Ultrastructural and Extracellular Protein Changes in Cell Suspension Cultures of Populus euphratica Associated with Low Temperature-induced Cold Acclimation

Populus euphratica Olive is the only tree species that can grow in the saline land and also survive cold winters in northwest China, and it plays a very important role in stabilizing the vulnerable ecosystem there. A cell suspension culture was initiated from callus derived from plantlets of Populus euphratica. Cold acclimation was induced (LT50 of 17.5 ℃) in cell suspension at 45 ℃ in the dark for 30 days and the freezing tolerance increased from LT50 of 12.5 ℃ in nonacclimated cells to LT50 of 17.5 ℃ in cold-acclimated cells. Microvacuolation, cytoplasmic augmentation and accumulation of starch granules were observed in cells that were cold-acclimated by exposure to low temperatures. Several qualitative and quantitative changes in proteins were noted during cold acclimation. Antibodies to carrot extracellular (apoplastic) 36 kD antifreeze protein did not cross react on immunoelectroblots with extracellular proteins in cell suspension culture medium of Populus euphratica, indicating no common epitopes in the carrot 36 kD antifreeze protein and P. euphratica extracellular proteins. The relationship of these changes to cold acclimation in Populus euphratica cell cultures was discussed.

Cold acclimation increases Asian citrus psyllid Diaphorina citri (Hemiptera: Liviidae) survival during exposure to freezing temperatures

The Asian citrus psyllid,Diaphorina citri,is the vector of the pathogenic bacteria that causes Huanglongbing,the most devasting disease affecting citrus worldwide.As cultivation of citrus tends to expand northward,the tolerance of psyllids to freezing temperatures needs to be investigated.While mortality of D.citri to cold temperature has been previously studied,much less is known regarding the acclimation potential of psyllids to cold temperatures.We first evaluated cold resistance of D.citri depending on color morph and sex.Subsequently,we compared mortality of D.citri at−4°C for 10 hours between unacclimated psyllids,psyllids that were gradually acclimated to cold over 1 or 2 weeks,and psyllids intermittently acclimated also for 1 or 2 weeks.We did not find major differences in cold resistance between color morphs and sex.We found that after 1 week of gradual acclimation,the survival of psyllids at−4°C increased to 86%,compared to 20%in the control group.Survival did not increase after the second week of gradual acclimation.Although intermittent acclimation improved survival compared to the control group,it was less effective than gradual acclimation with a survival at 30%and 70%after 1 and 2 weeks of acclimation,respectively,although this difference with gradual acclimation was not significant at week 2.These data show that gradual cold acclimation allows D.citri to survive brief periods below freezing.It may serve as a mechanism responsible for increasing northern establishment of this pest.

Effect of Putrescine on Low-Temperature Acclimation in Chlamydomonas reinhardtii

Putrescine is reported to be necessary for cold acclimation under low-temperature stress.In this study,the effect of low-temperature on some physiological and biochemical parameters has been investigated using the green algae Chlamydomonas reinhardtii.The lipid peroxidation rate,amount of Rubisco protein,activities of antioxidant enzymes and gene expression of polyamine biosynthesis(odc2,and spd1),heat shock proteins(hsp70c,hsp90a,and hsp90c),and PSII repair mechanisms(psba,rep27,and tba1)were determined to understand the low-temperature response.Exogenous putrescine application significantly increased Rubisco protein concentration and catalase enzyme activities under low-temperature stress.Moreover,real-time RT-PCR results and gene expression analysis showed that polyamine metabolism induced gene expression at low-temperatures in the first 24 h.In the same way,the gene expression of heat shock proteins(hsp70c,hsp90a,and hsp90c)decreased under low-temperature treatment for 72 h;however,application of putrescine enhanced the gene expression in the first 24 h.The results obtained indicated that molecular response in the first 24 h could be important for cold acclimation.The psba and tba1 expressions were reduced under low-temperatures depending on the exposure time.In contrast,the exogenous putrescine enhanced the expression level of the psba response to low-temperature at 24 and 72 h.The results obtained in this study indicate that putrescine could play a role in the PS II repair mechanisms under low-temperature stress.

The role of temperature as a driver of metabolic flexibility in the Red-billed Leiothrix(Leiothrix lutea)

Background:The thermoregulatory ability of animals is strongly influenced by the temperature of their environment.Acclimation to cold requires a range of physiological and morphological adjustments.In this study,we tested the hypothesis that a small passerine,the Red-billed Leiothrix(Leiothrix lutea),can maintain homeothermy in cold conditions by adjusting the physiology and biochemistry of its tissue and organs and return to its former physiological and biochemical state when moved to a warm temperature.Methods:Phenotypic variation in thermogenic activity of the Red-billed Leiothrixs(Leiothrix lutea)was investigated under warm(35℃),normal(25℃)or cold(15℃)ambient temperature conditions.Oxygen consumption was measured using an open-circuit respirometry system.Mitochondrial state-4 respiration and cytochrome-c oxidase(COX)activity in liver,kidney heart and pectoral muscle were measured with a Clark electrode.Results:Birds acclimated to an ambient temperature of 15℃ for 4 weeks significantly increased their basal metabolic rate(BMR)compared to a control group kept at 25℃.Birds acclimated to 35℃ decreased their BMR,gross energy intake(GEI)and digestible energy intake(DEI).Furthermore,birds acclimated to 15℃ increased state-4 respiration in their pectoral muscles and cytochrome-c oxidase(COX)activity in their liver and pectoral muscle,compared to the 25℃ control group.Birds acclimated to 35℃ also displayed lower state-4 respiration and COX activity in the liver,heart and pectoral muscles,compared to those kept at 25℃.There was a positive correlation between BMR and state-4 respiration,and between BMR and COX activity,in all of the above organs except the liver and heart.Conclusions:Our study illustrates that the morphological,physiological,and enzymatic changes are associated with temperature acclimation in the Red-billed Leiothrix,and supports the notion that the primary means by which small birds meet the energetic challenges of cold conditions is through metabolic adjustments.

Theplant cytosolic m^(6)A RNA methylome stabilizes photosynthesis in the cold

The sessile lifestyle of plants requires an immediate response to environmental stressors that affect photosynthesis,growth,and crop yield.Here,we showed that three abiotic perturbations—heat,cold,and high light—triggered considerable changes in the expression signatures of 42 epitranscriptomic factors(writers,erasers,and readers)with putative chloroplast-associated functions that formed clusters of commonly expressed genes in Arabidopsis.The expression changes under all conditions were reversible upon deacclimation,identifying epitranscriptomic players as modulators in acclimation processes.Chloroplast dysfunctions,particularly those induced by the oxidative stress-inducing norflurazon in a largely GENOME UNCOUPLED-independent manner,triggered retrograde signals to remodel chloroplastassociated epitranscriptomic expression patterns.N6-methyladenosine(m^(6)A)is known as the most prevalent RNA modification and impacts numerous developmental and physiological functions in living organisms.During cold treatment,expression of components of the primary nuclear m^(6)A methyltransferase complex was upregulated,accompanied by a significant increase in cellular m^(6)A mRNA marks.In the cold,the presence of FIP37,a core component of the writer complex,played an important role in positive regulation of thylakoid structure,photosynthetic functions,and accumulation of photosystemⅠ,the Cytb6f complex,cyclic electron transport proteins,and Curvature Thylakoid1 but not that of photosystemⅡcomponents and the chloroplast ATP synthase.Downregulation of FIP37 affected abundance,polysomal loading,and translation of cytosolic transcripts related to photosynthesis in the cold,suggesting m^(6)Adependent translational regulation of chloroplast functions.In summary,we identified multifaceted roles of the cellular m^(6)A RNA methylome in coping with cold;these were predominantly associated with chloroplasts and served to stabilize photosynthesis.

Climate chamber investigation of the effect of indoor thermal histories on thermal adaptation in different seasons

The indoor thermal history of residents in the hot summer and cold winter climate zone in China have undergone a significant change in recent years,which also changes their seasonal thermal adaptations and this has formed feed-back effects to the increasing usage of air conditioning units in this area.To study the seasonal variations of thermal adaptation,the thermal comfort experiments were conducted on two groups of participants.Each groups included 20 participants who had indoor history mainly with natural ventilation(NV group)and air-conditioning(AC group),respectively.The results demonstrated that the thermal sensation vote(TSV)in warm environments did not differ between AC and NV subjects in summer.However,the TSV of AC subjects were much lower than that of NV subjects in the same standard effective temperature and skin temperature in a cold environment in winter.Overall,the participants who spent most of their time in AC space in winter showed a low level of thermal adaptation with a narrower acceptable skin temperature range of 32.6-33.5℃.Thus,this study presents the basic information regarding the seasonal effects on human thermal adaptation due to different long-term indoor thermal histories.

The direct targets of CBFs:In cold stress response and beyond

Cold acclimation in Arabidopsis thaliana triggers a significant transcriptional reprogramming altering the expression patterns of thousands of cold-responsive(COR) genes. Essential to this process is the C-repeat binding factor(CBF)-dependent pathway, involving the activity of AP2/ERF(APETALA2/ethylene-responsive factor)-type CBF transcription factors required for plant cold acclimation. In this study, we performed chromatin immunoprecipitation assays followed by deep sequencing(ChIP-seq) to determine the genomewide binding sites of the CBF transcription factors. Cold-induced CBF proteins specifically bind to the conserved C-repeat(CRT)/dehydrationresponsive elements(CRT/DRE;G/ACCGAC) of their target genes. A Gene Ontology enrichment analysis showed that 1,012 genes are targeted by all three CBFs. Combined with a transcriptional analysis of the cbf1,2,3 triple mutant, we define 146 CBF regulons as direct CBF targets. In addition, the CBF-target genes are significantly enriched in functions associated with hormone, light,and circadian rhythm signaling, suggesting that the CBFs act as key integrators of endogenous and external environmental cues. Our findings not only define the genome-wide binding patterns of the CBFs during the early cold response, but also provide insights into the role of the CBFs in regulating multiple biological processes of plants.

Orthogroup and phylotranscriptomic analyses identify transcription factors involved in the plant cold response:Acase study of Arabidopsis BBX29

C-repeat binding factors(CBFs)are well-known transcription factors(TFs)that regulate plant cold acclimation.RNA sequencing(RNA-seq)data from diverse plant species provide opportunities to identify other TFs involved in the cold response.However,this task is challenging because gene gain and loss has led to an intertwined community of co-orthologs and in-paralogs between and within species.Using orthogroup(closely related homologs)analysis,we identified 10,549 orthogroups in five representative eudicots.A phylotranscriptomic analysis of cold-treated seedlings from eudicots identified 35 high-confidence conserved cold-responsive transcription factor orthogroups(CoCoFos).These 35 CoCoFos included the well-known cold-responsive regulators CBFs,HSFC1,ZAT6/10,and CZF1 among others.We used Arabidopsis BBX29 for experimental validation.Expression and genetic analyses showed that cold-induction of BBX29 is CBF-and abscisic acid-independent,and BBX29 is a negative regulator of cold tolerance.Integrative RNA-seq and Cleavage Under Targets and Tagmentation followed by sequencing analyses revealed that BBX29 represses a set of cold-induced TFs(ZAT12,PRR9,RVE1,MYB96,etc.).Altogether,our analysis yielded a library of eudicot CoCoFos and demonstrated that BBX29 is a negative regulator of cold tolerance in Arabidopsis.

Innovations and stepwise evolution of CBFs/DREB1s and their regulatory networks in angiosperms

The C-repeat binding factors/dehydrationresponsive element binding protein 1 s(CBFs/DREB1 s)have been identified as major regulators of cold acclimation in many angiosperm plants.However,their origin and evolutionary process associated to cold responsiveness are still lacking.By integrating multi-omics data of genomes,transcriptomes,and CBFs/DREB1 s genome-wide binding profiles,we unveil the origin and evolution of CBFs/DREB1 s and their regulatory network.Gene collinearity and phylogeny analyses show that CBF/DREB1 is an innovation evolved from tandem duplication-derived DREBⅢgene.A subsequent event of e-whole genome duplication led to two CBF/DREB1 archetypes(CladesⅠandⅡ)in ancient angiosperms.In contrast to cold-insensitivity of Clade I and their parent DREBⅢgenes,CladeⅡevolved a further innovation in cold-sensitive response and was stepwise expanded in eudicots and monocots by independent duplications.In geological time,the duplication events were mainly enriched around the Cretaceous-Paleogene(K-Pg)boundary and/or in the Late Cenozoic Ice Age,when the global average temperature significantly decreased.Consequently,the duplicated CBF/DREB1 genes contributed to the rewiring of CBFs/DREB1 s-regulatory network for cold tolerance.Altogether,our results highlight an origin and convergent evolution of CBFs/DREB1 s and their regulatory network probably for angiosperms adaptation to global cooling.

Transcriptional memory and response to adverse temperatures in plants

Temperature is one of the major environmental signals controlling plant development,geographical distribution,and seasonal behavior.Plants perceive adverse temperatures,such as high,low,and freezing temperatures,as stressful signals that can cause physiological defects and even death.As sessile organisms,plants have evolved sophisticated mechanisms to adapt to recurring stressful environments through changing gene expression or transcriptional reprogramming.Transcriptional memory refers to the ability of primed plants to remember previously experienced stress and acquire enhanced tolerance to similar or different stresses.Epigenetic modifications mediate transcriptional memory and play a key role in adapting to adverse temperatures.Understanding the mechanisms of the formation,maintenance,and resetting of stress-induced transcriptional memory will not only enable us to understand why there is a trade-off between plant defense and growth,but also provide a theoretical basis for generating stress-tolerant crops optimized for future climate change.In this review,we summarize recent advances in dissecting the mechanisms of plant transcriptional memory in response to adverse temperatures,based mainly on studies of the model plant Arabidopsis thaliana.We also discuss remaining questions that are important for further understanding the mechanisms of transcriptional memory during the adverse temperature response.