Hot Spot Position Research Articles

A study on the type-b hot spot stress

Stress distributions in the vicinity of type-b hot spot in the in-plane gusset welded joints and in the free edge of the stiffened cruciform welded joints were evaluated according to the solid model, the fine mesh shell model, and the coarse mesh shell model. The hot spot stress (HSS) of each FE model was evaluated by extrapolating the stresses at the read out point (ROP) to the hot spot position according to the recommendations of IIW and were compared. Regarding the in-plane gusset welded joints, it is confirmed that the HSS by the solid model and the shell model are corresponding in the case of in-plane notch in which the thicknesses of the main plate and the gusset plate are same. It is observed that the shell model estimates lower HSS than that by the solid model when the main plate is thicker than the gusset plate. The reason of this difference is considered that the shell model does not evaluate the stress distribution in the thickness direction of the main plate in front of the in-plane gusset plate but evaluate the averaging stress. A simple correction factor to the HSS evaluated by the shell model based on the ratio of both plate thicknesses was examined. Regarding the free edge of the loaded attached plate in the stiffened cruciform welded joints, it is confirmed that the hot spot is on the corner edge of the attached plate. It is observed that the shell model estimates lower HSS than that by the solid model.

De novo missense variants in the RAP1B gene identified in two patients with syndromic thrombocytopenia.

We present two independent cases of syndromic thrombocytopenia with multiple malformations, microcephaly, learning difficulties, dysmorphism and other features. Exome sequencing identified two novel de novo heterozygous variants in these patients, c.35G>T p.(Gly12Val) and c.178G>C p.(Gly60Arg), in the RAP1B gene (NM_001010942.2). These variants have not been described previously as germline variants, however functional studies in literature strongly suggest a clinical implication of these two activating hot spot positions. We hypothesize that pathogenic missense variants in the RAP1B gene cause congenital syndromic thrombocytopenia with a spectrum of associated malformations and dysmorphism, possibly through a gain of function mechanism.

Understanding the impact of DIS3 cancer-associated mutations by in silico structure modeling

DIS3 gene is reported as a driver gene specifically mutated in a few cancer types including multiple myeloma. Unraveling the molecular effects of DIS3 mutations on protein structure may provide insights into the functional effect of different substitutions. Hence, the aim of this study is to analyze the structural and functional consequences of hotspot mutations in DIS3 by using various sequence and structure-based bioinformatics methods. In addition to sequence-based programs, the effect of hotspot mutations on protein stability and the overall pathogenicity was investigated using SDM and Missense3D. Moreover, mutations were modeled on protein structure using UCSF Chimera. Changes in residue interactions after mutations were investigated using RINalyzer. In total 6 positions in DIS3 are reported as hotspot positions. Our analysis showed all hotspots were located in highly conserved positions and their substitution resulted in clashes or disruption in interaction with the Mg2+ or nucleic acid. Our findings confirmed analysis of the residue interaction network provides a better insight into the functional and structural impact of the DIS3 mutations. We expect our analysis will be used for designing future experimental evaluation of the mutations in DIS3 gene and it will also help evaluate other cancer driver genes' mutations.

Abstract A24: Identifying the genomic and clinical features of AKT1/PIK3CA mutant metastatic prostate cancer using circulating tumor DNA

Abstract Background: Hotspot activating mutations in AKT1 and PIK3CA represent a rare but potentially unique subset of metastatic prostate cancers (mPCa). The accompanying genomic and clinical features of these patients are currently unknown. Preclinical evidence suggests that unconstrained PI3K signaling via PTEN loss may render androgen receptor (AR)-targeted therapy less effective. Given the availability of agents that target nodes within the PI3K pathway, elucidating the genomic properties of AKT1/PIK3CA mutant patients and their response to AR-targeted therapy will be critical for therapeutic selection. Methods: We performed deep targeted sequencing on 1,381 cell-free DNA samples from 608 patients with mPCa. Analysis was restricted to patients with hotspot AKT1 or PI3KCA mutations of presumed clonal origin, requiring a variant allele frequency >25% of a sample’s ctDNA fraction. Activating AKT1 and PIK3CA mutations were defined as recurrent hotspot mutations with a cBioPortal annotation, as well as functionally equivalent variants within 5bp of these canonical hotspot positions. Patient records were reviewed for baseline clinical characteristics, as well as time from androgen deprivation therapy (ADT) initiation to castration resistance and overall survival (OS). Results: 5.9% (36/608) of patients harbored at least 1 clonal hotspot mutation in either AKT1 or PIK3CA, of which p.E17K and p.E545K/Q/A were most common. This population had a significantly higher ctDNA fraction compared to a control cohort of AKT1/PIK3CA wild-type mPCa patients (Mann-Whitney U test, 0.48 vs. 0.21, p < 0.001). There were no differences in PTEN copy number or mutation frequency compared to the control cohort. Although AR mutations and copy number amplifications were observed at similar frequencies, patients harboring AKT1 or PI3KCA mutations had fewer additional copies of AR (median 4.71 vs. 10.33, p=0.011, Mann-Whitney U Test). 30 patients with activating AKT1/PIK3CA mutations had clinical outcomes available. Interestingly, there were no significant differences in time to castration resistance or OS compared to ctDNA-positive patients without activating PI3K defects. Conclusions: AKT1/PIK3CA mutant mPCa is defined by a genomic landscape featuring high ctDNA-fraction and lower levels of AR amplification. Response to standard-of-care treatment among these patients is typical for ctDNA-positive mPCa. These findings may nominate patients who may benefit from PI3K-targeted therapeutics following resistance on AR-targeted therapy. Citation Format: Cameron M. Herberts, Andrew J. Murtha, Simon Fu, Gang Wang, Elena Schönlau, Anna Gleave, Steven Yip, Arkhjamil Angeles, Sebastien Hotte, Abdulraheem Alshangiti, Ben Tran, Scott North, Sinja Taavitsainen, Kevin Beja, Gillian Vandekerkhove1, Elie Ritch, Fred Saad, Nayyer Iqbal, Martin E. Gleave, Matti Annala, Kim N. Chi, Alexander W. Wyatt. Identifying the genomic and clinical features of AKT1/PIK3CA mutant metastatic prostate cancer using circulating tumor DNA [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr A24.

Acquisitions and applications of forest canopy hyperspectral imageries at hotspot and multiview angle using unmanned aerial vehicle platform

The accurate retrieval of forest functional and structural parameters is of great significance in the scientific research of ecosystem, global change, and carbon and nitrogen cycles. Recently, an unmanned aerial vehicle (UAV) hyperspectral imaging system provides a cost-effective way to capture the hyperspectral imageries from any points of the hemisphere above a forest canopy. However, compared with single-angle hyperspectral images, the multiangle hyperspectral images provide more information about forest functional and structural characteristics. We developed a semiautomatic multiangle observation method using a UAV hyperspectral imaging system and successfully collected the multiangle hyperspectral imageries with clear hotspots of broadleaf and coniferous forest canopies. Our results indicated that the hotspot of a forest canopy had a great effect on the reflectance, normalized difference vegetation index (NDVI), and enhanced vegetation index (EVI) of forests. The maximum values of canopy reflectance and EVI were found at the hotspot position, while the minimum NDVI was at the hotspot. Moreover, the hotspot effect was similar in both broadleaf and coniferous forests. Although the hotspot had no obvious effects on the photochemical reflectance index (PRI), different view zenith angles had a great effect on PRI. Our findings provide a solid foundation for retrieving forest structural parameters using fully automatic multiangle hyperspectral imaging system at both aerial and satellite platforms.

Tuning Enzyme Activity for Nonaqueous Solvents: Engineering an Enantioselective "Michaelase" for Catalysis in High Concentrations of Ethanol.

Enzymes have evolved to function under aqueous conditions and may not exhibit features essential for biocatalytic application, such as the ability to function in high concentrations of an organic solvent. Consequently, protein engineering is often required to tune an enzyme for catalysis in non‐aqueous solvents. In this study, we have used a collection of nearly all single mutants of 4‐oxalocrotonate tautomerase, which promiscuously catalyzes synthetically useful Michael‐type additions of acetaldehyde to various nitroolefins, to investigate the effect of each mutation on the ability of this enzyme to retain its “Michaelase” activity in elevated concentrations of ethanol. Examination of this mutability landscape allowed the identification of two hotspot positions, Ser30 and Ala33, at which mutations are beneficial for catalysis in high ethanol concentrations. The “hotspot” position Ala33 was then randomized in a highly enantioselective, but ethanol‐sensitive 4‐OT variant (L8F/M45Y/F50A) to generate an improved enzyme variant (L8F/A33I/M45Y/F50A) that showed great ethanol stability and efficiently catalyzes the enantioselective addition of acetaldehyde to nitrostyrene in 40 % ethanol (permitting high substrate loading) to give the desired γ‐nitroaldehyde product in excellent isolated yield (89 %) and enantiopurity (ee=98 %). The presented work demonstrates the power of mutability‐landscape‐guided enzyme engineering for efficient biocatalysis in non‐aqueous solvents.

Research and Experiment on Quench Detection Technology of HTS Current Leads Based on Distributed Optical Fibers

High-temperature superconductor (HTS) current leads are important components of the EAST and CFETR tokamaks, which are responsible for operating the high parametric current. HTS current leads are made of Bi-2223/Ag-Au alloy tapes, which have the characteristics of slow quench propagation speed and weak quench signal. Traditional thermometers are easily damaged by the high voltage from the current leads, and the terminal voltage signal cannot reflect the hot spot changes of current leads in real time. In this paper, a novel quench detection method based on optical frequency domain reflection technology is proposed. Temperature variations of HTS can be obtained in real time by demodulating the Rayleigh scattered spectrum from the distributed optical fiber attached to the surface of HTS stacks. This paper describes a quenching experiment for one pair of 1-kA small current leads. The external thermal disturbance is increased to explore the quench propagation of HTS current leads under the condition of a self-field, 77 to 80 K air and conduction combined cooling method. From the experimental results, the temperature distribution map of the whole HTS lead is obtained. Compared with the quench voltage, the spectral shift of the fiber appeared to be about 2 s ahead, and the hot spot position can be located with 1-cm accuracy.

CRISPR-sub: Analysis of DNA substitution mutations caused by CRISPR-Cas9 in human cells

CRISPR-Cas9 induces DNA cleavages at desired target sites in a guide RNA-dependent manner; DNA editing occurs through the resulting activity of DNA repair processes including non-homologous end joining (NHEJ), which is dominant in mammalian cells. NHEJ repair frequently causes small insertions and deletions (indels) near DNA cleavage sites but only rarely causes nucleotide substitutions. High-throughput sequencing is the primary means of assessing indel and substitution frequencies in bulk populations of cells in the gene editing field. However, it is difficult to detect bona fide substitutions, which are embedded among experimentally-induced substitution errors, in high-throughput sequencing data. Here, we developed a novel analysis method, named CRISPR-Sub, to statistically detect Cas9-mediated substitutions in high-throughput sequencing data by comparing Mock- and CRISPR-treated samples. We first pinpointed ‘hotspot positions’ in target sequences at which substitution mutations were quantitatively observed much more often (p > 0.001) in CRISPR- versus Mock-treated samples. We refer to the substitution mutations in defined hotspot positions as ‘apparent substitutions’ and ultimately calculated ‘apparent substitution frequencies’ for each target. By examining 51 endogenous target sites in HeLa cells, we found that the average apparent substitution frequency was 0.8% in all queries, that apparent substitutions frequently occur near CRISPR-Cas9 cleavage sites, and that nucleotide conversion showed no meaningful nucleotide preference patterns. Furthermore, we generated NHEJ-inhibited cell lines (LIG4−/−) by knockout of the gene encoding ligase IV and found that the apparent substitution frequencies were significantly decreased in LIG4−/− cells, strongly suggesting that DNA substitutions are generated by the NHEJ pathway.

Thermal Field Analysis for New AlGaN/GaN HEMT With Partial Etched AlGaN Layer

In order to explore the distribution of the device temperature field, this paper takes the new AlGaN/GaN HEMT with partial etched AlGaN layer as the research object. First, the ISE TCAD software is used to simulate the temperature field of AlGaN/GaN HEMT in two dimensions. The simulation results show a decrease of the maximum temperature and average junction temperature in the device channel with shorter thickness of the etched AlGaN layer. As the etching length increases, the hot spot temperature near the gate edge decreases, and the new hot spot position gradually shifts to the drain, which results in a more uniform distribution in the channel. Then, Raman spectroscopy is used to measure the channel junction temperature of AlGaN/GaN HEMTs. The results indicate that when the power dissipation is 0.3W and the etching depth is 5, 10, 15nm, the average channel junction temperature of the new etched AlGaN/GaN HEMT is 9.7%, 17.4% and 30.3% lower than that of the ordinary AlGaN/GaN HEMT, respectively. And the highest temperature of the new AlGaN/GaN HEMT also decreases. We can see that etching the barrier layer of AlGaN/GaN HEMT can not only improve the breakdown voltage, but also effectively reduce the channel junction temperature, and establish the relationship between the high power and thermal stability of the AlGaN/GaN HEMTs.

Flow and temperature distributions in a disc type winding-part I: Forced and directed cooling modes

The thermal design of a power transformer determines its lifetime and loading capability, so correct modelling of liquid flow and temperature distributions in the winding is of vital importance. Existing standards and widely used simple calculation models can seriously under/overestimate winding hotspot temperatures under certain circumstances. In this paper, liquid flow and temperature distributions in a physical model representing a disc-type winding in a liquid forced and directed cooling mode are investigated experimentally using Particle Image Velocimetry (PIV) and a temperature measurement system and numerically using Computational Fluid Dynamics (CFD). Dimensional analysis is used to generalise the results into a form useful for design review. The operating conditions investigated include different liquid inlet velocities, inlet temperatures, power losses in individual disc segments, and the effect of alternative liquids. It is shown that hotspot temperature and position within the winding are a non-linear function of liquid inlet velocity, with stagnation and reverse flow demonstrated in both experiments and CFD models. Comparisons of liquid flow and temperature distributions between measurements and CFD simulation results show that 2D CFD results are representative when there are no reverse flows and 3D CFD simulations are needed when reverse flows occur. The results are presented first in dimensional forms to show the effect of each parameter, and then in non-dimensional forms to provide a generalised insight into transformer thermal design.

Distributed temperature detection of transformer windings with externally applied distributed optical fiber.

The local overheating fault of power transformer winding threatens the safe, stable operation of a power system. Conventional winding temperature detection methods are point-type temperature measurements, and the hotspot position and the temperature distribution on the winding cannot be accurately obtained. A distributed temperature measurement method of transformer winding based on Raman scattering was thus proposed, and a fiber composite winding model was designed. The influence of fiber on electric field distribution and temperature transfer in oil was simulated, and the insulation performance of fiber composite winding was studied. The distributed optical fiber temperature measurement platform was built, and a temperature sensing test was conducted. The results show that the distributed temperature detection system of transformer winding has high temperature sensing and positioning performance with temperature measurement error less than 1°C and positioning accuracy error within 1.2 m. This means that this method can satisfy the requirements of transformer winding temperature measurement and provide a new, to the best of our knowledge, approach for online monitoring of high-power equipment.

3D Temperature Distribution Reconstruction in Furnace Based on Acoustic Tomography

3D temperature distribution measurement in a furnace based on acoustic tomography (AT) calculates temperature field through multipath acoustic time-of-flight (TOF) data. In this paper, a new 3D temperature field reconstruction model based on radial basis function approximation with polynomial reproduction (RBF-PR) is proposed for solving the AT inverse problem. In addition, the modified reconstruction method that integrates the advantages of the TSVD and Tikhonov regularization methods is presented to reduce the sensitivity of noise on perturbations with the ill-posed problems and improve the reconstruction quality (RQ). Numerical simulations are implemented to evaluate the effectiveness of the proposed reconstruction method using different 3D temperature distribution models, which include the one-peak symmetry distribution, one-peak asymmetry distribution, and two-peak symmetry distribution. To study the antinoise ability of our method, noises are added to the value of TOF. 3D display of reconstructed temperature fields and reconstruction errors is given. The results indicate that our model can reconstruct the temperature distribution with higher accuracy and better antinoise ability compared with the truncated generalized singular value decomposition (TGSVD). Besides that, the proposed method can determine the hot spot position with higher precision, and the temperature error of the hot spot is lower than the other compared methods.

Prediction of quaternary structure by analysis of hot spot residues in protein-protein interfaces: the case of anthranilate phosphoribosyltransferases.

It is an important goal of computational biology to correctly predict the association state of a protein based on its amino acid sequence and the structures of known homologues. We have pursued this goal on the example of anthranilate phosphoribosyltransferase (AnPRT), an enzyme that is involved in the biosynthesis of the amino acid tryptophan. Firstly, known crystal structures of naturally occurring homodimeric AnPRTs were analyzed using the Protein Interfaces, Surfaces, and Assemblies (PISA) service of the European Bioinformatics Institute (EBI). This led to the identification of two hydrophobic "hot spot" amino acids in the protein-protein interface that were predicted to be essential for self-association. Next, in a comprehensive multiple sequence alignment (MSA), naturally occurring AnPRT variants with hydrophilic or charged amino acids in place of hydrophobic residues in the two hot spot positions were identified. Representative variants were characterized in terms of thermal stability, enzymatic activity, and quaternary structure. We found that AnPRT variants with charged residues in both hot spot positions exist exclusively as monomers in solution. Variants with hydrophilic amino acids in one hot spot position occur in both forms, monomer and dimer. The results of the present study provide a detailed characterization of the determinants of the AnPRT monomer-dimer equilibrium and show that analysis of hot spots in combination with MSAs can be a valuable tool in prediction of protein quaternary structures.

Geometric description of self-interaction potential in symmetric protein complexes

Proteins can self-associate with copies of themselves to form symmetric complexes called homomers. Homomers are widespread in all kingdoms of life and allow for unique geometric and functional properties, as reflected in viral capsids or allostery. Once a protein forms a homomer, however, its internal symmetry can compound the effect of point mutations and trigger uncontrolled self-assembly into high-order structures. We identified mutation hot spots for supramolecular assembly, which are predictable by geometry. Here, we present a dataset of descriptors that characterize these hot spot positions both geometrically and chemically, as well as computer scripts allowing the calculation and visualization of these properties for homomers of choice. Since the biological relevance of homomers is not readily available from their X-ray crystallographic structure, we also provide reliability estimates obtained by methods we recently developed. These data have implications in the study of disease-causing mutations, protein evolution and can be exploited in the design of biomaterials.

Divergent Fine-Scale Recombination Landscapes between a Freshwater and Marine Population of Threespine Stickleback Fish.

Meiotic recombination is a highly conserved process that has profound effects on genome evolution. At a fine-scale, recombination rates can vary drastically across genomes, often localized into small recombination “hotspots” with highly elevated rates, surrounded by regions with little recombination. In most species studied, the location of hotspots within genomes is highly conserved across broad evolutionary timescales. The main exception to this pattern is in mammals, where hotspot location can evolve rapidly among closely related species and even among populations within a species. Hotspot position in mammals is controlled by the gene, Prdm9, whereas in species with conserved hotspots, a functional Prdm9 is typically absent. Due to a limited number of species where recombination rates have been estimated at a fine-scale, it remains unclear whether hotspot conservation is always associated with the absence of a functional Prdm9. Threespine stickleback fish (Gasterosteus aculeatus) are an excellent model to examine the evolution of recombination over short evolutionary timescales. Using a linkage disequilibrium-based approach, we found recombination rates indeed varied at a fine-scale across the genome, with many regions organized into narrow hotspots. Hotspots had highly divergent landscapes between stickleback populations, where only ∼15% of these hotspots were shared. Our results indicate that fine-scale recombination rates may be diverging between closely related populations of threespine stickleback fish. Interestingly, we found only a weak association of a PRDM9 binding motif within hotspots, which suggests that threespine stickleback fish may possess a novel mechanism for targeting recombination hotspots at a fine-scale.

LDJump: Estimating variable recombination rates from population genetic data.

As recombination plays an important role in evolution, its estimation and the identification of hotspot positions is of considerable interest. We propose a novel approach for estimating population recombination rates based on genotyping or sequence data that involves a sequential multiscale change point estimator. Our method also permits demography to be taken into account. It uses several summary statistics within a regression model fitted on suitable scenarios. Our proposed method is accurate, computationally fast, and provides a parsimonious solution by ensuring a type I error control against too many changes in the recombination rate. An application to human genome data suggests a good congruence between our estimated and experimentally identified hotspots. Our method is implemented in the R‐package LDJump, which is freely available at https://github.com/PhHermann/LDJump.

Influences of Rod Diameter and Sand-Mould Strength on Hot Tearing in Mg WE43A Constrained Rod Castings

Under conditions of changing sand-mould strength and rod diameter, hot tearing susceptibility of Mg WE43A alloys was studied using constrained rod casting solidified in a sand mould. Variations of temperature and shrinkage force with time during solidification of Mg WE43A alloy were recorded by means of a thermocouple and a force sensor. Susceptibility to hot tearing at hot section was decreased with increasing rod diameter from 10 mm to 20 mm. The rod with 10 mm diameter fractured, and the rod with 15 mm diameter presented hot tearing. No hot tearing was noted for the rod with 20 mm diameter. For the resin content of 1%, 1.5%, 2%, 2.5% and 3%, the tensile strength of sand mould was measured as 0.12, 0.18, 0.28, 0.17 and 0.15 MPa, respectively. The casting fractured at hot spot position for the sand mould with strength of 0.12 MPa. Hot tearing occurred at hot spot for the sand mould with strength of 0.15 MPa. No hot tearing was found at hot spot for the sand mould with strength of 0.28 MPa. The present research confirms that increasing the sand-mould strength and avoiding the entrapment of sand particles and the formation of gas pores during casting increase the resistance to hot tearing.

"Site and Mutation"-Specific Predictions Enable Minimal Directed Evolution Libraries.

Directed evolution experiments designed to improve the activity of a biocatalyst have increased in sophistication from the early days of completely random mutagenesis. Sequence-based and structure-based methods have been developed to identify "hotspot" positions that when randomized provide a higher frequency of beneficial mutations that improve activity. These focused mutagenesis methods reduce library sizes and therefore reduce screening burden, accelerating the rate of finding improved enzymes. Looking for further acceleration in finding improved enzymes, we investigated whether two existing methods, one sequence-based (Protein GPS) and one structure-based (using Bioluminate and MOE), were sufficiently predictive to provide not just the hotspot position, but also the amino acid substitution that improved activity at that position. By limiting the libraries to variants that contained only specific amino acid substitutions, library sizes were kept to less than 100 variants. For an initial round of ATA-117 R-selective transaminase evolution, we found that the methods used produced libraries where 9% and 18% of the amino acid substitutions chosen were amino acids that improved reaction performance in lysates. The ability to create combinations of mutations as part of the initial design was confounded by the relatively large number of predicted mutations that were inactivating (30% and 45% for the sequence-based and structure-based methods, respectively). Despite this, combining several mutations identified within a given method produced variant lysates 7- and 9-fold more active than the wild-type lysate, highlighting the capability of mutations chosen this way to generate large advances in activity in addition to the reductions in screening.

Heat dissipation characteristics from photovoltaic cells within the partitioned or non-partitioned glazed cavity to the windy environment

Abstract A numerical model, whose computation domain contains the glazed shallow cavity and surrounding wind field, has been proposed to investigate the characteristics of heat dissipation from photovoltaic cells within the glazed cavity to the windy environment. To reflect the actual working characteristics, both the photoelectric conversion characteristic and surface radiation were considered. The temperature distribution of PV cells particularly the hot spot temperature has been displayed to reveal the effect of tilt angle (α) and wind velocity (v). Then a thorough analysis was presented in virtue of visualized results, convection Nusselt number inside the cavity (Nucin) and total heat loss coefficient (Ut). Meanwhile, efforts were also performed to compare the partitioned and non-partitioned cavities. Results show that hot spot position at α = 15° has the obvious diversity due to the variation of wind velocity. Unlike the increased Nucin, total heat loss coefficient of PV cells at v = 5 m/s decreases when the tilt angle shifts from 15° to 30°. Further, it is found that hot spot temperature difference between the three cells shrinks in the partitioned cavity, and the partition indeed contributes to the improvement of PV cells temperature uniformity, which plays a positive role in the protection of PV cell property.

Distinct DICER1 Hotspot Mutations Identify Bilateral Tumors as Separate Events.

DICER1 syndrome1 predisposes to a variety of cancers, including pleuropulmonary blastoma (PPB),2 ovarian Sertoli-Leydig cell tumor (SLCT),3 embryonal rhabdomyosarcoma,4 and kidney tumors.5–8 In DICER1 syndrome, most patients bear a germline null mutation in DICER1, and the tumors uniformly bear a second-hit missense substitution at one of five hotspot positions (1705E, 1709D, 1809G, 1810D, and 1813E). A wide range of clinical phenotypes can be seen in DICER1 syndrome5,9; some patients are asymptomatic, whereas others develop multiple tumors. In the classic cases, patients with a germ-line null mutation develop different somatic hotspot mutations in each tumor.7,10–16 However, in some patients, multiple tumors arise from germline mosaicism of the hotspot mutation, with subsequent somatic loss of the wild-type allele.17–19 Here we report a patient with DICER1 syndrome who developed four tumor types at six anatomic sites over the course of 12 years. These tumors harbor four distinct hotspot mutations, which is one of the highest numbers of distinct somatic DICER1 mutations reported in a single patient. By identifying these mutations, we show that the patient’s bilateral renal tumors and bilateral ovarian SLCTs each constituted a new primary tumor. Because we found no other mutations to explain her particularly severe clinical course, we speculate that her subsequent tumors were a product of the intense chemotherapy and radiation regimens she received.