BLM and BRCA1-BARD1 coordinate complementary mechanisms of joint DNA molecule resolution.

The Bloom syndrome helicase BLM interacts with topoisomerase IIIα (TOP3A), RMI1, and RMI2 to form the BTR complex, which dissolves double Holliday junctions and DNA replication intermediates to promote sister chromatid disjunction before cell division. In its absence, structure-specific nucleases like the SMX complex (comprising SLX1-SLX4, MUS81-EME1, and XPF-ERCC1) can cleave joint DNA molecules instead, but cells deficient in both BTR and SMX are not viable. Here, we identify a negative genetic interaction between BLM loss and deficiency in the BRCA1-BARD1 tumor suppressor complex. We show that this is due to a previously overlooked role for BARD1 in recruiting SLX4 to resolve DNA intermediates left unprocessed by BLM in the preceding interphase. Consequently, cells with defective BLM and BRCA1-BARD1 accumulate catastrophic levels of chromosome breakage and micronucleation, leading to cell death. Thus, we reveal mechanistic insights into SLX4 recruitment to DNA lesions, with potential clinical implications for treating BRCA1-deficient tumors.

FANCM regulates repair pathway choice at stalled replication forks.

Repair pathway "choice" at stalled mammalian replication forks is an important determinant of genome stability; however, the underlying mechanisms are poorly understood. FANCM encodes a multi-domain scaffolding and motor protein that interacts with several distinct repair protein complexes at stalled forks. Here, we use defined mutations engineered within endogenous Fancm in mouse embryonic stem cells to study how Fancm regulates stalled fork repair. We find that distinct FANCM repair functions are enacted by molecularly separable scaffolding domains. These findings define FANCM as a key mediator of repair pathway choice at stalled replication forks and reveal its molecular mechanism. Notably, mutations that inactivate FANCM ATPase function disable all its repair functions and "trap" FANCM at stalled forks. We find that Brca1 hypomorphic mutants are synthetic lethal with Fancm null or Fancm ATPase-defective mutants. The ATPase function of FANCM may therefore represent a promising "druggable" target for therapy of BRCA1-linked cancer.

EXO5-DNA structure and BLM interactions direct DNA resection critical for ATR-dependent replication restart.

Stalled DNA replication fork restart after stress as orchestrated by ATR kinase, BLM helicase, and structure-specific nucleases enables replication, cell survival, and genome stability. Here we unveil human exonuclease V (EXO5) as an ATR-regulated DNA structure-specific nuclease and BLM partner for replication fork restart. We find that elevated EXO5 in tumors correlates with increased mutation loads and poor patient survival, suggesting that EXO5 upregulation has oncogenic potential. Structural, mechanistic, and mutational analyses of EXO5 and EXO5-DNA complexes reveal a single-stranded DNA binding channel with an adjacent ATR phosphorylation motif (T88Q89) that regulates EXO5 nuclease activity and BLM binding identified by mass spectrometric analysis. EXO5 phospho-mimetic mutant rescues the restart defect from EXO5 depletion that decreases fork progression, DNA damage repair, and cell survival. EXO5 depletion furthermore rescues survival of FANCA-deficient cells and indicates EXO5 functions epistatically with SMARCAL1 and BLM. Thus, an EXO5 axis connects ATR and BLM in directing replication fork restart.

Molecular forecasting of domoic acid during a pervasive toxic diatom bloom.

In 2015, the largest recorded harmful algal bloom (HAB) occurred in the Northeast Pacific, causing nearly 100 million dollars in damages to fisheries and killing many protected marine mammals. Dominated by the toxic diatom Pseudo-nitzschia australis, this bloom produced high levels of the neurotoxin domoic acid (DA). Through molecular and transcriptional characterization of 52 near-weekly phytoplankton net-tow samples collected at a bloom hotspot in Monterey Bay, California, we identified active transcription of known DA biosynthesis (dab) genes from the three identified toxigenic species, including P. australis as the primary origin of toxicity. Elevated expression of silicon transporters (sit1) during the bloom supports the previously hypothesized role of dissolved silica (Si) exhaustion in contributing to bloom physiology and toxicity. We find that coexpression of the dabA and sit1 genes serves as a robust predictor of DA one week in advance, potentially enabling the forecasting of DA-producing HABs. We additionally present evidence that low levels of iron could have colimited the diatom population along with low Si. Iron limitation represents an overlooked driver of both toxin production and ecological success of the low-iron-adapted Pseudo-nitzschia genus during the 2015 bloom, and increasing pervasiveness of iron limitation may fuel the escalating magnitude and frequency of toxic Pseudo-nitzschia blooms globally. Our results advance understanding of bloom physiology underlying toxin production, bloom prediction, and the impact of global change on toxic blooms.

DNA Bloom Filter enables anti-contamination and file version control for DNA-based data storage.

DNA storage is one of the most promising ways for future information storage due to its high data storage density, durable storage time and low maintenance cost. However, errors are inevitable during synthesizing, storing and sequencing. Currently, many error correction algorithms have been developed to ensure accurate information retrieval, but they will decrease storage density or increase computing complexity. Here, we apply the Bloom Filter, a space-efficient probabilistic data structure, to DNA storage to achieve the anti-error, or anti-contamination function. This method only needs the original correct DNA sequences (referred to as target sequences) to produce a corresponding data structure, which will filter out almost all the incorrect sequences (referred to as non-target sequences) during sequencing data analysis. Experimental results demonstrate the universal and efficient filtering capabilities of our method. Furthermore, we employ the Counting Bloom Filter to achieve the file version control function, which significantly reduces synthesis costs when modifying DNA-form files. To achieve cost-efficient file version control function, a modified system based on yin-yang codec is developed.

Mechanism of Bloom syndrome complex assembly required for double Holliday junction dissolution and genome stability.

The RecQ-like helicase BLM cooperates with topoisomerase IIIα, RMI1, and RMI2 in a heterotetrameric complex (the "Bloom syndrome complex") for dissolution of double Holliday junctions, key intermediates in homologous recombination. Mutations in any component of the Bloom syndrome complex can cause genome instability and a highly cancer-prone disorder called Bloom syndrome. Some heterozygous carriers are also predisposed to breast cancer. To understand how the activities of BLM helicase and topoisomerase IIIα are coupled, we purified the active four-subunit complex. Chemical cross-linking and mass spectrometry revealed a unique architecture that links the helicase and topoisomerase domains. Using biochemical experiments, we demonstrated dimerization mediated by the N terminus of BLM with a 2:2:2:2 stoichiometry within the Bloom syndrome complex. We identified mutations that independently abrogate dimerization or association of BLM with RMI1, and we show that both are dysfunctional for dissolution using in vitro assays and cause genome instability and synthetic lethal interactions with GEN1/MUS81 in cells. Truncated BLM can also inhibit the activity of full-length BLM in mixed dimers, suggesting a putative mechanism of dominant-negative action in carriers of BLM truncation alleles. Our results identify critical molecular determinants of Bloom syndrome complex assembly required for double Holliday junction dissolution and maintenance of genome stability.

Draft genome of novel cyanobacteria Sphaerothrix Gracilis isolated from coastal microplastics reveal insights to chemical ecology, bloom and plastic-utilization potential.

Cyanobacteria are oxygenic photosynthetic organisms which are found across many ecosystems, including freshwater and marine habitats. They are also found on natural and artificial surfaces. In this study, we cultured and characterise a novel cyanobacterium from the surfaces of foam microplastics of tropical coastal waters. We study the chemical ecology of this cyanobacterium, Sphaerothrix gracilis gen. et sp. nov., together with its potential to form harmful cyanobacterial blooms and bioremediation applications to combat plastic pollution. The genome of S. gracilis spanned 6.7 Mbp, with identification of antibiotic resistance, nitrogen-fixation, plastic-degrading and genes involved in harmful metabolite production. The transport of potentially harmful S. gracilis in coastal environments could have severe implications on human health and food security, especially in times of a cyanobacterial bloom.

The dominant white color trait of the melon fruit rind is associated with epicuticular wax accumulation.

MAIN CONCLUSION: Microscopic analyses and chemical profiling demonstrate that the white rind phenotype in melon fruit is associated with the accumulation of n-alkanes, fatty alcohols, aldehydes and wax esters. Serving as an indicator of quality, the rind (or external) color of fruit directly affects consumer choice. A fruit's color is influenced by factors such as the levels of pigments and deposited epicuticular waxes. The latter produces a white-grayish coating often referred to as "wax bloom". Previous reports have suggested that some melon (Cucumis melo L.) accessions may produce wax blooms, where a dominant white rind color trait was genetically mapped to a major locus on chromosome 7 and suggested to be inherited as a single gene named Wi. We here provide the first direct evidence of the contribution of epicuticular waxes to the dominant white rind trait in melon fruit. Our light and electron microscopy and gas chromatography-mass spectrometry (GC-MS) comparative analysis of melon accessions with white or green rinds reveals that the rind of melon fruit is rich in epicuticular waxes. These waxes are composed of various biochemical classes, including fatty acids, fatty alcohols, aldehydes, fatty amides, n-alkanes, tocopherols, triterpenoids, and wax esters. We show that the dominant white rind phenotype in melon fruit is associated with increased accumulation of n-alkanes, fatty alcohols, aldehydes and wax esters, which are linked with the deposition of crystal-like wax platelets on their surfaces. Together, this study broadens the understanding of natural variation in an important quality trait of melon fruit and promotes the future identification of the causative gene for the dominant white rind trait.

Environmental and molecular regulation of diapause formation in a scyphozoan jellyfish.

Understanding the mechanisms underlying diapause formation is crucial for gaining insight into adaptive survival strategies across various species. In this study, we aimed to uncover the pivotal role of temperature and food availability in regulating diapausing podocyst formation in the jellyfish Aurelia coerulea. Furthermore, we explored the cellular and molecular basis of diapause formation using single-cell RNA sequencing. Our results showed cell-type-specific transcriptional landscapes during podocyst formation, which were underscored by the activation of specific transcription factors and signalling pathways. In addition, we found that the heat shock protein-coding genes HSC70 and HSP90a potentially act as hub genes that regulate podocyst formation. Finally, we mapped the single-cell atlas of diapausing podocysts and identified cell types involved in metabolism, environmental sensing, defence and development that may collectively contribute to the long-term survival and regulated excystment of diapausing podocysts. Taken together, the findings of this study provide novel insights into the molecular mechanisms that regulate diapause formation and contributes to a better understanding of adaptive survival strategies in a variety of ecological contexts.

Genetic factors acting prior to dormancy in sour cherry influence bloom time the following spring.

Understanding the process of Prunus species floral development is crucial for developing strategies to manipulate bloom time and prevent crop loss due to climate change. Here, we present a detailed examination of flower development from initiation until bloom for early- and late-blooming sour cherries (Prunus cerasus) from a population segregating for a major bloom time QTL on chromosome 4. Using a new staging system, we show floral buds from early-blooming trees were persistently more advanced than those from late-blooming siblings. A genomic DNA coverage analysis revealed the late-blooming haplotype of this QTL, k, is located on a subgenome originating from the late-blooming P. fruticosa progenitor. Transcriptome analyses identified many genes within this QTL as differentially expressed between early- and late-blooming trees during the vegetative-to-floral transition. From these, we identified candidate genes for the late bloom phenotype, including multiple transcription factors homologous to Reproductive Meristem B3 domain-containing proteins. Additionally, we determined that the basis of k in sour cherry is likely separate from candidate genes found in sweet cherry-suggesting several major regulators of bloom time are located on Prunus chromosome 4.

Salinity decline promotes growth and harmful blooms of a toxic alga by diverting carbon flow.

Global climate change intensifies the water cycle and makes freshest waters become fresher and vice-versa. But how this change impacts phytoplankton in coastal, particularly harmful algal blooms (HABs), remains poorly understood. Here, we monitored a coastal bay for a decade and found a significant correlation between salinity decline and the increase of Karenia mikimotoi blooms. To examine the physiological linkage between salinity decreases and K. mikimotoi blooms, we compare chemical, physiological and multi-omic profiles of this species in laboratory cultures under high (33) and low (25) salinities. Under low salinity, photosynthetic efficiency and capacity as well as growth rate and cellular protein content were significantly higher than that under high salinity. More strikingly, the omics data show that low salinity activated the glyoxylate shunt to bypass the decarboxylation reaction in the tricarboxylic acid cycle, hence redirecting carbon from CO2 release to biosynthesis. Furthermore, the enhanced glyoxylate cycle could promote hydrogen peroxide metabolism, consistent with the detected decrease in reactive oxygen species. These findings suggest that salinity declines can reprogram metabolism to enhance cell proliferation, thus promoting bloom formation in HAB species like K. mikimotoi, which has important ecological implications for future climate-driven salinity declines in the coastal ocean with respect to HAB outbreaks.

Rare case of myelodysplastic syndrome with excess blasts 2 developing after adjuvant chemoradiotherapy for triple-negative breast cancer in a patient with Bloom syndrome.

INTRODUCTION: Bloom syndrome (BS) is a rare autosomal recessive disorder caused by a loss-of-function mutation in the BLM gene encoding an RecQ helicase involved in DNA repair and maintenance of chromosomal stability. In patients with BS, significant sensitivity to both DNA-damaging chemotherapy (CT) and ionizing radiation complicates the management of neoplasms by exacerbating comorbidities and predisposing to toxicities and poor outcomes. CASE REPORT: A 30-year-old female patient diagnosed with BS who presented with early-stage triple-negative breast cancer was treated with four cycles of doxorubicin (60 mg/m2) and cyclophosphamide (600 mg/m2) followed by weekly paclitaxel (80 mg/m2) for 12 weeks as the chemotherapy protocol and a total of 5000 cGy curative radiotherapy (RT). Due to pancytopenia 8 months after completion of therapy, bone marrow biopsy and aspiration were performed, and a diagnosis of myelodysplastic syndrome with excess blasts 2 (MDS-EB2) was made. Two courses of the azacitidine (75 mg/m2) protocol were administered every 28 days in the hematology clinic. Two weeks after CT the patient was transferred from the emergency department to the hematology clinic with the diagnosis of pancytopenia and febrile neutropenia. She died at the age of 33 due to sepsis that developed during follow-up. CONCLUSION: Due to the rarity of BS, there is no prospective trial in patients with cancer and no evidence base upon which to design treatment programs. For these reasons, it is strongly recommended that patients receive multidisciplinary care, with precise assessment and discussion of the indication and an adequate dose of DNA-damaging agents such as chemotherapy and ionizing radiation.

An assessment of a biosensor system for the quantification of microcystins in freshwater cyanobacterial blooms.

Microcystin-producing cyanobacterial blooms are a global issue threatening drinking water supplies and recreation on lakes and beaches. Direct measurement of microcystins is the only way to ensure waters have concentrations below guideline concentrations; however, analyzing water for microcystins takes several hours to days to obtain data. We tested LightDeck Diagnostics' bead beater cell lysis and two versions of the quantification system designed to give microcystin concentrations within 20 min and compared it to the standard freeze-thaw cycle lysis method and ELISA quantification. The bead beater lyser was only 30 % effective at extracting microcystins compared to freeze-thaw. When considering freeze-thaw samples analyzed in 2021, there was good agreement between ELISA and LightDeck version 2 (n = 152; R2 = 0.868), but the LightDeck slightly underestimated microcystins (slope of 0.862). However, we found poor relationships between LightDeck version 2 and ELISA in 2022 (n = 49, slopes 0.60 to 1.6; R2 < 0.6) and LightDeck version 1 (slope = 1.77 but also a high number of less than quantifiable concentrations). After the quantification issues are resolved, combining the LightDeck system with an already-proven rapid lysis method (such as microwaving) will allow beach managers and water treatment operators to make quicker, well-informed decisions.

Nutrient enrichment and phytoplankton toxicity influence a diversity of complex dynamics in a fear-induced plankton-fish model.

In this paper, we contemplate the dynamics of an aquatic system consisting of three interacting species, phytoplankton, zooplankton, and fish. We assume that the evading risk of fish predation induces fear in zooplankton species, which affects its growth dynamics radically. On the other hand, zooplankton develop an anti-predator defense by taking temporary refuge. Interestingly, the system potentially exhibits multi-stable configurations under identical ecological conditions by allowing different bifurcation scenarios, including multiple saddle-node and transcritical bifurcations with varying levels of nutrients, strength of phytoplankton toxicity, zooplankton refuge size and the cost of fear imposed by fish population. Further, by adding Gaussian white noise, we have extended the deterministic system to its stochastic version. We find that white noise appears to regulate the survival and extinction of model species. Comprehensive numerical simulations are consistent with mathematical results prognosticated by linear analysis. Overall, our study may provide a new insight into the mechanisms of emergence and mitigation of plankton blooms.

Oscillations of algal cell quota: Considering two-stage phosphate uptake kinetics.

Elucidating the mechanism of effect of phosphate (PO43-) uptake on the growth of algal cells helps understand the frequent outbreaks of algal blooms caused by eutrophication. In this study, we develop a comprehensive mathematical model that incorporates two stages of PO43- uptake and accounts for transport time delay. The model parameter values are determined by fitting experimental data of Prorocentrum donghaiense and the model is validated using experimental data of Karenia mikimotoi. The numerical results demonstrate that the model successfully captures the general characteristics of algal growth and PO43- uptake under PO43- sufficient conditions. Significantly, the experimental and mathematical findings suggest that the time delay associated with the transfer of PO43- from the surface-adsorbed PO43- (Ps) pool to the intracellular PO43- (Pi) pool may serve as a physiologically plausible mechanism leading to oscillations of algal cell quota. These results have important implications for resource managers, enabling them to predict and deepen their understanding of harmful algal blooms.

Sphingobium cyanobacteriorum sp. nov., isolated from fresh water.

A novel Gram-stain-negative, yellow-pigmented, short rod-shaped bacterial strain, HBC34T, was isolated from a freshwater sample collected from Daechung Reservoir, Republic of Korea. The results of 16S rRNA gene sequence analysis indicated that HBC34T was affiliated with the genus Sphingobium and shared the highest sequence similarity to the type strains of Sphingobium vermicomposti (98.01 %), Sphingobium psychrophilum (97.87 %) and Sphingobium rhizovicinum (97.59 %). The average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) values between HBC34T and species of the genus Sphingobium with validly published names were below 84.01 and 28.1 %, respectively. These values were lower than the accepted species-delineation thresholds, supporting its recognition as representing a novel species of the genus Sphingobium. The major fatty acids (>10 % of the total fatty acids) were identified as summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The main polar lipids were phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, two phospholipids and two unidentified polar lipids. The respiratory quinone was Q-10. The genomic DNA G+C content of HBC34T was 64.04 %. The polyphasic evidence supports the classification of HBC34T as the type strain of a novel species of the genus Sphingobium, for which the name Sphingobium cyanobacteriorum sp. nov is proposed. The type strain is HBC34T (= KCTC 8002T= LMG 33140T).

Beyond the Bloom: Unraveling the Diversity, Overlap, and Stability of Free-Living and Particle-Attached Bacterial Communities in a Cyanobacteria-Dominated Hypereutrophic Lake.

In aquatic ecosystems with low nutrient levels, organic aggregates (OAs) act as nutrient hotspots, hosting a diverse range of microbial species compared to those in the water column. Lake eutrophication, marked by intensified and prolonged cyanobacterial blooms, significantly impacts material and energy cycling processes, potentially altering the ecological traits of both free-living (FL) and particle-attached (PA) bacteria. However, the extent to which observed patterns of FL and PA bacterial diversity, community assembly, and stability extend to hypereutrophic lakes remains understudied. To address this gap, we investigated bacterial diversity, composition, assembly processes, and stability within hypereutrophic Lake Xingyun. Our results revealed that FL bacterial communities exhibited higher α-diversity than PA counterparts, coupled with discernible taxonomic compositions. Both bacterial communities showed distinct seasonality, influenced by cyanobacterial bloom intensity. Environmental factors accounted for 71.1% and 54.2% of the variation among FL and PA bacteria, respectively. The assembly of the PA bacterial community was predominantly stochastic, while FL assembly was more deterministic. The FL network demonstrated greater stability, complexity, and negative interactions, indicative of competitive relationships, while the PA network showed a prevalence of positive correlations, suggesting mutualistic interactions. Importantly, these findings differ from observations in oligotrophic, mesotrophic, and eutrophic lakes. Overall, this research provides valuable insights into the interplay among bacterial fractions, enhancing our understanding of nutrient status and cyanobacterial blooms in shaping bacterial communities.

CyanoStrainChip: A Novel DNA Microarray Tool for High-Throughput Detection of Environmental Cyanobacteria at the Strain Level.

Detecting cyanobacteria in environments is an important concern due to their crucial roles in ecosystems, and they can form blooms with the potential to harm humans and nonhuman entities. However, the most widely used methods for high-throughput detection of environmental cyanobacteria, such as 16S rRNA sequencing, typically provide above-species-level resolution, thereby disregarding intraspecific variation. To address this, we developed a novel DNA microarray tool, termed the CyanoStrainChip, that enables strain-level comprehensive profiling of environmental cyanobacteria. The CyanoStrainChip was designed to target 1277 strains; nearly all major groups of cyanobacteria are included by implementing 43,666 genome-wide, strain-specific probes. It demonstrated strong specificity by in vitro mock community experiments. The high correlation (Pearson's R > 0.97) between probe fluorescence intensities and the corresponding DNA amounts (ranging from 1-100 ng) indicated excellent quantitative capability. Consistent cyanobacterial profiles of field samples were observed by both the CyanoStrainChip and next-generation sequencing methods. Furthermore, CyanoStrainChip analysis of surface water samples in Lake Chaohu uncovered a high intraspecific variation of abundance change within the genus Microcystis between different severity levels of cyanobacterial blooms, highlighting two toxic Microcystis strains that are of critical concern for Lake Chaohu harmful blooms suppression. Overall, these results suggest a potential for CyanoStrainChip as a valuable tool for cyanobacterial ecological research and harmful bloom monitoring to supplement existing techniques.

Tracking Extensive Portfolio of Cyanotoxins in Five-Year Lake Survey and Identifying Indicator Metabolites of Cyanobacterial Taxa.

Cyanobacterial blooms require monitoring, as they pose a threat to ecosystems and human health, especially by the release of toxins. Along with widely reported microcystins, cyanobacteria coproduce other bioactive metabolites; however, information about their dynamics in surface waters is sparse. We investigated dynamics across full bloom successions throughout a five-year lake monitoring campaign (Greifensee, Switzerland) spanning 150 sampling dates. We conducted extensive suspect screening of cyanobacterial metabolites using the database CyanoMetDB. Across all 850 samples, 35 metabolites regularly co-occurred. Microcystins were present in 70% of samples, with [d-Asp3,(E)-Dhb7]MC-RR reaching concentrations of 70 ng/L. Anabaenopeptins, meanwhile, were detected in 95% of all samples with concentrations of Oscillamide Y up to 100-fold higher than microcystins. Based on LC-MS response and frequency, we identified indicator metabolites exclusively produced by one of three cyanobacteria isolated from the lake, these being [d-Asp3,(E)-Dhb7]MC-RR from Planktothrix sp. G2020, Microginin 761B from Microcystis sp. G2011, and Ferintoic acid B from Microcystis sp. G2020. These indicators showed distinct temporal trends and peaking seasons that reflect the variance in either the abundance of the producing cyanobacteria or their toxin production dynamics. Our approach demonstrates that selecting high LC-MS response and frequent and species-specific indicator metabolites can be advantageous for cyanobacterial monitoring.

Microcystin-LR Induces Estrogenic Effects at Environmentally Relevant Concentration in Black-Spotted Pond Frogs (Pelophylax nigromaculatus): In Situ, In Vivo, In Vitro, and In Silico Investigations.

Harmful cyanobacterial blooms are frequent and intense worldwide, creating hazards for aquatic biodiversity. The potential estrogen-like effect of Microcystin-LR (MC-LR) is a growing concern. In this study, we assessed the estrogenic potency of MC-LR in black-spotted frogs through combined field and laboratory approaches. In 13 bloom areas of Zhejiang province, China, the MC-LR concentrations in water ranged from 0.87 to 8.77 µg/L and were correlated with sex hormone profiles in frogs, suggesting possible estrogenic activity of MC-LR. Tadpoles exposed to 1 µg/L, an environmentally relevant concentration, displayed a female-biased sex ratio relative to controls. Transcriptomic results revealed that MC-LR induces numerous and complex effects on gene expression across multiple endocrine axes. In addition, exposure of male adults significantly increased the estradiol (E2)/testosterone (T) ratio by 3.5-fold relative to controls. Downregulation of genes related to male reproductive endocrine function was also identified. We also showed how MC-LR enhances the expression of specific estrogen receptor (ER) proteins, which induce estrogenic effects by activating the ER pathway and hypothalamic-pituitary-gonadal (HPG) axis. In aggregate, our results reveal multiple lines of evidence demonstrating that, for amphibians, MC-LR is an estrogenic endocrine disruptor at environmentally relevant concentrations. The data presented here support the need for a shift in the MC-LR risk assessment. While hepatoxicity has historically been the focus of MC-LR risk assessments, our data clearly demonstrate that estrogenicity is a major mode of toxicity at environmental levels and that estrogenic effects should be considered for risk assessments on MC-LR going forward.