Distinct cold tolerance traits independently vary across genotypes in Drosophila melanogaster

Cold tolerance, the ability to cope with low temperature stress, is a critical adaptation in thermally variable environments. An individual's cold tolerance comprises several traits including minimum temperatures for growth and activity, ability to survive severe cold, and ability to resume normal function after cold subsides. Across species, these traits are correlated, suggesting they were shaped by shared evolutionary processes or possibly share physiological mechanisms. However, the extent to which cold tolerance traits and their associated mechanisms covary within populations has not been assessed. We measured five cold tolerance traits—critical thermal minimum, chill coma recovery, short- and long-term cold tolerance, and cold-induced changes in locomotor behavior—along with cold-induced expression of two genes with possible roles in cold tolerance (heat shock protein 70 and frost)—across 12 lines of Drosophila melanogaster derived from a single population. We observed significant genetic variation in all traits, but few were correlated across genotypes, and these correlations were sex-specific. Further, cold-induced gene expression varied by genotype, but there was no evidence supporting our hypothesis that cold-hardy lines would have either higher baseline expression or induction of stress genes. These results suggest cold tolerance traits possess unique mechanisms and have the capacity to evolve independently.     

Interactive effects of heat shock and smoke on germination of nine species forming soil seed banks within the Sydney region

Abstract The germination response of seeds from fire‐prone vegetation to fire‐related cues such as heat shock and smoke has usually been studied by applying the cues singly. The few studies that have applied the cues in combination have shown that interactions between the cues are possible. Here, the response of seeds from a number of species to combined heat shock and smoke is reported. Heat shock (25, 50, 75 and 100°C) and aerosol smoke (0, 5, 10 and 20 min) were applied factorially to nine species that form soil seed banks in the Sydney region of south‐eastern Australia. These species were from Epacridaceae (four species), Myrtaceae (four species) and Cyperaceae (one species) and ranged from fire‐sensitive obligate seeders to fire‐tolerant facultative resprouters. Germination of Dracophyllum secundum R. Br and Sprengelia monticola (A. Cunn. ex DC.) Druce was low and did not respond to the germination cues. The positive response of Gahnia sieberiana Kunth and Kunzea ambigua (Sm.) Druce to heat shock and smoke was independent and additive. The positive response of Kunzea capitata Rchb. to the interaction between heat shock and smoke was synergistic, and the response of Baeckea diosmifolia Rudge and Baeckea imbricata (Gaertn.) Druce was unitive, with germination increase only occurring following combined heat and smoke application. Epacris coriacea A. Cunn. ex DC. and Epacris obtusifolia Sm. had low levels of dormancy and hence it was not possible to find a fire response. Gahnia sieberiana and K. capitata responded differently to the combination of heat shock and smoke than has previously been reported. Germination of species from habitats that are infrequently burnt was not affected by heat shock or smoke. Low‐intensity fire or patches within fire may be important for seedling recruitment as the 50°C heat shock stimulated germination in four of the five species that responded to the heat cue, and germination of Baeckea imbricata declined within the 100°C heat shock treatment. Germination of one species, Baeckea imbricata, was only stimulated by a specific combination of cues, indicating that regeneration niches may be narrow for some species and that the application of a range of heat and smoke doses is required to find such responses. Of the species positively responding to heat shock and smoke, a requirement for both cues was prevalent, therefore the response to these cues in isolation cannot be relied upon to give a true indication of the fire response of a species.     

Physiological adaptation along environmental gradients and replicated hybrid zone structure in swordtails (Teleostei: Xiphophorus)

Local adaptation is often invoked to explain hybrid zone structure, but empirical evidence of this is generally rare. Hybrid zones between two poeciliid fishes, Xiphophorus birchmanni and X. malinche, occur in multiple tributaries with independent replication of upstream‐to‐downstream gradients in morphology and allele frequencies. Ecological niche modelling revealed that temperature is a central predictive factor in the spatial distribution of pure parental species and their hybrids and explains spatial and temporal variation in the frequency of neutral genetic markers in hybrid populations. Among populations of parentals and hybrids, both thermal tolerance and heat‐shock protein expression vary strongly, indicating that spatial and temporal structure is likely driven by adaptation to local thermal environments. Therefore, hybrid zone structure is strongly influenced by interspecific differences in physiological mechanisms for coping with the thermal environment.     

Neighbour signals perceived by phytochrome B increase thermotolerance in Arabidopsis

Due to the preeminence of reductionist approaches, understanding of plant responses to combined stresses is limited. We speculated that light‐quality signals of neighbouring vegetation might increase susceptibility to heat shocks because shade reduces tissue temperature and hence the likeness of heat shocks. In contrast, plants of Arabidopsis thaliana grown under low‐red/far‐red ratios typical of shade were less damaged by heat stress than plants grown under simulated sunlight. Neighbour signals reduce the activity of phytochrome B (phyB), increasing the abundance of PHYTOCHROME‐INTERACTING FACTORS (PIFs). The phyB mutant showed high tolerance to heat stress even under simulated sunlight, and a pif multiple mutant showed low tolerance under simulated shade. phyB and red/far‐red ratio had no effects on seedlings acclimated with nonstressful warm temperatures before the heat shock. The phyB mutant showed reduced expression of several fatty acid desaturase (FAD) genes and less proportion of fully unsaturated fatty acids and electrolyte leakage of membranes exposed to heat shocks. Red‐light‐activated phyB also reduced thermotolerance of dark‐grown seedlings but not via changes in FADs expression and membrane stability. We propose that the reduced photosynthetic capacity linked to thermotolerant membranes would be less costly under shade, where the light input limits photosynthesis.     

Genotype-specific heat shock proteins in two maize inbreds

Summary Leaf blade tissue of maize inbred lines B73 and Mo17 was analyzed for intraspecific genetic variability in the heat shock response. The maize inbreds were characterized for acquired thermal tolerance and patterns of heat shock protein synthesis. The leakage conductivity assay of membrane stability during stress indicated that Mol7 possesses greater potential than B73 to acquire thermal tolerance. Poly(A)⁺ RNA, extracted from leaf blades, was translated in vitro in the presence of ³⁵S-methionine and the translation products separated by twodimensional gel electrophoresis. Major genotypic differences were observed in the translation products. Mo 17 synthesized twelve unique heat shock proteins in the 15–18 kD range, but B73 synthesized only three unique heat shock proteins in the same range. DNA polymorphisms were observed between the maize lines using ³²P labeled heat shock protein gene probes.Abbreviations HKT Heat-killing time - HS Heat shock - HSP Heat shock protein - HMW High molecular weight - LMW Low molecular weight Contribution of the College of Agricultural Sciences, Texas Tech University, Journal No. T-4-333     

The actin gene in Paracoccidioides brasiliensis: organization, expression and phylogenetic analyses

PbrACT1, the gene responsible for the synthesis of actin in Paracoccidioides brasiliensis, was found as a single copy, organized into six exons and five introns. Its open reading frame (ORF) codes for a putative protein of 375 amino acids, with a molecular mass of 41.5 kDa and an isoelectric point of 5.6. Analysis of the nucleotide sequence revealed a high homology to other fungal actins, the presence of characteristic fungal actin sequences, and heat shock elements at the 5′ untranslated region (UTR). Phylogenetic analyses with deduced amino acid sequences of fungal actins grouped P. brasiliensis within the phylum Ascomycota, order Onygenales, in concordance with a few previous reports. Patterns of expression through the temperature-induced morphological transitions from mycelial to yeast-like shapes and reverse, suggests that PbrACT1 is regulated in this process. The PbrACT1 gene sequence is available at the GenBank database under accession number AY383732.     

草菇热激蛋白60基因(Vvhsp60)生物信息学分析及低温下的表达

【背景】生物受到温度胁迫时,热激蛋白被诱导并在短时间内大量产生,可以使受损的蛋白质恢复正常构象,增强生物对逆境胁迫的耐受性。【目的】初步探究草菇热激蛋白60(Vvhsp60)与低温耐受性的关系,为深入开展草菇不耐低温特性的遗传改良奠定理论基础。【方法】对Vvhsp60进行生物信息学分析,以低温敏感型草菇菌株V23及耐低温菌株VH3为实验材料,利用实时荧光定量PCR技术分析低温胁迫及热激诱导后在低温下草菇菌丝体中Vvhsp60基因的表达水平。【结果】草菇Vvhsp60编码蛋白不存在信号肽,不属于分泌蛋白,在线粒体和细胞质内发挥生物学作用,属于双向跨膜蛋白。低温处理显著提高了V23与VH3菌丝体中Vvhsp60基因的表达量,而且VH3中的表达量显著高于V23,推测Vvhsp60基因的表达量高可能有助于增强草菇对低温胁迫的耐受性。经热激处理后两菌株Vvhsp60基因的表达量显著高于各自未热激处理的对照组,表明热激处理可诱导Vvhsp60基因的表达。【结论】Vvhsp60与草菇低温耐受性相关,并且热激可以诱导Vvhsp60基因的表达。     

Effect of calcium ions on the Hsp104 synthesis and heat tolerance of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Effect of calcium ions on heat tolerance of Saccharomyces cerevisiae and on the induction of Hsp104 synthesis by this microorganism was studied. Short-term (30 min) treatment with CaCl₂ at 30°C enhanced the heat tolerance to the lethal heat shock (50°C); the synthesis of Hsp104 was induced as well. The effect of Ca²⁺ on the heat tolerance and Hsp104 synthesis was shown to be ion-specific and was inhibited by LaCl₃, which is known to block calcium ion channels on the cytoplasmic membrane. The effect of Ca²⁺ depended on the potential of the inner mitochondrial membrane. When the cells were treated with sodium azide, which reduced the electrochemical potential, the effect of calcium both on heat tolerance and Hsp104 synthesis was suppressed. Depending on the concentration of exogenous Ca²⁺ and the ambient conditions, calcium ions may either induce or inhibit the expression of the stress genes and cell viability.     

Cloning and expression of heat shock protein genes in two whitefly species in response to thermal stress

Two whitefly species, Trialeurodes vaporariorum and Bemisia tabaci biotype B were shown to have different temperature tolerance and seasonal dynamics. To determine whether this variation in thermal tolerance is related to different expression patterns of heat shock protein (hsp) genes during temperature stress, we obtained complete cDNA sequences for hsp90, hsp70 and hsp20, and analysed their expression profiles across temperature gradients by real‐time quantitative polymerase chain reaction (PCR). Six full‐length cDNAs were cloned and sequenced from these two species. The full‐length cDNAs of hsp90s contain 2166 and 2157 bp open‐reading frames (ORF) which encode proteins with calculated molecular weights of 83 013 and 82 857 Da in T. vaporariorum and B. tabaci, respectively. The 1947 and 1959 bp ORFs of whitefly hsp70s comprise 649 and 653 amino acids with the calculated masses of 70 885 and 71 008 Da in T. vaporariorum and B. tabaci, respectively. Both complete cDNAs of hsp20 of T. vaporariorum and B. tabaci contain 585 bp ORFs and deduced amino acid sequences had molecular weights of 21 559 and 21 539 Da, respectively. The hsp expression profile results showed that temperatures for onset (T_on) or maximal (T_max) induction of hsp expression in T. vaporariorum were generally 2–6°C lower than those in B. tabaci. These results suggest that the T_on (or T_max) of hsps can represent the differences in temperature tolerance of these two whitefly species, and may be used to determine their natural geographical distribution and natural population seasonal dynamics. Significant upregulation of most hsps were observed when temperature stress was lifted, except that hsp70 and hsp20 of B. tabaci did not respond to the cold stress, indicating that response to heat and cold stress may have a different genetic and physiological basis in two whitefly species. These results highlight the importance of understanding the complexity of the heat shock response across multiple isoforms while attempting to link them to whole‐organism traits such as thermal tolerance.     

Analysis of the Heat-Shock Response Displayed by Two Chaetomium Species Originating from Different Thermal Environments

Three features of the heat shock response, reorganization of protein expression, intracellular accumulation of trehalose, and alteration in unsaturation degree of fatty acids were investigated in the thermophilic fungus Chaetomium thermophile and compared to the response displayed by a closely related mesophilic species, C. brasiliense. Thermophilic heat shock response paralleled the mesophilic response in many respects like (i) the temperature difference observed between normothermia and the upper limit of translational activity, (ii) the transient nature of the heat shock response at the level of protein expression including both the induction of heat shock proteins (HSPs) as well as the repression of housekeeping proteins, (iii) the presence of representatives of high-molecular-weight HSPs families, (iv) intracellular accumulation of trehalose, and finally (v) modifications in fatty acid composition. On the other hand, a great variability between the two organisms was observed for the proteins expressed during stress, in particular a protein of the HSP60 family that was only observed in C. thermophile. This peptide was also present constitutively at normal temperature and may thus fulfil thermophilic functions. It is shown that accumulation of trehalose does not play a part in thermophily but is only a stress response. C. thermophile contains less polyunsaturated fatty acids at normal temperature than C. brasiliense, a fact that can be directly related to thermophily. When subjected to heat stress, both organisms tended to accumulate shorter and less unsaturated fatty acids.     

Thermal tolerance,evaporative water loss,air-water oxygen consumption and zonation of intertidal prosobranchs: a new synthesis

Duration of emergence increases with tidal height on rocky shores therefore, emergence adaptations in intertidal species such as littorine and other prosobranch gastropods have been considered correlated with zonation patterns; temperature tolerance, desiccation resistance and aerial respiration rate all commonly assumed to increase progressively with increasing zonation level. Such direct correlations are rarely observed in nature. Maximal aerial gas exchange occurs in mid-shore, not high shore species. Temperature tolerance and desiccation resistance do not increase directly with shore height. Thus, hypotheses regarding physiological correlates of zonation require revaluation. A new hypothesis is presented that the high tide mark presents a single major physiological barrier on rocky shores. Above it, snails experience prolonged emergence and extensive desiccation; below it, predictable submergence and rehydration with each tidal cycle. Thus, desiccation stress is minimal below the high tide mark and maximal above it. Therefore, species restricted below high tide (the eulittoral zone) should display markedly different adaptive strategies to emergence than those above it (the eulittoral fringe). A review of the literature indicated that adaptations in eulittoral species are dominated by those allowing maintenance of activity and foraging in air including: evaporative cooling; low thermal tolerance; elevated aerial O₂ uptake rates; and high capacity for radiant heat absorption. Such adaptations exacerbate evaporative water loss. In contrast, species restricted to the eulittoral fringe display adaptive strategies that minimize desiccation and prolong survival of emergence including: foot withdrawal, preventing heat conduction from the substratum; aestivation in air; elevated thermal tolerance reducing necessity for evaporative cooling; position maintenance by cementation to the substratum and increased capacity for heat dissipation. In order to test of this hypothesis the upper thermal limits, tissue and substratum temperatures on emergence in direct sunlight and evaporative water loss and tissue temperatures on emergence in 40 °C were evaluated for specimens of six species of eulittoral and eulittoral fringe gastropods from a granite shore on Princess Royal Harbour near Albany, Western Australia. The results were consistant with adaptation to the proposed desiccation barrier at high tide. The eulittoral species, Austrocochlea constricta, Austrocochlea concamerata, Nerita atramentosa and Lepsiella vinosa, displayed adaptations dominated by maintenance of activity and foraging during emergence while the eulittoral fringe littorine species, Bembicium vittatum and Nodilittorina unifasciata displayed adaptations dominated by minization of activity and evaporative water loss during emergence. The evolution of adaptations allowing tolerance of prolonged desiccation have allowed littorine species to dominate high intertidal rocky shore gastropod faunas throughout the world's oceans.     

One- and two-dimensional polyacrylamide gel analysis of the heat shock proteins of the virilis group of Drosophila

The heat shock proteins of the virilis group of Drosophila are analyzed by one- and two-dimensional polyacrylamide gel analysis. This group consists of the two closely related but distinct virilis and montana phylads. The analysis reveals that some of the heat shock proteins are highly conserved among the two phylads while others are not. The 83-, 72-, and 69-kdalton proteins comigrate in all species examined. There is, however, a noticable trend toward greater molecular weight variability in the smaller heat shock proteins. In general, the heat shock protein patterns within each phylad follow the proposed phylogenetic relationships with some exceptions. D. ezoana and D. littoralis, both members of the montana phylad, exhibit heat shock protein patterns more similar to those of the virilis phylad. The data also demonstrate that the montana phylad has almost two times the heat shock allele members that the virilis phylad has. It is also shown that F₁ and F₂ hybrid flies of crosses between Drosophila species having different patterns of heat shock proteins show Mendelian segregation of alleles. After several generations of inbred growth, however, the pattern of heat shock protein synthesis in reciprocal hybrids each resembles that of the paternal parent. The implications of these findings are discussed.This research was supported in part by Damon Runyon-Walter Winchell Grant DRG-233F to R.M.S. and NIH Grant GM 27611 to R.V.S. R.V.S. is the recipient of an NIH Research Career Development Award.     

Isolation and characterization of Zymomonas mobilis mutants resistant to octadecyltrimethylammonium chloride,a detergent acting on hopanoid-producing bacteria

Growth of the hopanoid-producing bacterium Zymomonas mobilis was inhibited at low concentrations of the cationic detergent octadecyltrimethylammoniumchloride (OTAC). A relationship between sensitivity of Zymomonas mobilis to OTAC, presence of hopanoids and ethanol tolerance was postulated. Mutants resistant to OTAC were isolated from strains ZM1 and ZM4. They did not present any alteration of the hopanoid content and their squalene cyclases showed the same sensitity to OTAC as the parent enzymes. Resistance to OTAC paralleled pleiotropic effects including, enhanced accessibility of the membrane-bound alkaline phosphatase, important release of proteins from cells by Tris/HCl treatment, increased resistance to antibiotics and increased sensitivity to ethanol. In addition, OTAC^R mutants were also characterized by the synthesis or the overproduction of an outer membrane protein (F53) not detected on 2D-PAGE maps of parent strains and by a normal heat shock response. The role of hopanoids, heat shock proteins, protein F53 and membrane organization in ethanol tolerance is discussed.Abbreviations OTAC octadecyltrimethylammoniumchloride - SLS sodium lauryl sarcosinate     

Genome-wide expression analysis of yeast response during exposure to 4°C

Adaptation to temperature fluctuation is essential for the survival of all living organisms. Although extensive research has been done on heat and cold shock responses, there have been no reports on global responses to cold shock below 10°C or near-freezing. We examined the genome-wide expression in Saccharomyces cerevisiae, following exposure to 4°C. Hierarchical cluster analysis showed that the gene expression profile following 4°C exposure from 6 to 48 h was different from that at continuous 4°C culture. Under 4°C exposure, the genes involved in trehalose and glycogen synthesis were induced, suggesting that biosynthesis and accumulation of those reserve carbohydrates might be necessary for cold tolerance and energy preservation. The observed increased expression of phospholipids, mannoproteins, and cold shock proteins (e.g., TIP1) is consistent with membrane maintenance and increased permeability of the cell wall at 4°C. The induction of heat shock proteins and glutathione at 4°C may be required for revitalization of enzyme activity, and for detoxification of active oxygen species, respectively. The genes with these functions may provide the ability of cold tolerance and adaptation to yeast cells.     

The Giardia lamblia actin gene and the phylogeny of eukaryotes

The single-copy actin gene of Giardia lamblia lacks introns; it has an average of 58% amino acid identity with the actin of other species; and 49 of its amino acids can be aligned with the amino acids of a consensus sequence of heat shock protein 70. Analysis of the potential RNA secondary structure in the transcribed region of the G. lamblia actin gene and of the single-copy actin gene of nine other species did not reveal any conserved structures. The G. lamblia actin sequence was used to root the phylogenetic trees based on 65 actin protein sequences from 43 species. This tree is congruent with small-subunit rRNA trees in that it shows that oomycetes are not related to higher fungi; that kinetoplatid protozoans, green plants, fungi and animals are monophyletic groups; and that the animal and fungal lineages share a more recent common ancestor than either does with the plant lineage. In contrast to smalls-ubunit rRNA trees, this tree shows that slime molds diverged after the plant lineage. The slower rate of evolution of actin genes of slime molds relative to those of plants, fungi, and animals species might be responsible for this incongruent branching. Correspondence to: G. Drouin