Do plants contain typical GPCRs?” How is G-protein signaling operating in plants. G-proteins are universal signal transducers mediating many cellular responses. In animal systems the G-protein signaling cycle is activated by seven transmembrane-spanning G-protein coupled receptors (or GPCRs, popularly known as “serpentine receptors”). Whether typical G protein-coupled receptors (GPCRs) exist in plants or not is a fundamental question. In contrast to the animal system, the existence of these types of receptors in plants still remains controversial. While in animals ligand binding causes a change in receptor conformation that activate a particular G Protein, in plants, such mechanism is unknown. In fact, it is considered that the plants G-Proteins are self-activating. The G Proteins have their respective GPCRs in animal system. A lot of information is already accumulated in animal system and hence the animal GPCRs are considered “canonical.” Thus, from the very beginning, plant G-proteins have been compared with the animal counterparts and studied as an extrapolation of the animal model. This presentation provides an insight into the molecular mechanisms of G Protein activation in plants as well as whether “canonical” GPCRs are present in any plant species or not.

1. UNIVERSITY OF AGRICULTURALSCIENCES,BANGALORE
Presentation on
DEPARTMENTOF Cropphysiology
CPH-602- Signal Perceptions and transduction and
regulation of physiological process
ANANYA
1ST PhD
PAMB0077
“Do plants contain typical GPCRs?” How is G-
protein signaling operating in plants?

2. Nobel prize in Chemistry (2012) for studies on
GPCR
ROBERT J. LEFKOWITZ BRIAN K . KOBILKA

3. • Guanosine nucleotide binding protein
(G-protein)
• Serpentine receptor
• 7 transmembrane protein
What are GPCRs?

4. Structure of G protein

5. 7 transmembrane helices connected by
alternating cytosolic and extra cellular loop
C terminal: inside the cell
N terminal : extra cellular region
Extra cellular portion has unique messenger
binding site
Cytosolic loop allow receptor to interact with G
protein
• Ligand binding causes a change in receptor
conformation that activate a particular G Protein
• Eg: Olfactory receptors Norepinephrine receptors

6. G protein contain 3different subunits
• G Alpha
• G Beta
• G Gamma
G alpha – largest , binds to GTP or GDP and G
beta & G gamma are permanently bound together

17. Activity of G Protein persists as long as
• GTP is bound to G alpha subunit
• G alpha and g beta –g gamma complex remain
separated
This feature allows the signal transduction pathway
to shutdown when the messenger is utilized
• RGS –Regulators of G Protein signaling Protein

19. Do plants contain true/typical GPCRs?

20. Evolutionary Support for the Lack
of Plant GPCRs
• GPCRs appear to be strictly limited to the
Eukaryota
• yeast (Blumer et al, 1988), coral (Anctil et al,
2007) nematodes (Carre-Pierrat et al, 2006) ,
arthropods (Stafflinger et al, 2005),
human(Nathans et al, 1984) and even from the
preserved DNA of the woolly mammoth
(Römpler et al, 2006)

21. • Over a decade, research on plant G proteins
has revealed a basic difference between plant
and animal G protein activation and led to the
conclusion that the animal half for G
activation is probably limited to one small
corner of the eukaryotic kingdom
Animal G-protein activation versus Plants….

22. Does plant have true/typical GPCRs?

23. (a) The animal model. An animal G protein forms an inactive heterotrimer in the steady state. Ligand-bound G
protein–coupled receptors (GPCRs) promote nucleotide exchange on the Gα subunit, and GTP-bound Gα separates
from the Gβγ dimer. Both the GTP-bound Gα and the freed Gβγ regulate the activity of the effectors. Gα hydrolyzes
GTP, returns to the GDP-bound state, and then re-forms the inactive heterotrimer with Gβγ. Regulator of G protein
signaling (RGS) proteins accelerate GTP hydrolysis by Gα. The numbers (min−1) beside the black arrows show the
intrinsic rates of GDP/GTP exchange and GTP hydrolysis. (b) The Arabidopsis model. The Arabidopsis Gα protein,
AtGPA1, spontaneously exchanges its GDP for GTP without GPCRs but does not readily hydrolyze GTP without
GTPase-accelerating proteins (GAPs). A seven-transmembrane (7TM) RGS protein, AtRGS1, constitutively
promotes the intrinsically slow hydrolysis reaction by AtGPA1. (c) A structural basis for the self-activating property
of AtGPA1.
10.1146/annurev-arplant-050213-040133

24. The Ras domain (red) has
similarity to small GTPases. It
contains sites for binding to
guanine nucleotides, effectors,
and RGS proteins. The helical
domain (yellow) shields the
guanine nucleotide (blue) bound
on the Ras domain. Ligand-bound
GPCRs in animals or spontaneous
fluctuations
in Arabidopsis change the
orientation of the helical domain,
leaving the guanine nucleotide
exposed, which leads to
dissociation from the Ras domain.
(Urano et al, 2016)
10.1146/annurev-arplant-050213-040133

25. Models of potential regulators of G proteins. The active G protein is shown as a “G” with a
bound GTP. The inactive G protein is bound by GDP. (a) In animals, activation of G proteins is
regulated by a guanine nucleotide exchange factor (GEF) that speeds up the release of bound
GDP. (b) In plants other than cereals, a seven-transmembrane (7TM) regulator of G protein
signaling (RGS) protein speeds up the rate-limiting reaction of hydrolysis. Plants may also
utilize a GDP dissociation inhibitor (GDI), which slows nucleotide exchange. (c) Cereals lack
canonical RGS proteins; therefore, if the rate-limiting GTP hydrolysis is regulated, it is by an
unknown mechanism and protein. (d) In liverworts, both nucleotide exchange and hydrolysis
are fast. The mechanism for regulating the active state of G proteins is unknown and without
precedent. 10.1146/annurev-arplant-050213-040133

26. • In contrast to animal G proteins, plant G proteins
are self-activating,
• regulation of G activation in plants occurs at the
deactivation step (by RGS)
• The self-activating property also means that plant G
proteins “do not need”? (#may need in future)
and therefore do not have typical animal G
protein–coupled receptors
• The precedents of signal transduction activated
plant G proteins, also known as effectors, are unlike
effectors in animal cells (Urano et al, 2016)

27. • Warpeha et al., 2007,reviewed that for the past
decade there has been only one putative GPCR
(GCR1) identified and experimentally investigated in
Arabidopsis GPCR
• GCR2, has been reported in Arabidopsis (Liu et al,
2007) , although the protein sequence does not
appear to have the canonical or typical seven
transmembrane (TM) topology of known GPCRs
and some disputes exist regarding its plant hormone
signaling function (Gao et al, 2007)
• Arabidopsis also has a single regulator of G-protein
signaling (RGS) protein (RGS1), which directly
accelerates the intrinsic guanosine triphosphatase
activity of Gα (Jones et al, 2004)

28. • Whole genome sequencing efforts have shown that
heterotrimeric G-protein signaling can be highly complex
• The human proteome is known to contain 23 Gα, 5 Gβ,
and 12 Gγ subunits (McCudden et al,2005), leading to
over 1,300 known heterotrimeric complexes
• Over 850 human GPCRs are predicted (Bai et al, 2004),
many of which are known to homo- and heterodimerize
(Fredriksson et al, 2005) the number of potential
signaling pathways or functions becomes huge
• While,the number of known heterotrimeric signaling
complex components in plants is potentially less. The
fully sequenced model plant Arabidopsis thaliana has
only one canonical Gα subunit (GPA1), one Gβ subunit
(AGB1), and two identified Gγ subunits (AGG1 and
AGG2) (Assmann et al, 2005)

29. • Candidate plant GPCRs has so far been limited
to the discovery of Arabidopsis GCR1(Hooley
et al, 1998) and its homolog in pea (Misra et
al., 2007), Arabidopsis RGS1 (Temple et al.,
2007)and, Arabidopsis GCR2 (Liu et al, 2007)

30. Is it reasonable that GCR1,
and potentially RGS1, are the
only candidate GPCRs in
Arabidopsis, or are there
other as yet undiscovered
candidate GPCRs?

31. • LANCL1, another lanthionine synthetase, was
initially identified as a GPCR (Prohaska et al, 2000)
prior to biochemical experimentation, which
confirmed its subcellular localization as a peripheral
membrane protein
• Additional controversies have also arisen regarding
the description of GCR2 as a GPCR that functions as
a receptor for the plant hormone abscisic acid.
• Gao et al., 2007, report that GCR2 is not genetically
or physiologically coupled to GPA1 and is not
required for abscisic acid perception during seed
germination and seedling development

32. • They used a combinatorial approach to identify novel GPCRs
based on the direct prediction of GPCRs by the QFC algorithm and
GPCRHMM; signal peptide detection by Phobius; transmembrane
domain prediction by TMHMM2, HMMTOP2, and Phobius; and
subsequent GPCR classification by GPCRsIdentifier and coupling
specificity prediction by Pred-Couple 2

33. • Within the Arabidopsis genome no other genes have
any appreciable similarity to GCR1 or RGS1 by
BLAST analysis
• GCR2 and its two homologs within the Arabidopsis
genome are homologous to the lanthionine
synthetase C family (Gao et al, 2007)
• Results - whole proteome analysis using multiple
topology prediction approach did not predict a
single seven TM domain within this protein

34. Plant G proteins are self-activating
(Conclusion)
• In contrast to animal G proteins, plant G proteins are
self-activating, and therefore regulation of G activation
in plants occurs by deactivation ( Urano and Jones,
2013)
• The self-activating property also means that plant G
proteins “do not need”? (#may need in future) and
therefore do not have typical animal G protein–
coupled receptors

35. PLANTS DO NOT HAVE CANONICAL GPCRs
• In vitro, animal G proteins bind GDP, and removal of this
nucleotide to allow GTP to bind requires a receptor
having GEF activity. Plant G proteins on it’s own
release GDP and bind GTP in vitro, and thus are self-
activating. Self-activation removes the requirement for
a receptor GEF. Plants do not need and therefore do
not have GPCRs. This idea is difficult for many to grasp
because plants have 7TM proteins. There are
approximately 50 proteins in Arabidopsis and rice that
potentially have the same structure as human GPCRs
(Moriyama et al, 2006), but topology and sequence alone
does not define a GPCR. These GPCR may look like
animal GPCRs but are not plant GPCRs, and we
should not call them plant GPCRs

36. SUMMARY
• G protein–coupled signaling in plants is profoundly different
than in it’s animal counterpart, but both plant and animal cells
contain the same G protein core elements
• Plant G proteins are self-activating; specifically, they bind
GTP without the need for a G protein–coupled receptor (GPCR)
• Plants do not have canonical/typical GPCRs
• In most plants, regulation of the activation state is by the GTP
hydrolysis (RGS Protein)
• A new protein architecture comprising a seven-transmembrane
(7TM) domain and a regulator of G protein signaling (RGS)
domain was first identified in plants ( Example - Arabidopsis
RGS1, serves as the regulatory point of G activation)

37. • The precedents of signal transduction activated plant G
proteins, also known as effectors which are well
targetted, in animal cells do not exist in plants. Plant
effectors are under investigation for now!
• The primary function of G signaling in plants is
nutrient sensing, and this information impacts signaling
by several plant hormones, light, pathogen-associated
molecular patterns, and probably other signals (these
also may be the activators of G signaling in
plants…..WE NEVER KNOW!)