4.. Conclusions and future challenges
Clearly, our knowledge of thermosensory systems of plants has greatly
expanded in the past decade. Important discoveries in ambient
temperature signaling include the identification of photo/thermal
sensors, an RNA switch and self-coalescence of ELF3 through its
prion-like domain. These three systems unambiguously translate high
ambient temperatures into altered gene expression. The reprogramming of
development by these factors assists plants to avoid damaging high
temperatures. In Arabidopsis, the inhibition of phyB activity by warm
temperatures has been shown in detail. It is however still unknown if
other photoreceptors function in a similar way. Phototropin plays a role
in low temperature signalling in Physcomitrella but it is not known
whether this temperature-dependent activity also stretches to warm
temperatures. Other photoreceptors undergo thermal reversion and so
could conceptually also function in warm temperature sensing, but this
remains to be demonstrated. The finding that PIF7 RNA translation
is enhanced at warm temperatures opens the possibility that other RNAs
act in a similar way. Indeed, the authors of the PIF7 study also show
that HSF2 RNA may also be regulated through a comparable
mechanism (Chung et al., 2020). The finding that ELF3 contains a
prion-like domain that undergoes temperature-dependent coalescence
likely has effects beyond just evening complex transcriptional
repression. ELF3 acts as a scaffold protein for large protein complexes
(Huang et al., 2016) and directly binds to PIF4 to inhibit its
transcriptional activity (Nieto et al., 2015). Both of these functions
are likely inhibited at warm temperatures.
Upon moderate temperature increases, plants trigger a heat stress
response for acclimation, but the sensing mechanism is still largely
unknown. Rather than unfolded proteins, the increased membrane fluidity
at high temperatures is speculated to be a molecular basis of sensing.
While no fluidity sensor has been found in plants, evidence is
accumulating for thermosensory mechanisms based on heat-induced phase
changes in lipids and proteins.
Under heat stress, thylakoid membranes locally undergo transition to
non-bilayer, HII phases, which are essential for heat acclimation, since
they compartmentalize and activate the enzymes of the xanthophyll cycle.
At the plasma membrane, microdomains are formed containing lipids in a
liquid-ordered phase. These domains harbor potential signaling lipids
and proteins, including RbohD, which is activated in response to heat
stress. Formation of microdomains and HII phases could constitute a
basis for thermosensitive regulation of enzymes. Simultaneously, they
provide potential avenues for the rapid trafficking of lipids between
phases, in order to preserve membrane integrity under heat stress. For
the latter function, a membrane fluidity sensor would thus not be
required.
The regulation of ELF3 has unveiled a novel type of ambient temperature
sensing mechanism, based on liquid-liquid phase separation (Jung et al.,
2020). Under heat stress, the formation of spherical, condensed liquid
phases within a bulk dilute phase can be triggered by the coalescence of
proteins through their intrinsically disordered, prion-like domains. The
resulting liquid droplets, also called membraneless organelles,
constitute compartments that can contain proteins with associated
regulatory functions. The reversible process of droplet formation is
highly temperature sensitive. Could liquid-liquid phase separation also
function, at higher temperatures, in the activation of the heat stress
response? Such a function was recently proposed for the yeast
RNA-binding protein, Pab1 (poly(A)-binding protein), which displayed
self-coalescence and phase separation upon a shift to a temperature that
induces the heat-shock response (Riback et al., 2017). The extreme
thermosensitivity of this process was quantified using the temperature
coefficient Q 10, the ratio of biological properties measured 10°C
apart. With a Q 10 of 350, it exceeds by far any other known
biological thermosensory process. This indicates the potential of
liquid-liquid phase separation of proteins as a thermosensing mechanism.
The sharp threshold temperature above which phase separation is
triggered, which is determined by the amino acid side chains in the
prion-like domain, allows for precise temperature-dependent regulation
of responses. Pab1 was speculated to activate the heat stress response
by sequestering a negative regulator of HSF in liquid droplets, calling
into question the requirement of unfolded proteins for activation
(Riback et al., 2017). It seems plausible that similar regulation
governs heat stress responses in plants.
The stress-induced clustering of proteins into membrane microdomains
could trigger liquid-liquid phase separation in the adjacent cytosol.
This could result in coupled lipid and liquid compartments, that
assemble selected response components, allowing for specific channeling
of sensory signals to downstream responses (Jaillais & Ott, 2020).
Similarly, plasma membrane organization could respond to changes in the
cell wall, which may also adopt different biophysical states dependent
on temperature (Wu et al., 2018). As yet, such potential interactions
are unexplored territory.
Identifying plant proteins that could act as thermosensors through
liquid-liquid phase separation will be challenging. Previously, heat
stress was found to induce relocalization of splicing factors with
disordered domains, e.g. serine/arginine-rich protein SR45, into
enlarged nuclear speckles (Ali et al., 2003; Reddy et al., 2012), which
could underlie alternative splicing of pre-mRNAs. Many of the,
approximately, 500 proteins in plants with predicted prion-like domains
are transcription factors with potential roles in temperature
signalling. There are prion-like domains in HSFA1b, and several PIFs,
auxin response factors and ABRE-binding factors (Chakrabortee et al.,
2016). Investigating the effect of temperature on the coalescence of
these factors in vitro could yield interesting results.
Thermosensing appears to be a highly distributed capacity, based on a
range of mechanisms which are only just beginning to come to light. Most
strikingly, the temperature-dependent behavior of phyB, the PIF7 RNA
hairpin, and both lipid and liquid phase separations, provides an
impressive spectrum of potential heat sensing and responding modes,
essential for plants to acclimate and survive.