Manipulation of photosensory and circadian signaling restricts phenotypic plasticity in response to changing environmental conditions in Arabidopsis.

Plants exploit phenotypic plasticity to adapt their growth and development to prevailing environmental conditions. Interpretation of light and temperature signals is aided by the circadian system, which provides a temporal context. Phenotypic plasticity provides a selective and competitive advantage in nature but is obstructive during large-scale, intensive agricultural practices since economically important traits (including vegetative growth and flowering time) can vary widely depending on local environmental conditions. This prevents accurate prediction of harvesting times and produces a variable crop. In this study, we sought to restrict phenotypic plasticity and circadian regulation by manipulating signaling systems that govern plants' responses to environmental signals. Mathematical modeling of plant growth and development predicted reduced plant responses to changing environments when circadian and light signaling pathways were manipulated. We tested this prediction by utilizing a constitutively active allele of the plant photoreceptor phytochrome B, along with disruption of the circadian system via mutation of EARLY FLOWERING3. We found that these manipulations produced plants that are less responsive to light and temperature cues and thus fail to anticipate dawn. These engineered plants have uniform vegetative growth and flowering time, demonstrating how phenotypic plasticity can be limited while maintaining plant productivity. This has significant implications for future agriculture in both open fields and controlled environments.

Spatial and cumulative organochlorine and mercury exposure assessments in Steller Sea lions of Alaska: Emphasizing pups.

Steller sea lions (SSL) are sentinels for monitoring environmental contaminants in remote areas of the Aleutian Islands, Alaska. Therefore, concentrations of several organochlorines (OCs) were measured in blood from 123 SSL pups sampled from 3 regions; the western Aleutian Islands (WAI), central Aleutian Islands (CAI), and the central Gulf of Alaska. Blood, blubber, and milk from 12 adult female SSL from WAI, CAI and southeast Alaska also were analyzed. Findings included the following. SSL pups had higher concentrations of some OCs and mercury (Hg) on rookeries in the WAI than those more easterly. Pups had significantly higher blood concentrations of many OC classes than adult females sampled within the same region; some pups had PCB concentrations exceeding thresholds of concern (∑PCBs >8600 ng/g lw). ∑PCB concentration in pup whole blood was positively correlated with the trophic marker, δ15N within the regions sampled, along with two PCB congeners (PCB138 and PCB153). This suggests that the dams of pups with higher ∑PCBs, PCB138, and PCB153 concentrations were feeding on more predatory prey. Adult female blubber ∑DDT and hexachlorocyclohexane concentrations were also positively correlated with δ15N values. Several pups (mostly from WAI) had blood Hg concentrations and/or blood PCB concentrations (surrogate for overall OC exposures) of concern. The finding that WAI SSL pups have been exposed to multiple contaminants calls for future investigation of their cumulative exposure to a mixture of contaminants especially their transplacental and then transmammary exposure routes.

Integrating analog and digital modes of gene expression at Arabidopsis FLC.

Quantitative gene regulation at the cell population level can be achieved by two fundamentally different modes of regulation at individual gene copies. A 'digital' mode involves binary ON/OFF expression states, with population-level variation arising from the proportion of gene copies in each state, while an 'analog' mode involves graded expression levels at each gene copy. At the Arabidopsis floral repressor FLOWERING LOCUS C (FLC), 'digital' Polycomb silencing is known to facilitate quantitative epigenetic memory in response to cold. However, whether FLC regulation before cold involves analog or digital modes is unknown. Using quantitative fluorescent imaging of FLC mRNA and protein, together with mathematical modeling, we find that FLC expression before cold is regulated by both analog and digital modes. We observe a temporal separation between the two modes, with analog preceding digital. The analog mode can maintain intermediate expression levels at individual FLC gene copies, before subsequent digital silencing, consistent with the copies switching OFF stochastically and heritably without cold. This switch leads to a slow reduction in FLC expression at the cell population level. These data present a new paradigm for gradual repression, elucidating how analog transcriptional and digital epigenetic memory pathways can be integrated.

Feeling Every Bit of Winter - Distributed Temperature Sensitivity in Vernalization.

Temperature intrinsically influences all aspects of biochemical and biophysical processes. Organisms have therefore evolved strategies to buffer themselves against thermal perturbations. Many organisms also use temperature signals as cues to align behavior and development with certain seasons. These developmentally important thermosensory mechanisms have generally been studied in constant temperature conditions. However, environmental temperature is an inherently noisy signal, and it has been unclear how organisms reliably extract specific temperature cues from fluctuating temperature profiles. In this context, we discuss plant thermosensory responses, focusing on temperature sensing throughout vernalization in Arabidopsis. We highlight many different timescales of sensing, which has led to the proposal of a distributed thermosensing paradigm. Within this paradigm, we suggest a classification system for thermosensors. Finally, we focus on the longest timescale, which is most important for sensing winter, and examine the different mechanisms in which memory of cold exposure can be achieved.

Natural variation in autumn expression is the major adaptive determinant distinguishing Arabidopsis FLC haplotypes.

In Arabidopsis thaliana, winter is registered during vernalization through the temperature-dependent repression and epigenetic silencing of floral repressor FLOWERING LOCUS C (FLC). Natural Arabidopsis accessions show considerable variation in vernalization. However, which aspect of the FLC repression mechanism is most important for adaptation to different environments is unclear. By analysing FLC dynamics in natural variants and mutants throughout winter in three field sites, we find that autumnal FLC expression, rather than epigenetic silencing, is the major variable conferred by the distinct Arabidopsis FLChaplotypes. This variation influences flowering responses of Arabidopsis accessions resulting in an interplay between promotion and delay of flowering in different climates to balance survival and, through a post-vernalization effect, reproductive output. These data reveal how expression variation through non-coding cis variation at FLC has enabled Arabidopsis accessions to adapt to different climatic conditions and year-on-year fluctuations.

Publisher Correction: Temperature-dependent growth contributes to long-term cold sensing.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Temperature-dependent growth contributes to long-term cold sensing.

Temperature is a key factor in the growth and development of all organisms1,2. Plants have to interpret temperature fluctuations, over hourly to monthly timescales, to align their growth and development with the seasons. Much is known about how plants respond to acute thermal stresses3,4, but the mechanisms that integrate long-term temperature exposure remain unknown. The slow, winter-long upregulation of VERNALIZATION INSENSITIVE 3 (VIN3)5-7, a PHD protein that functions with Polycomb repressive complex 2 to epigenetically silence FLOWERING LOCUS C (FLC) during vernalization, is central to plants interpreting winter progression5,6,8-11. Here, by a forward genetic screen, we identify two dominant mutations of the transcription factor NTL8 that constitutively activate VIN3 expression and alter the slow VIN3 cold induction profile. In the wild type, the NTL8 protein accumulates slowly in the cold, and directly upregulates VIN3 transcription. Through combining computational simulation and experimental validation, we show that a major contributor to this slow accumulation is reduced NTL8 dilution due to slow growth at low temperatures. Temperature-dependent growth is thus exploited through protein dilution to provide the long-term thermosensory information for VIN3 upregulation. Indirect mechanisms involving temperature-dependent growth, in addition to direct thermosensing, may be widely relevant in long-term biological sensing of naturally fluctuating temperatures.

Regional variations and drivers of mercury and selenium concentrations in Steller sea lions.

Mercury (Hg) can be neurotoxic to mammals and impact reproduction, whereas selenium (Se) is an important antioxidant known to ameliorate some adverse effects of Hg. Total Hg concentrations ([THg]) were measured in lanugo (pelage grown in utero) of 812 Steller sea lion (Eumetopias jubatus) pups across Alaska and Russia to assess fetal exposure during late gestation. The molar ratio of total Se to THg (TSe:THg) was determined in whole blood collected from 291 pups. Stable isotope ratios of carbon and nitrogen were measured in sections of vibrissae (whiskers, n = 498) and in lanugo (n = 480) of pups grown during late gestation to track diet variations among adult females that can drive Hg and Se exposure during this critical fetal development period. Lanugo [THg] ranged from 1.4 to 73.7 µg/g dry weight with the lowest median [THg] in Southeast Alaska. Pups from the Western Aleutian Islands had higher median lanugo [THg] than pups from other metapopulations in Alaska. Over 25% of pups in the Western Aleutian Islands had [THg] above published risk thresholds (20 µg/g) for other mammals. Whole blood molar TSe:THg was significantly lower in the Western Aleutian Islands and in some parts of the Central Aleutian Islands with higher molar ratios found in the Eastern Aleutian Islands and Central Gulf of Alaska. This suggests a limitation on potential protective functions of Se in the western regions with the highest relative [THg]. The Central Aleutian Island pups with [THg] over 20 µg/g had higher δ15N ratios than pups with lower [THg] suggesting dams consuming higher trophic level prey is a key driver for Hg exposure. However, regional differences likely reflect variability in diet of the dam during gestation and in Hg food web dynamics between oceanic regimes east and west of key passes in the Aleutian Islands.

Noncoding SNPs influence a distinct phase of Polycomb silencing to destabilize long-term epigenetic memory at Arabidopsis FLC.

In Arabidopsis thaliana, the cold-induced epigenetic regulation of FLOWERING LOCUS C (FLC) involves distinct phases of Polycomb repressive complex 2 (PRC2) silencing. During cold, a PHD-PRC2 complex metastably and digitally nucleates H3K27me3 within FLC On return to warm, PHD-PRC2 spreads across the locus delivering H3K27me3 to maintain long-term silencing. Here, we studied natural variation in this process in Arabidopsis accessions, exploring Lov-1, which shows FLC reactivation on return to warm, a feature characteristic of FLC in perennial Brassicaceae This analysis identifies an additional phase in this Polycomb silencing mechanism downstream from H3K27me3 spreading. In this long-term silencing (perpetuated) phase, the PHD proteins are lost from the nucleation region and silencing is likely maintained by the read-write feedbacks associated with H3K27me3. A combination of noncoding SNPs in the nucleation region mediates instability in this long-term silencing phase with the result that Lov-1 FLC frequently digitally reactivates in individual cells, with a probability that diminishes with increasing cold duration. We propose that this decrease in reactivation probability is due to reduced DNA replication after flowering. Overall, this work defines an additional phase in the Polycomb mechanism instrumental in natural variation of silencing, and provides avenues to dissect broader evolutionary changes at FLC.

Viral emergence in marine mammals in the North Pacific may be linked to Arctic sea ice reduction.

Climate change-driven alterations in Arctic environments can influence habitat availability, species distributions and interactions, and the breeding, foraging, and health of marine mammals. Phocine distemper virus (PDV), which has caused extensive mortality in Atlantic seals, was confirmed in sea otters in the North Pacific Ocean in 2004, raising the question of whether reductions in sea ice could increase contact between Arctic and sub-Arctic marine mammals and lead to viral transmission across the Arctic Ocean. Using data on PDV exposure and infection and animal movement in sympatric seal, sea lion, and sea otter species sampled in the North Pacific Ocean from 2001-2016, we investigated the timing of PDV introduction, risk factors associated with PDV emergence, and patterns of transmission following introduction. We identified widespread exposure to and infection with PDV across the North Pacific Ocean beginning in 2003 with a second peak of PDV exposure and infection in 2009; viral transmission across sympatric marine mammal species; and association of PDV exposure and infection with reductions in Arctic sea ice extent. Peaks of PDV exposure and infection following 2003 may reflect additional viral introductions among the diverse marine mammals in the North Pacific Ocean linked to change in Arctic sea ice extent.

Temperature Sensing Is Distributed throughout the Regulatory Network that Controls FLC Epigenetic Silencing in Vernalization.

Many organisms need to respond to complex, noisy environmental signals for developmental decision making. Here, we dissect how Arabidopsis plants integrate widely fluctuating field temperatures over month-long timescales to progressively upregulate VERNALIZATION INSENSITIVE3 (VIN3) and silence FLOWERING LOCUS C (FLC), aligning flowering with spring. We develop a mathematical model for vernalization that operates on multiple timescales-long term (month), short term (day), and current (hour)-and is constrained by experimental data. Our analysis demonstrates that temperature sensing is not localized to specific nodes within the FLC network. Instead, temperature sensing is broadly distributed, with each thermosensory process responding to specific features of the plants' history of exposure to warm and cold. The model accurately predicts FLC silencing in new field data, allowing us to forecast FLC expression in changing climates. We suggest that distributed thermosensing may be a general property of thermoresponsive regulatory networks in complex natural environments.

Temporal records of diet diversity dynamics in individual adult female Steller sea lion (Eumetopias jubatus) vibrissae.

Detailed information on the nutrition of free-ranging mammals contributes to the understanding of life history requirements, yet is often quite limited temporally for most species. Reliable dietary inferences can be made by analyzing the stable carbon (C) and nitrogen (N) isotopic values (δ13C and δ15N) of some consumer tissues; exactly which tissue is utilized dictates the inferential scope. Steller sea lion (SSL) vibrissae are grown continuously without shedding and thus provide a continuous multi-year record of dietary consumption. We applied a novel kernel density approach to compare the δ13C and δ15N values along the length of SSL vibrissae with δ13C and δ15N distributions of potential prey species. This resulted in time-series of proportion estimates of dietary consumption for individual SSL. Substantial overlap in δ13C and δ15N distributions for prey species prevented a discrete species-scale assessment of SSL diets; however, a post hoc correlational analysis of diet proportion estimates revealed grouping by trophic level. Our findings suggest that adult female SSL diets in the western and central Aleutian Islands shift significantly according to season: diets contain a higher proportion of lower trophic level species (Pacific Ocean perch, northern rockfish, Atka mackerel and walleye pollock) in the summer, whereas in the winter SSL consume a much more diverse diet which includes a greater proportion of higher trophic level species (arrowtooth flounder, Kamchatka flounder, darkfin sculpin, Pacific cod, Pacific octopus, rock sole, snailfish, and yellow Irish lord).

Absence of warmth permits epigenetic memory of winter in Arabidopsis.

Plants integrate widely fluctuating temperatures to monitor seasonal progression. Here, we investigate the temperature signals in field conditions that result in vernalisation, the mechanism by which flowering is aligned with spring. We find that multiple, distinct aspects of the temperature profile contribute to vernalisation. In autumn, transient cold temperatures promote transcriptional shutdown of Arabidopsis FLOWERING LOCUS C (FLC), independently of factors conferring epigenetic memory. As winter continues, expression of VERNALIZATION INSENSITIVE3 (VIN3), a factor needed for epigenetic silencing, is upregulated by at least two independent thermosensory processes. One integrates long-term cold temperatures, while the other requires the absence of daily temperatures above 15 °C. The lack of spikes of high temperature, not just prolonged cold, is thus the major driver for vernalisation. Monitoring of peak daily temperature is an effective mechanism to judge seasonal progression, but is likely to have deleterious consequences for vernalisation as the climate becomes more variable.

Buckling as an origin of ordered cuticular patterns in flower petals.

The optical properties of plant surfaces are strongly determined by the shape of epidermal cells and by the patterning of the cuticle on top of the cells. Combinations of particular cell shapes with particular nanoscale structures can generate a wide range of optical effects. Perhaps most notably, the development of ordered ridges of cuticle on top of flat petal cells can produce diffraction-grating-like structures. A diffraction grating is one of a number of mechanisms known to produce 'structural colours', which are more intense and pure than chemical colours and can appear iridescent. We explore the concept that mechanical buckling of the cuticle on the petal epidermis might explain the formation of cuticular ridges, using a theoretical model that accounts for the development of compressive stresses in the cuticle arising from competition between anisotropic expansion of epidermal cells and isotropic cuticle production. Model predictions rationalize cuticle patterns, including those with long-range order having the potential to generate iridescence, for a range of different flower species.

Quality of life of patients with newly diagnosed poor prognosis M1 prostate cancer undergoing orchiectomy without or with mitomycin C. Results from the EORTC Phase-III trial 30893.

OBJECTIVES: To compare the quality of life (QL) of patients with poor prognosis M1 prostate cancer treated with orchiectomy alone (ORCH) or orchiectomy combined with adjuvant mitomycin C (MMC; 15 mg/m(2) i.v. q 6 weeks: ORCH+MMC; EORTC trial 30893). METHODS: Patients with newly diagnosed M1 poor prognosis prostate cancer completed a truncated version of the EORTC QLQ-C30 (V 1.0) at randomization (baseline) and every 6-12 weeks thereafter until going off the protocol. Five ad hoc questions assessing lower urinary tract symptoms were included in the QL questionnaire. RESULTS: At least one QL form was completed by 177 of the 189 patients included in the trial, with baseline questionnaires available for 113 patients (ORCH n = 52; ORCH+MMC n = 61). In both arms, pain and urinary dysfunction improved during treatment. Compared with patients from the ORCH arm, the use of adjuvant MMC was associated with a significant reduction in global health status/QL and with impairment in 7 of 11 QL dimensions covered by the questionnaire. Some improvement in QL was observed after discontinuation of MMC. A survival benefit was not observed in the ORCH+MMC arm. CONCLUSIONS: Intravenous MMC (15 mg/m(2) q 6 weeks) cannot be recommended as adjuvant treatment in M1 poor prognosis prostate cancer due to its negative impact on QL and lack of efficacy. In general, QL assessments should be mandatory when adjuvant chemotherapy is evaluated in patients with metastatic prostate cancer.

Orchiectomy and orchiectomy plus mitomycin C for metastatic prostate cancer in patients with poor prognosis: the final results of a European Organization for Research in Cancer Therapy Genitourinary Group Trial.

PURPOSE: The outcome of patients with symptomatic metastatic prostate cancer is poor and improved treatment regimens are urgently needed. Theoretically, the combination of orchiectomy and chemotherapy could reduce androgen sensitive and insensitive cells in the prostate. This European Organization for Research in Cancer Therapy Genitourinary Group randomized, multicenter phase III trial demonstrates the outcome of orchiectomy alone versus orchiectomy followed by intravenous mitomycin C. MATERIALS AND METHODS: A total of 189 patients with metastatic prostate cancer and poor prognostic factors were randomized in this trial by 42 institutions. Of these patients 184 (97%) were eligible for study, including 90 treated with orchiectomy alone (orchiectomy only arm) and 94 treated with orchiectomy followed by 15 mg./m.2 mitomycin C in 1 week (combined treatment arm). Mitomycin C was administered every 6 weeks and treatment was continued as long as tolerance and patient compliance allowed, and no progression was observed. Objective and subjective criteria for progression were clearly defined in the protocol. RESULTS: Patient and tumor characteristics were well balanced between the 2 treatment arms. At a median followup of 4.2 years 144 patients had died, including 112 of prostate cancer. No significant differences for time to overall (p = 0.17), subjective (p = 0.25) and objective (p = 0.08) progression were found between the 2 treatment groups. For progression-free survival no difference was noted (p = 0.67) between the 2 treatment groups but a trend in favor of orchiectomy alone was observed for overall survival (p = 0.04). Mitomycin C induced considerable hematological, gastrointestinal, renal and pulmonary toxicity leading to discontinuation in 31% of patients with pulmonary toxicity and 7% with renal deterioration. In addition, the quality of life evaluation revealed significant reduction in the combined treatment arm. CONCLUSIONS: Based on the results of this randomized phase III study orchiectomy plus mitomycin C for metastatic prostate cancer in patients with poor prognostic factors cannot be recommended due to failure of improvement in survival and reduced quality of life parameters.

Health status of young Alaska Steller sea lion pups (Eumetopias jubatus) as indicated by blood chemistry and hematology.

Blood chemistry and hematology were examined in 238 Steller sea lion pups (Eumetopias jubatus) to assess the health status of pups <1 month of age. Failure of juvenile recruitment (possibly due to nutritionally or physiologically compromised pups) into breeding populations has been proposed as a cause of recent declines of this endangered species in Alaska. To identify potential correlations with areas of high population decline, blood chemistry data were considered for three areas: eastern Aleutian Islands (low rates of population decline to stable populations), Gulf of Alaska (high rates of decline), and Southeast Alaska (stable to increasing population). Southeast Alaska pups showed elevated ketone body concentrations (beta-hydroxybutyrate,(beta-HBA)) and depressed glucose levels than pups in the Gulf of Alaska. Over 40% of the pups from Southeast Alaska had elevated beta-HBA concentrations suggesting they underwent longer periods of fasting than seen in pups from other areas. Hematocrit (Hct), hemoglobin concentration (Hb) and water content of the blood exhibited typical mammalian relationships. In summary, blood chemistry and hematology data showed no indication that Steller sea lion pups <1 month old from areas of population decline were nutritionally compromised.

Importance of bleomycin in combination chemotherapy for good-prognosis testicular nonseminoma: a randomized study of the European Organization for Research and Treatment of Cancer Genitourinary Tract Cancer Cooperative Group.

PURPOSE: This prospective randomized trial was designed to compare the efficacy of etoposide plus cisplatin (EP) versus bleomycin, etoposide, and cisplatin (BEP) chemotherapy in patients with good-prognosis metastatic nonseminomatous testicular cancer. PATIENTS AND METHODS: Four hundred nineteen patients with good-prognosis nonseminomatous testicular cancer were randomized to receive four cycles of cisplatin 20 mg/m2 on days 1 to 5 plus etoposide 120 mg/m2 on days 1, 3, and 5 with or without bleomycin 30 mg weekly. RESULTS: Of 395 eligible patients, 169 of 195 patients allocated to EP (87%) and 189 of 200 patients allocated to BEP (95%) achieved a complete response with chemotherapy alone or after postchemotherapy surgery. These results are significantly different (P = .0075). After a median follow-up duration of 7.3 years, eight patients (4%) on each treatment arm relapsed. In view of the low number of unfavorable treatment outcomes (11%), no significant differences were detected in time to progression (P = .136) and survival (P = .262). Both the acute and late pulmonary toxicity and neurotoxicity were significantly greater in patients who received BEP, whereas Raynaud's phenomenon occurred exclusively in patients who received BEP (P < .001). Two patients treated with BEP died of bleomycin pulmonary toxicity. CONCLUSION: BEP is the most effective combination regimen in the treatment of disseminated nonseminomatous germ cell cancer. In this particular BEP regimen with etoposide at a dose of 360 mg/m2 per cycle, even in good-prognosis patients, bleomycin cannot be deleted without compromising treatment efficacy, but its use is associated with more toxicity (particularly pulmonary) and efforts to reduce this merit further exploration.

Plasma haptoglobin levels in threatened Alaskan pinniped populations.

We evaluated the plasma concentration of the acute phase protein haptoglobin (Hp) from Steller sea lions (Eumetopias jubatus) and harbor seals (Phoca vitulina) in regions of Alaska (USA) where the populations of these pinnipeds were declining and compared the values with concentrations of Hp from the same species in areas where the populations were stable. Samples were collected from 1992 through 1994 at sites in Southeast Alaska, Prince William Sound, the Gulf of Alaska, and the Aleutian Islands. Significantly higher levels of Hp were found in the samples from the areas of decline compared to those from stable populations. Based on these findings, we propose that one may be able to distinguish these compromised pinniped populations using Hp as a biomedical indicator.