Retallack 1993), but internal
leaf anatomy is not preserved, and their photosynthetic
type remains in question (Cerling 1999). These fossils
highlight a major difficulty in identifying fossil plants on
the basis of anatomical features; in most cases these are
simply not preserved, an issue especially acute for C4
plants, which inhabit seasonally dry environments
where fossilization is unlikely. Direct evidence of
Kranz anatomy is therefore rare, and the origins of C4
photosynthesis must be inferred by integrating data
from stable carbon isotope analyses and investigations
of plant molecular genetics.
Despite intriguing isotopic evidence from the
Cretaceous (90 Ma, Kuypers et al. 1999), general
consensus currently places the earliest origins of C4
photosynthesis within the family Poaceae (grasses)
(Kellogg 2000). Grasses first appear in the fossil record
as pollen in the Palaeocene (55–60 Ma), with
additional, more equivocal records of grass-like pollen
in the latest Cretaceous (70 Ma; Jacobs et al. 1999).
Estimates of when C4 photosynthesis arose in this
group come from molecular genetic techniques, in
which evolutionary histories are retraced by comparing
differences in DNA sequences between species. Since
most DNA mutations are selectively neutral, with no
effect on Darwinian fitness, they accumulate over time
by ‘genetic drift’ and therefore indicate evolutionary
distance between species. By using the number of
mutations between species and a mutation rate
calibrated using fossils, the ‘molecular clock’ technique
dates the appearance of C4photosynthesis in the grass
sub-family Panicoideae at 25–32 Ma (Gaut & Doebley
1997). However, phylogenetic data are unable to
distinguish with confidence whether this event was
followed by multiple reversions back to the C3type, up
to six further C4origination events, or the persistence
of genes allowing flipping between types (Kellogg
2000; Duvall et al. 2001, 2003; Guissani et al. 2001).
additional, and independent, origination events occur-
ring earlier than 25 Ma, within the cluster of related
grass lineages Aristidoideae, Eriachneae and Chlor-
idoideae, although these are not precisely dated at
present (Kellogg 2000, 2001). Each origination co-
opted a slightly different set of biochemical pathways to
achieve a functional C4cycle (Sinha & Kellogg 1996),
and followed an adaptation to open habitats in grass
groups whose ancestors were confined to forest shade
habitats, like today’s bamboos (Kellogg 2001). How-
ever, the location of this evolutionary innovation
remains mysterious, because high diversity and ancient
origins in the grasses make geographic centres of C4
evolution hard to pinpoint (Sage 2004).
C4 photosynthesis also proliferated within the
Cyperaceae (sedges), and numerous families of Eudi-
cots, including the Asteraceae (daisies), Brassicaceae
(cabbages), Euphorbiaceae, but especially the Cheno-
podiaceae and related Amaranthaceae, where 13
(Kadereit et al. 2003). Molecular genetic evidence
points to 37 independent evolutionary origins for C4
photosynthesis outside the grasses (Sage 2004),
starting as early as 14–21 Ma in the Chenopodiaceae
(Kadereit et al. 2003). Biogeographical analysis of
diversity in these groups suggest diverse centres of C4
origin located across the world, in southern Texas-
central Mexico, central Asia, sub-tropical Africa and
sub-tropical South America (Sage 2004).
Molecular genetics, therefore, predicts that the
carbon isotope signature of C4grasses should be picked
up from the Oligocene (23–35 Ma) onwards, with C4
Eudicots contributing soon afterwards (from 14 to
21 Ma). The signal is faint, but present in carbonates
from fossil soils (palaeosols) dating to 23 Ma in the
southern Great Plains of North America (Fox & Koch
2003). After ruling out a number of potential biases,
reconstructions for this region suggest that C4plants
made up 12–34% ofthe biomass from 23 to 7 Ma in the
Late Miocene (Fox & Koch 2003), and grew in open
woodland ecosystems (Stro ¨mberg 2004). Similar
claims from East Africa of a persistent, but relatively
low, abundance of C4plants in a savannah or woodland
ecosystem from 15 to 7 Ma are disputed, on the
grounds of conflicting evidence from mammalian teeth
and possible misidentification of palaeosols (Cerling
1999). What is clear, however, is that prior to 9 Ma
there is no isotopic signature of a C4-dominated
ecosystem anywhere in the world. Data from Africa,
South America, the Indian sub-continent, and China
all show ecosystems comprised entirely of C3plants,
although a low C4 presence is difficult to exclude
against a varying C3d13C background (Cerling et al.
1997). For up to 25 Myr from their inferred origins, C4
plants were rare or absent from the tropics. The C4
revolution was a long-time coming.
3. CO2AND THE RISE OF C4PLANTS TO
C4 plants came to dominate terrestrial ecosystems
abruptly in the Late Miocene (5–8 Ma). A revolu-
tionary expansion of C4plants is identified by major
shifts in d13C (figure 3) across the southern US Great
Nature’s green revolution
C. P.Osborne & D. J. Beerling 175
Phil. Trans. R. Soc. B (2006)
Plains, Argentina, Bolivia, India, Pakistan, Nepal and
Kenya (reviewed by Cerling et al. 1997). Isotopic
changes are recorded in palaeosol carbonate, organic
matter, and in the diets of large mammals and flightless
birds via tooth, bone and egg shell d13C (figure 3;
reviewed by Cerling et al. 1997; Cerling 1999). The
revolution continued into the Pliocene (2–5 Ma), with
later C4expansions in the northern US Great Plains
(Cerling et al. 1997), China (Ding & Yang 2000), Chad
(Zazzo et al. 2000), and across East Africa (Levin et al.
2004). It transformed ecosystems from the tropical to
the warm temperate climate zones across four con-
tinents, with C4biomass increasing from near zero to
more than 80% of vegetation in just 2–4 Myr (figure 3).
The near-synchronous expansion of C4plants across
diverse geographical regions in the Late Miocene
points towards a global trigger for the phenomenon,
and led Ehleringer, Cerling and co-workers (Ehleringer
et al. 1991; Cerling et al. 1997) to propose the net
decline in atmospheric CO2over the last 150 Myr as
the mechanism. According to this proposal, falling CO2
gradually lowered the crossover temperature of C3
plants (figure 2) until it fell below tropical tempera-
tures, making C4photosynthesis progressively more
advantageous and allowing C4 plants to achieve
ecological dominance in ever-cooler climates (Ehler-
inger et al. 1991; Cerling et al. 1997). Circumstantial
evidence for this decline in crossover temperature with
CO2comes from aslight asynchronyin the timing of C4
expansion, with dominance achieved first in hot,
equatorial Kenya, followed by the southern Great
Plains and Pakistan (20–378N), and finally the
northern Great Plains (40–438N; Cerling et al. 1997).
Further indirect support is provided by the inverse
relationship between tropical C4plant abundance from
d13C records during recent ice age (glacial) cycles
(Ehleringer et al. 1997), and fluctuations of atmos-
pheric CO2from 180 p.p.m during glacial intervals to
280 p.p.m during interglacial periods (Petitet al.1999).
The CO2 starvation mechanism rapidly achieved
widespread acceptance, but geological data are now
beginning to challenge its proposed role in C4success
(Keeley & Rundel 2003). Palaeo-CO2reconstructions
from three independent proxies indicate low CO2
concentrations for at least 15 Myr before the Late
Miocene expansion of C4grasslands (figure 4; Pagani
et al. 1999; Pearson & Palmer 2000; Royer et al. 2001).
The inferred levels of CO2 vary between 180 and
320 p.p.m (figure 4), and correspond to crossover
temperatures of less than 10–22 8C (figure 2), well
within tropical temperature limits. According to the
CO2hypothesis, the C4mechanism would, therefore,
have presented a substantial photosynthetic advantage
in the tropics as early as the Oligocene (23 Ma).
Critically, we note that CO2was not declining during
the Late Miocene period of ecological change, and one
proxy even indicates an increase at 8 Ma (figure 4;
Pagani et al. 1999). Further evidence from the last
glacial cycle also challenges the primary role of CO2as
a driver of ancient C4successes, with uncoupling of C4
plant abundance from CO2being attributed to lower
summer rainfall (Huang et al. 2001; Scott 2002).
On the basis of our assessment, we reject the
hypothesis of CO2-starvation as the proximate driver
of Miocene C4expansions. However, we still see C4
physiology as an adaptation to low CO2atmospheres,
because it only provides a photosynthetic advantage at
tropical temperatures of less than 30 8C when CO2
concentrations are lower than 500 p.p.m (figure 2;
Ehleringer et al. 1997). Consequently, although declin-
ing CO2 was not the direct trigger for Miocene
expansions of C4plants, a decrease in its concentration
was a necessary pre-condition for this widespread C4
success (Sage 2001). Three independent palaeo-CO2
proxies all show a drop in CO2from a high point of
greater than 1000 p.p.m in the mid-Cretaceous to its
in CO2from between 1000 and 1500 p.p.m to around
300 p.p.m during the Oligocene (23–35 Ma; Pagani
et al. 2005), coincident with continental ice-sheet
initiation on Antarctica (Zachos et al. 2001). This
decrease corresponds to a fall in crossover temperature
from greater than 40 to 17–21 8C (figure 2), and
d13C (tooth enamel)
05 10 15 20
d13C (tooth enamel)
Figure 3. Increase in d13C from palaeosols and tooth enamel
showing apparent synchronicity in the transition to C4-
dominated terrestrial ecosystems across continents. Data for
(a) from Cerling et al. (1997), (b) from Quade & Cerling
(1995) and (c) from Passey et al. (2002).
176C. P.Osborne & D. J. Beerling
Nature’s green revolution
Phil. Trans. R. Soc. B (2006)
Farquhar, G. D. 1983 On the nature of carbon isotope
discrimination in C4species. Aus. J. Plant Physiol. 10,
Fox, D. L. & Koch, P. L. 2003 Tertiary history of C4biomass
in the Great Plains, USA. Geology 31, 809–812. (doi:10.
Fox, D. L. & Koch, P. L. 2004 Carbon and oxygen isotope
variability in Neogene paleosol carbonates: constraints on
the evolution of the C4-grasslands of the Great Plains,
Furumoto, T., Hata, S. & Izui, K. 2000 Isolation and
characterization of cDNAs for differentially accumulated
transcripts between mesophyll cells and bundle sheath
strands of maize leaves. Plant Cell Physiol. 41, 1200–1209.
Gaut, B. S. & Doebley, J. F. 1997 DNA sequence evidence for
the segmental allotetraploid origin of maize. Proc. Natl
Acad. Sci. USA 94, 6809–6814. (doi:10.1073/pnas.94.13.
Guissani, L. M., Cota-Sa ´nchez, J. H., Zuloaga, F. O. &
Kellogg, E. A. 2001 A molecular phylogeny of the grass
subfamily Panicoideae (Poaceae) shows multiple origins of
C4photosynthesis. Am. J. Bot. 88, 1993–2012.
Hatch, M. D. 1971 Mechanism and function of the C4
pathway of photosynthesis. In Photosynthesis and photo-
respiration (ed. M. D. Hatch, C. B. Osmond & R. O.
Slayter), pp. 139–152. San Diego, CA: Academic Press.
Hattersley, P. W. 1983 The distribution of C3and C4grasses
in Australia in relation to climate. Oecologia 57, 113–128.
Hattersley, P. W. 1992 C4photosynthetic pathway variation
in grasses (Poaceae): its significance for arid and semi-arid
lands. In Desertified grasslands: their biologyand management
(ed. G. P. Chapman), pp. 181–212. London: Academic
Hibberd, J. M. & Quick, W. P. 2002 Characteristics of C4
photosynthesis in stems and petioles of C3 flowering
plants. Nature 415, 451–454. (doi:10.1038/415451a)
Hoorn, C., Ohja, T. & Quade, J. 2000 Palynological evidence
for vegetation development and climatic change in the
Sub-Himalayan Zone (Neogene,Central Nepal). Palaeo-
geogr. Palaeoclimatol. Palaeoecol. 163, 133–161. (doi:10.
Huang, Y., Street-Perrott, F. A., Metcalfe, S. E., Brenner,
M., Moreland, M. & Freeman, K. H. 2001 Climate
change as the dominant control on glacial-interglacial
variations in C3and C4plant abundance. Science 293,
Jacobs, B. F., Kingston, J. D. & Jacobs, L. L. 1999 The origin
of grass-dominated ecosystems. Ann. Missouri Bot. Gard.
Janis, C. M., Damouth, J. & Theodor, J. M. 2000 Miocene
ungulates and terrestrial primary productivity: where have
all the browsers gone? Proc. Natl Acad. Sci. USA 97,
Jones, T. P. 199413C enriched lower Carboniferous fossil
plants from Donegal. Ireland: carbon isotope constraints
on taphonomy, diagenesis and palaeoenvironments. Rev.
Pal. Pal. 81, 53–64.
Kadereit, G., Borsch, T., Weising, K. & Freitag, H. 2003
Phylogeny of Amaranthaceae and Chenopodiaceae and
the evolution of C4photosynthesis. Int. J. Plant Sci. 164,
Kanai, R. & Edwards, G. E. 1999 The biochemistry of C4
photosynthesis. In C4plant biology (ed. R. F. Sage & R. K.
Monson), pp. 49–87. San Diego, CA: Academic Press.
Keeley, J. E. & Rundel, P. W. 2003 Evolution of CAM and C4
carbon-concentrating mechanisms. Int. J. Plant. Sci. 164,
Keeley, J. E. & Rundel, P. W. 2005 Fire and the Miocene
expansion of C4grasslands. Ecol Lett. 8, 683–690. (doi:10.
Kellogg, E. A. 2000 The grasses: a case study in macro-
evolution. Ann. Rev. Ecol. Syst. 31, 217–238. (doi:10.
Kellogg, E. A. 2001 Evolutionary history of the grasses. Plant
Physiol. 125, 1198–1205. (doi:10.1104/pp.125.3.1198)
Knapp, A. K. & Medino, E. 1999 Success of C4photosyn-
thesis in the field: lessons from communities dominated by
C4plants. In C4plant biology (ed. R. F. Sage & R. K.
Monson), pp. 251–283. San Diego, CA: Academic Press.
Kuypers, M.M. M., Pancost, R. D. & Damste ´, J.S. S. 1999 A
large and abrupt fall in atmospheric CO2concentration
during Cretaceous times. Nature 399, 342–345. (doi:10.
Larcher, W. 1994 Photosynthesis as a tool for indicating
temperature stress events. In Ecophysiology of photosynthesis
(ed. E. D. Schulze & M. M. Caldwell), pp. 261–277.
Latorre, C., Quade, J. & McIntosh, W. C. 1997 The
expansion of C4grasses and global change in the late
Miocene: stable isotope evidence from the Americas.
Earth Planet. Sci. Lett. 146, 83–96. (doi:10.1016/S0012-
Levin, N. E., Quade, J., Simpson, S. W., Semaw, S. & Rogers,
M. 2004 Isotopic evidence for Plio-Pleistocene environ-
mental change at Gona, Ethiopia. Earth Planet. Sci. Lett.
219, 93–110. (doi:10.1016/S0012-821X(03)00707-6)
Lloyd, J. & Farquhar, G. D. 199413C discrimination during
CO2assimilation by the terrestrial biosphere. Oecologia 99,
Long, S. P. 1983 C4photosynthesis at low temperatures.
Plant Cell Environ. 6, 345–363.
Long, S. P. 1999 Environmental responses. In C4plant biology
(ed. R. F. Sage & R. K. Monson), pp. 215–249. San
Diego, CA: Academic Press.
MacFadden, B. J., Cerling, T. E., Harris, J. M. & Prado, J.
1999 Ancient latitudinal gradients of C3/C4 grasses
interpreted from stable isotopes of New World Pleistocene
horse (Equus) teeth. Global Ecol. Biogeog. 8, 137–149.
Maier-Reimer, E., Mikolajewicz, U. & Crowley, T. J. 1990
Ocean general circulation model sensitivity experiment
with an open central American isthmus. Paleoceanography
McLoughlin, S. & Drinnan, A. N. 1997 Revised stratigraphy
of the Permian Bainmedart Coal Measures, northern
Prince Charles Mountains, East Antarctica. Geol. Mag.
134, 335–353. (doi:10.1017/S0016756897006870)
Mikolajewicz, U. & Crowley, T. J. 1997 Response of a
coupled ocean/energy balance model to restricted flow
through the central America isthmus. Paleoceanography 12,
Monson, R. K. 1999 The origins of C4 genes and
evolutionary pattern in the C4metabolic phenotype. In
C4 plant biology (ed. R. F. Sage & R. K. Monson),
pp. 377–410. San Diego, CA: Academic Press.
Nambuduri, E. M. V., Tidwell, W. D., Smith, B. N. &
Hebbert, N. P. 1978 A C4plant from the Pilocene. Nature
276, 816–817. (doi:10.1038/276816a0)
Ogle, K. 2003 Implications of interveinal distance for
quantum yield in C4 grasses: a modeling and meta-
analysis. Oecologia 136, 532–542. (doi:10.1007/s00442-
Osmond, C. B. 1971 The absence of photorespiration in C4
plants: real or apparent? In Photosynthesis and photorespira-
tion (ed. M. D. Hatch, C. B. Osmond & R. O. Slayter),
pp. 472–482. San Diego, CA: Academic Press.
192C. P.Osborne & D. J. Beerling
Nature’s green revolution
Phil. Trans. R. Soc. B (2006)
Otto-Bliesner, B. L. 1995 Continental drift, runoff, and
weathering feedbacks: implications from climate model
experiments. J. Geophys. Res. 100, 11537–11548. (doi:10.
Otto-Bliesner, B. L., Brady, E. C. & Shields, C. 2000 Late
Cretaceous ocean: coupled simulations with the National
Centre for Atmospheric Research climate system model.
Owensby, C. E., Ham, J. M., Knapp, A. K. & Auen, L. M.
1999 Biomass production and species composition change
in a tallgrass prairie ecosystem after long-term exposure to
elevated atmospheric CO2. Global Change Biol. 5,
Pagani, M., Freeman, K. H. & Arthur, M. A. 1999 Late
Miocene atmospheric CO2concentrations and the expan-
sion of C4grasses. Science 285, 876–879. (doi:10.1126/
Pagani, M., Zachos, J., Freeman, K. H., Tipple, B. &
Boharty, S. 2005 Marked decline in atmospheric carbon
dioxide concentrations during the Paleogene. Science 309,
Passey, B. H., Cerling, T., Perkins, M., Voorhies, M., Harris,
J. & Tucker, S. 2002 Environmental change in the Great
Plains: an isotopic record from fossil horses. J. Geol. 110,
Pearson, P. N. & Palmer, M. R. 2000 Atmospheric carbon
dioxide concentrations over the past 60 million years.
Nature 406, 695–699. (doi:10.1038/35021000)
Pearson, P. N., Ditchfield, P. W., Singano, J., Harcourt-
Brown, K. G., Nicholas, C. J., Olsson, R. K., Shackleton,
N. J. & Hall, M. A. 2001 Warm tropical sea surface
temperatures in the Late Cretaceous and Eocene epochs.
Nature 413, 481–487. (doi:10.1038/35097000)
Petit, J. R. et al. 1999 Climate and atmospheric history of the
past 420,000 years from the Vostok ice core, Antarctica.
Nature 399, 429–436. (doi:10.1038/20859)
Quade, J. & Cerling, T. E. 1995 Stable isotopes in paleosols
and the expansion of C4grasses in the late Miocene of
Northern Pakistan. Palaeogeogr. Palaeoclimatol. Palaeoecol.
115, 91–116. (doi:10.1016/0031-0182(94)00108-K)
Quade, J., Cater, J.M. L., Ojha, T. P., Adam, J. & Harrison,
T. M. 1995 Late Miocene environmental change in Nepal
and the northern Indian subcontinent: stable isotope
evidence from paleosols. GSA Bulletin 107, 1381–1397.
Raven, J. A. 2002 Evolutionary options. Nature 415,
Royer, D. L., Wing, S. L., Beerling, D. J., Jolley, D. W., Koch,
P. L., Hickey, L. J. & Berner, R. A. 2001 Paleobotanical
evidence for near present-day levels of atmospheric CO2
during part of the Tertiary. Science 292, 2310–2313.
Royer, D. L., Berner, R. A., Montan ˜ez, I. P., Tabor, N. J. &
Beerling, D. J. 2004 CO2 as a primary driver of
Phanerozoic climate. GSA Today 14, 4–10. (doi:10.1130/
Rye, R., Kuo, P. H. & Holland, H. D. 1995 Atmospheric
carbon dioxide concentrations before 2.2 billion years ago.
Nature 378, 603–605. (doi:10.1038/378603a0)
Sage, R. F. 2001 Environmental and evolutionary precondi-
tions for the origin and diversification of the C4
Sage, R. F. 2004 The evolution of C4photosynthesis. New
Phytol. 161, 341–370. (doi:10.1111/j.1469-8137.2004.
Sage, R. F. & Pearcy, R. W. 2000 The physiological ecology of
C4 photosynthesis. In Photosynthesis: physiology and
metabolism (ed. R. C. Leegood, T. D. Sharkey &
S. von Caemmerer), pp. 497–532. The Netherlands:
Sage, R. F., Wedin, D. A. & Li, M. 1999 The biogeography of
C4photosynthesis: patterns and controlling factors. In C4
plant biology (ed. R. F. Sage & R. K. Monson),
pp. 313–373. San Diego, CA: Academic Press.
Scotese, C. R. & McKerrow, W. S. 1990 Revised world maps
and introduction. In Palaeozoic palaeogeography and
biogeography (ed. W. S. McKerrow & C. R. Scotese)
Geological Society Memoir No. 12, pp. 1–21. London:
Geological Society of London.
Scott, L. 2002 Grassland development under glacial and
interglacial conditions in southern Africa: review of pollen,
phytolith and isotope evidence. Palaeogeogr. Palaeoclimatol.
Palaeoecol. 177, 47–57. (doi:10.1016/S0031-0182(01)
Sinha, N. R. & Kellogg, E. A. 1996 Parallelism and diversity
in multiple origins of C4photosynthesis in grasses. Am.
J. Bot. 83, 1458–1470.
Spicer, R. A., Harris, N.B. W., Widdowson, M., Herman, A.,
Guo, S., Valdes, P. J., Wolfe, J. A. & Kelly, S. P. 2003
Constant elevation of southern Tibet over the past 15
Steffen, W. et al. 2004 Global change and the Earth System. A
planet under pressure. Berlin: Springer.
Stro ¨mberg, C.A. E. 2004 Using phytolith assemblages to
reconstruct the origin and spread of grass-dominated
habitats in the great plains of North America during the
late Eocene to early Miocene. Palaeogeogr. Palaeoclimatol.
Taub, D. R. 2000 Climate and the US distribution of C4grass
subfamilies and decarboxylation variants of C4photosyn-
thesis. Am. J. Bot. 87, 1211–1215.
Thomasson, J. R., Nelson, M. E. & Zakrzewski, R. J. 1986 A
fossil grass (Gramineae: Chloridoideae) from the Miocene
with Kranz anatomy. Science 233, 876–878.
Valdes, P. J. & Crowley, T. J. 1998 A climate model
intercomparison for the Carboniferous. Palaeoclimates:
Data Modelling 2, 219–238.
Veizer, J. et al. 199987Sr/86Sr, d13C and d18O evolution of
Phanerozoic seawater. Chem. Geol. 161, 59–88. (doi:10.
Veizer, J., Godderis, Y. & Franc ¸ois, L. M. 2000 Evidence for
the decoupling of atmospheric CO2and global climate
during the Phanerozoic eon. Nature 408, 698–701.
Vellinga, M. & Wood, R. A. 2002 Global climatic impacts of
a collapse of the Atlantic thermohaline circulation.
Climatic Change 54, 251–267. (doi:10.1023/A:101616
Voznesenskaya, E. V., Franceschi, V. R., Kiirats, O., Freitag,
H. & Edwards, G. E. 2001 Kranz anatomy is not essential
for terrestrial C4 plant photosynthesis. Nature 414,
Watson, A. J., Lovelock, J. E. & Margulis, L. 1978
Methanogenesis, fires and the regulation of atmospheric
oxygen. Biosystems 10, 293–298. (doi:10.1016/0303-
Wildman, R. A., Hickey, L. J., Dickinson, M. B., Berner,
R. A., Robinson, J. M., Dietrich, M., Essenhigh, R. H. &
Wildman, C. B. 2004 Burning of forest materials under
late Palaeozoic high O2 levels. Geology 32, 457–460.
Wright, V. P. & Vanstone, S. D. 1991 Assessing the carbon
dioxide content of ancient atmospheres using palaeocal-
cretes: theoretical and empirical constraints. J. Geol. Soc.
Nature’s green revolution
C. P.Osborne & D. J. Beerling 193
Phil. Trans. R. Soc. B (2006)
Wyrich, R., Dressen, U., Brockmann, S., Streubel, M.,
Chang, C., Qiang, D., Paterson, A. H. & Westhoff, P. 1998
The molecular basis of C4photosynthesis in sorghum:
isolation, characterization and RFLP mapping of meso-
phyll- and bundle-sheath-specific cDNAs obtained by
differential screening. Plant Mol. Biol. 37, 319–335.
Zachos, J., Pagani, M., Sloan, L., Thomas, E. & Billups, K.
2001 Trends, rhythms, and aberrations in global climate
65 Ma to present. Science 292, 686–693. (doi:10.1126/
Zazzo,A., Bocherens, H., Brunet, M., Beauvilian, A., Billiou,
D., Taisso Mackaye, H., Vignaud, P. & Mariotti, A. 2000
Herbivore paleodiet and paleoenvironmental changes in
Chad during the Pliocene using stable carbon isotope
Zhisheng, A., Kutzbach, J. E., Prell, W. L. & Porter,
S. C. 2001 Evolution of Asian monsoons and phased
uplift of the Himalaya–Tibetan plateau since Late
194C. P.Osborne & D. J. Beerling
Nature’s green revolution
Phil. Trans. R. Soc. B (2006)