Abstract
A species’ response to thermal stress is an essential physiological
trait that can determine occurrence and temporal succession in nature,
including response to climate change. Environmental temperature affects
zooplankton performance by altering life-spans and population growth
rates, but the molecular mechanisms underlying these alterations are
largely unknown. To compare temperature-related demography, we performed
cross-temperature life-table experiments in closely related
heat-tolerant and heat-sensitive Brachionus rotifer species that
occur in sympatry. Within these same populations, we examined the
genetic basis of physiological variation by comparing gene expression
across increasing temperatures. We found significant cross-species and
cross-temperature differences in heat response, with the heat-sensitive
species adopting a strategy of high survival and low population growth,
while the heat-tolerant followed an opposite strategy. Comparative
transcriptomic analyses revealed both shared and opposing responses to
heat. Most notably, expression of heat shock proteins (hsps ) is
strikingly different in the two species. In both species, hspresponses mirrored differences in population growth rates, showing thathsp genes are likely a key component of a species’ adaptation to
different temperatures. Temperature induction caused opposing patterns
of expression in further functional categories such as energy,
carbohydrate and lipid metabolism, and in genes related to ribosomal
proteins. In the heat-sensitive species, elevated temperatures caused
up-regulation of genes related to induction of meiotic division as well
as genes responsible for post-translational histone modifications. This
work demonstrates the sweeping reorganizations of biological functions
that accompany temperature adaptation in these two species and reveals
potential molecular mechanisms that might be activated for adaptation to
global warming.
Keywords: Brachionus calyciflorus , Brachionus
fernandoi , thermal adaptation, RNA-seq, heat-stress
INTRODUCTION
On a global scale, a species’ occurrence is related to its tolerance of
a particular range of environmental parameters such as temperature,
salinity and precipitation. In aquatic ecosystems, temperature has a
profound impact on an organism’s survival and performance, and can
affect species abundance, spatio-temporal distribution, and habitat
colonization (Paaijmans et al., 2013; Parmesan, 2006). There is great
variation in the thermal tolerance among aquatic taxa. Many species can
tolerate a broad range of temperature, while others have specific and
narrow temperature limits (Cullum,
2008; Hershey, Lamberti, Chaloner & Northington, 2010). Importantly,
this can impact temporal occurrence, and temperature-dependent seasonal
succession has been well documented among genetically similar species
that might have evolved species-specific temperature specializations
(Papakostas, Michaloudi, Triantafyllidis, Kappas, & Abatzopoulos, 2013;
Wen, Xi, Zhang, Xue, & Xiang, 2016; Xiang et al., 2011a; Zhang et al.,
2017). Therefore, understanding species thermal boundaries is essential
for comprehending how species have adapted to their environment and how
they may respond to climate change.
Zooplankton are important components of aquatic ecosystems, as they
transfer organic compounds and energy from primary producers (e.g.
phytoplankton) to higher trophic levels (Segers, 2008). Among
zooplankton, monogonont rotifers are of particular interest because of
their high, often cryptic, diversity, their frequent adaptation to
specific environmental conditions, and their high dispersal capability
(Fontaneto, Kaya, Herniou & Barraclough, 2009; Mills et al., 2017).
Species complexes formerly assumed to be ubiquitous generalists have
been found to comprise cryptic species adapted to specific ecological
conditions regarding temperature, habitat type, or salinity (Gabaldón,
Fontaneto, Carmona, Montero-Pau, & Serra, 2017). As their dispersal
capabilities can be large, distribution and diversification seems less
dependent on geographical barriers and historical factors, suggesting
that ecological specialization is more likely to drive speciation
(Gómez, Serra, Carvalho & Lunt, 2002; Mills et al., 2017; Serra &
Fontaneto, 2017; Suatoni, Vicario, Rice, Snell & Caccone, 2006). As
evidence of specialization, co-occurrence of differentially adapted,
closely related species in a single locality is a common phenomenon in
rotifers (Papakostas, Michaloudi, Triantafilidis, Kappas &
Abatzopoulos, 2013; Xiang, Xi, Wen, & Ge, 2017; Zhang et al., 2017). In
these cases, morphologically similar species might have evolved
different ecological specialties to reduce competition over resources in
space or time (Fontaneto, Giordani, Melone & Serra, 2007; Montero-Pau,
Ramos-Rodríguez, Ciros-Pérez, Serra & Gómez, 2011; Serra & Fontaneto,
2017).
The best studied freshwater monogonont rotifer is the Brachionus
calyciflorus species complex that has recently been resolved to four
different species: Brachionus calyciflorus sensu stricto (s.s.),Brachionus fernandoi , Brachionus dorcas, andBrachionus elevatus , using integrative taxonomy (Michaloudi et
al., 2018; Papakostas et al., 2016). The species of this complex exhibit
temporal succession, and their occurrences have been related to
temperature in several studies (Li, Niu, & Ma, 2010; Wen et al., 2016;
Zhang et al., 2017). More specifically, temperature constraints were
shown to affect the temporal occurrence and abundance of B.
calyciflorus cryptic species in different habitats in China, withB. fernandoi occurring in winter and spring, and B.
calyciflorus s.s. in summer (Xiang et al., 2017; Zhang et al., 2017).
Comparative laboratory studies on heat tolerance between B.
calyciflorus s.s. and B. fernandoi have shown higher
heat-tolerance of the former, thus confirming that temperature tolerance
likely plays a role in their temporal distribution (Paraskevopoulou,
Tiedemann & Weithoff, 2018).
Regulation of gene expression is an
essential mechanism underlying phenotypic plasticity (Wray et al.,
2003). As selection acts on expression, transcriptome data are
particularly useful in revealing genes contributing to phenotypic
plasticity. Differences in gene expression might have been evolved via
divergent selection as a consequence of ecological differentiation
(Romero, Ruvinsky, & Gilad, 2012). Aquatic taxa use a variety of
physiological mechanism to cope with temperature changes. Transcription
studies on heat shock response have often focused on genes encoding
for
heat shock proteins (hsps ). Heat shock proteins are divided into
several groups (families) of different molecular weights (kDa): e.g.hsp90, hspP70, hsp60, hsp40 , and small proteins. Hsp70s ,
in combination with other proteins, play a vital role in stress
tolerance and survival under adverse conditions. They reduce
accumulation of peptide aggregates and promote the correct folding of
newly synthesized proteins (Mayer & Bukau, 2005). Hsp90s play
also a major role in stress tolerance, mainly by removing incorrectly
folded proteins. Furthermore, they regulate the activity of other
proteins (e.g., kinases) and stabilize the cytoskeleton (Csermely,
Schnaider, Soti, Prohászka, & Nardai, 1998). Induction of hspgenes is an evolutionary old and conserved mechanism, and is described
from prokaryotes to higher eukaryotes
(Feder
& Hofmann, 1999). However, the specific genes involved and the
conditions of induction vary among taxa (Parsell & Lindquist, 1993). In
rotifers, particularly Brachionus species, members of thehsp70 and hsp40 families increase heat shock survival,
suggesting that there may be coordination among heat shock proteins in
which hsp40 works synergistically to regulate hsp70 ’s
activity (as shown in B. manjavacas ; Smith, Burns, Shearer, &
Snell, 2012).
To investigate the marked variation in thermal tolerance between two
closely related species in the former B. calyciflorus species
complex (Paraskevopoulou et al., 2018), we compared life-history
demography and gene expression under mild to high temperature
conditions. We use life-table experiments to examine survival,
fecundity, and population growth rate differences between the two
species. We collect transcriptomic data (RNA-seq) to examine the genetic
basis of physiological response and its difference between the two
species. Identifying the mechanisms involved in thermal tolerance and
investigating their variation among heat-tolerant and heat-sensitive
species is important to understand how physiology determines species’
temporal distribution, and how this might be affected by different
scenarios of climate change.
MATERIALS AND METHODS