Evolutionary game study of crowdsourcing open innovation synergy mechanism.

Open innovation crowdsourcing can help enterprises meet the challenges of a rapidly changing environment and improve their innovation performance. This study introduces network externalities as influencing factors of the crowdsourcing open innovation synergy mechanism. This study constructed the game payment matrix of the crowdsourcing open innovation synergy mechanism, and the evolutionary game method obtained the equilibrium solution of the crowdsourcing open innovation synergy mechanism. The impact of changes in the main influencing factors on the issuers' and receivers' willingness to collaborate and innovate was explored through numerical and case studies. The study shows that the higher the synergy benefit and its allocation coefficient need to be within a reasonable range for the willingness to collaborate and innovate to increase; the lower the original cost of both parties, and the higher the cost reduction coefficient under the policy support of the crowdsourcing platform, the higher the willingness to collaborate and innovate; the higher the network externality and the lower the penalty for breach of contract, the higher the desire to collaborate and innovate. The study recommends strengthening non-school education to guide innovation for all, and refining relevant policies to tailor innovation to local conditions. This study provides a new perspective and theoretical guidance for enterprises to build a crowdsourcing open innovation synergy mechanism and is a valuable reference for open innovation management.

The Effect of Defect Charge and Parasitic Surface Conductance on Aluminum Nitride RF Filter Circuit Loss.

When AlN thin films are deposited directly on the high-resistance silicon (HR-Si) substrate, a conductive layer will be formed on the HR-Si surface. This phenomenon is called the parasitic surface conduction (PSC) effect. The presence of the PSC effect will increase the power consumption of electronic components. Therefore, it is necessary to reduce the PSC effect. In prior technology, the polysilicon layer is usually used as the trap-rich layer to reduce the PSC effect. Experiments show that compared to AlN films deposited directly on HR-Si, the AlN substrates with polysilicon introduced on HR-Si have less radio frequency (RF) loss. To verify the effect of polysilicon on RF loss, polysilicon films of three different thicknesses and several different roughnesses were introduced. The results show that the thickness of the polysilicon will affect the RF loss, while the roughness has almost no effect on it. The polysilicon trap-rich layer can reduce the RF loss, which gradually becomes smaller as the polysilicon thickness increases.

Study on the Evolution of the Game of Willingness to Cooperate between Residents and Separation Enterprises in Waste Separation Considering the Convenience of Separation Facilities.

The distributivity and complexity of separation facilities in waste separation cooperation are incorporated into the factors influencing the payoff of waste separation cooperation. The game payment matrix of waste separation cooperation is constructed based on the distributivity and complexity of separation facilities. The equilibrium solution of waste separation cooperation is obtained through the evolutionary game. The influence of different changes in distributivity and complexity of separation facilities on the willingness to cooperate in waste separation is explored through numerical analysis of cases. The study shows that when the distributivity of separation facilities is certain, the lower the complexity of separation facilities, the higher the willingness of residents and enterprises to cooperate; when the complexity of separation facilities is certain, the willingness of residents and enterprises to cooperate rises and then falls with the increase of distributivity of separation facilities; finally, when the distributivity and complexity of separation facilities change at the same time, the willingness of residents and enterprises to cooperate shows different changes with the different changes of two separation facilities convenience factors.

Disease-driven engineering of peptide-targeted DM1 loaded liposomal nanoparticles for enhanced efficacy in treating multiple myeloma by exploring DM1 prodrug chemistry.

Here, we report a CD138 receptor targeting liposomal formulation (TNP[Prodrug-4]) that achieved efficacious tumor growth inhibition in treating multiple myeloma by overcoming the dose limiting severe toxicity issues of a highly potent drug, Mertansine (DM1). Despite the promising potential to treat various cancers, due to poor solubility and pharmacokinetic profile, DM1's translation to the clinic has been unsatisfactory. We hypothesized that the optimal prodrug chemistry would promote efficient loading of the prodrug into targeted nanoparticles and achieve controlled release following endocytosis by the cancer cells, consequently, accomplish the most potent tumor growth inhibition. We evaluated four functional linker chemistries for synthesizing DM1-Prodrug molecules and evaluated their stability and cancer cell toxicity in vitro. It was determined that the phosphodiester moiety, as part of nanoparticle formulations, demonstrated most favorable characteristics with an IC50 of ∼16 nM. Nanoparticle formulations of Prodrug-4 enabled its administration at 8-fold higher dosage of equivalent free drug while remaining below maximum tolerated dose. Importantly, TNP[Prodrug-4] achieved near complete inhibition of tumor growth (∼99% by day 10) compared to control, without displaying noticeable systemic toxicity. TNP[Prodrug-4] promises a formulation that could potentially make DM1 treatment available for wider clinical applications with a long-term goal for better patient outcomes.

Evolutionary Game Analysis of Government and Residents' Participation in Waste Separation Based on Cumulative Prospect Theory.

Government and residents' participation in waste separation is a complex non-cooperative game process, and the evolutionary game can explain the behavior of participating subjects well. Considering that the traditional evolutionary game cannot satisfactorily explain the irrational psychology and risk preference factors of the participating issues, this study combines the prospect theory and evolutionary game, uses the prospect value function to supplement and improve the parameters of the evolutionary game payment matrix, and analyzes the evolutionary stabilization strategy. To verify the theoretical results, simulation experiments and impact analysis were conducted, and meaningful results were obtained: There are two stable evolutionary strategies in the system, namely higher participation benefits for residents and lower participation costs and opportunity costs, and reasonable direct benefit distribution coefficients all help to increase the participation rate of waste separation. This study can provide some scientific suggestions for the government to design and build a waste-separation system.

Distinct dynamics of Vibrio parahaemolyticus populations in two farming models.

AIMS: Despite the recent prosperity of shrimp cultivation in China, very little is known about how different shrimp farming models influence the dynamics of Vibrio parahaemolyticus populations and the antibiotic resistance of this bacterium. METHODS AND RESULTS: To this end, we conducted continuous surveillance of V. parahaemolyticus on four farms over 3 years: two traditional shrimp farms with daily water exchange and two farms operated in the recirculating aquaculture systems (RAS). No antibiotics were used in these farms to exclude the potential impacts of antibiotics on the emergence of antibacterial resistance. Multilocus sequence typing was utilized to characterize the dynamics of V. parahaemolyticus populations. Whole-genome sequencing (WGS) was conducted to determine the representative sequence types (STs) at each farm. Results revealed that the population structure of V. parahaemolyticus remained stable over time in both RAS farms, with only nine and four STs observed at each. In contrast, annual replacement of V. parahaemolyticus populations was observed in traditional farms with 26 and 28 STs identified in rearing water. WGS of 50 isolates divided them into five clusters, of which ST917a isolates harboured a genomic island that disrupted the gene recA. Pair-wised genomic comparison of isolates from the same STs showed that they were genetically related but belonged to different clones associated with geographical distribution. CONCLUSIONS: These results suggested that RAS presented a specific ecological niche by minimizing the water exchanges with the external environment. In contrast, traditional farming might pose a food safety issue by introducing new V. parahaemolyticus populations with antibiotic resistance genes. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results expose the potential food safety issue associated with conventional agriculture and should encourage the development of preventive strategies to reduce the emergence of resistant V. parahaemolyticus populations.

Low Temperature Hydrophilic SiC Wafer Level Direct Bonding for Ultrahigh-Voltage Device Applications.

SiC direct bonding using O2 plasma activation is investigated in this work. SiC substrate and n- SiC epitaxy growth layer are activated with an optimized duration of 60s and power of the oxygen ion beam source at 20 W. After O2 plasma activation, both the SiC substrate and n- SiC epitaxy growth layer present a sufficient hydrophilic surface for bonding. The two 4-inch wafers are prebonded at room temperature followed by an annealing process in an atmospheric N2 ambient for 3 h at 300 °C. The scanning results obtained by C-mode scanning acoustic microscopy (C-SAM) shows a high bonding uniformity. The bonding strength of 1473 mJ/m2 is achieved. The bonding mechanisms are investigated through interface analysis by transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDX). Oxygen is found between the two interfaces, which indicates Si-O and C-O are formed at the bonding interface. However, a C-rich area is also detected at the bonding interface, which reveals the formation of C-C bonds in the activated SiC surface layer. These results show the potential of low cost and efficient surface activation method for SiC direct bonding for ultrahigh-voltage devices applications.

Metagenomics combined with comprehensive validation as a public health risk assessment tool for urban and agricultural run-off.

Early detection of emerging and life-threatening pathogens circulating in complex environments is urgently required to combat infectious diseases. This study proposed a public health risk assessment workflow with three stages, pathogen screening, pathogen genotyping, and risk assessment. In stage one, pathogens were screened with metagenomic sequencing, microfluidic chip, and qPCR. In stage two, pathogens were isolated and genotyped with multi-locus sequence typing (MLST) or conventional PCR. Finally, virulence genes from metagenomic data were assessed for pathogenicity. Two regions (Donggang and Zhanjiang) with potential public health concerns were selected for evaluation, each of which comprised of one urban and one farming wastewater sampling location. Overall, metagenomic sequencing reflected the variation in the relative abundance of medically important bacteria. Over 90 bacterial pathogens were monitored in the metagenomic dataset, of which 56 species harbored virulence genes. In Donggang, a pathogenic Acinetobacter sp. reached high abundances in 2018 and 2020, whereas all pathogenic Vibrio spp. peaked in October 2019. In Zhanjiang, A. baumanni, and other Enterobacteriaceae species were abundantly present in 2019 and 2020, whereas Aeromonas and Vibrio spp. peaked in November-2017. Forty species were subsequently isolated and subtyped by MLST, half of which were prevalent genotypes in clinical data. Additionally, we identified the African Swine Fever Virus (ASFV) in water samples collected in 2017, ahead of the first reported ASFV outbreak in 2018 in China. RNA viruses like Hepatitis A virus (HAV) and Enterovirus 71 (EV71) were also detected, with concentrations peaking in April 2020 and April 2018, respectively. The dynamics of HAV and EV71 were consistent with local epidemic trends. Finally, based on the virulence gene profiles, our study identified the risk level in wastewater of two cities. This workflow illustrates the potential for an early warning of local epidemics, which helps to prioritize the preparedness for specific pathogens locally.

In vivo evaluation of CD38 and CD138 as targets for nanoparticle-based drug delivery in multiple myeloma.

BACKGROUND: Drug-loaded nanoparticles have established their benefits in the fight against multiple myeloma; however, ligand-targeted nanomedicine has yet to successfully translate to the clinic due to insufficient efficacies reported in preclinical studies. METHODS: In this study, liposomal nanoparticles targeting multiple myeloma via CD38 or CD138 receptors are prepared from pre-synthesized, purified constituents to ensure increased consistency over standard synthetic methods. These nanoparticles are then tested both in vitro for uptake to cancer cells and in vivo for accumulation at the tumor site and uptake to tumor cells. Finally, drug-loaded nanoparticles are tested for long-term efficacy in a month-long in vivo study by tracking tumor size and mouse health. RESULTS: The targeted nanoparticles are first optimized in vitro and show increased uptake and cytotoxicity over nontargeted nanoparticles, with CD138-targeting showing superior enhancement over CD38-targeted nanoparticles. However, biodistribution and tumor suppression studies established CD38-targeted nanoparticles to have significantly increased in vivo tumor accumulation, tumor cell uptake, and tumor suppression over both nontargeted and CD138-targeted nanoparticles due to the latter's poor selectivity. CONCLUSION: These results both highlight a promising cancer treatment option in CD38-targeted nanoparticles and emphasize that targeting success in vitro does not necessarily translate to success in vivo.

Stiff stroma increases breast cancer risk by inducing the oncogene ZNF217.

Women with dense breasts have an increased lifetime risk of malignancy that has been attributed to a higher epithelial density. Quantitative proteomics, collagen analysis, and mechanical measurements in normal tissue revealed that stroma in the high-density breast contains more oriented, fibrillar collagen that is stiffer and correlates with higher epithelial cell density. microRNA (miR) profiling of breast tissue identified miR-203 as a matrix stiffness-repressed transcript that is downregulated by collagen density and reduced in the breast epithelium of women with high mammographic density. Culture studies demonstrated that ZNF217 mediates a matrix stiffness- and collagen density-induced increase in Akt activity and mammary epithelial cell proliferation. Manipulation of the epithelium in a mouse model of mammographic density supported a causal relationship between stromal stiffness, reduced miR-203, higher levels of the murine homolog Zfp217, and increased Akt activity and mammary epithelial proliferation. ZNF217 was also increased in the normal breast epithelium of women with high mammographic density, correlated positively with epithelial proliferation and density, and inversely with miR-203. The findings identify ZNF217 as a potential target toward which preexisting therapies, such as the Akt inhibitor triciribine, could be used as a chemopreventive agent to reduce cancer risk in women with high mammographic density.

Aquaporin-7 Regulates the Response to Cellular Stress in Breast Cancer.

The complex yet interrelated connections between cancer metabolism, gene expression, and oncogenic driver genes have the potential to identify novel biomarkers and drug targets with prognostic and therapeutic value. Here we effectively integrated metabolomics and gene expression data from breast cancer mouse models through a novel unbiased correlation-based network analysis. This approach identified 35 metabolite and 34 gene hubs with the most network correlations. These hubs have prognostic value and are likely integral to tumor metabolism and breast cancer. The gene hub Aquaporin-7 (Aqp7), a water and glycerol channel, was identified as a novel regulator of breast cancer. AQP7 was prognostic of overall survival in patients with breast cancer. In mouse breast cancer models, reduced expression of Aqp7 caused reduced primary tumor burden and lung metastasis. Metabolomics and complex lipid profiling of cells and tumors with reduced Aqp7 revealed significantly altered lipid metabolism, glutathione metabolism, and urea/arginine metabolism compared with controls. These data identify AQP7 as a critical regulator of metabolic and signaling responses to environmental cellular stresses in breast cancer, highlighting AQP7 as a potential cancer-specific therapeutic vulnerability. SIGNIFICANCE: Aquaporin-7 is identified as a critical regulator of nutrient availability and signaling that responds to cellular stresses, making it an attractive therapeutic target in breast cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/19/4071/F1.large.jpg.

Engineering peptide-targeted liposomal nanoparticles optimized for improved selectivity for HER2-positive breast cancer cells to achieve enhanced in vivo efficacy.

Here, we report rationally engineered peptide-targeted liposomal doxorubicin nanoparticles that have an enhanced selectivity for HER2-positive breast tumor cells with high purity, reproducibility, and precision in controlling stoichiometry of targeting peptides. To increase HER2-positive tumor cell selective drug delivery, we optimized the two most important design parameters, peptide density and linker length, via systematic evaluations of their effects on both in vitro cellular uptake and in vivo tumor accumulation and cellular uptake. The optimally designed nanoparticles were finally evaluated for their tumor inhibition efficacy using in vivo MMTV-neu transplantation mouse model. In vitro, we demonstrated that ~1% peptide density and EG8 linker were optimal parameters for targeted nanoparticle formulations to enhance HER2-positive cancer cellular uptake while preventing non-selectivity. In vivo results demonstrated that at 0.5% peptide density, enhancement of tumor cell uptake over non-targeted nanoparticles was ~2.7 fold and ~3.4 fold higher for targeted nanoparticles with EG8 and EG18 linker, respectively, while their accumulation levels at tumor tissue were similar to the non-targeted nanoparticles. These results were consistent with in vivo efficacy outcomes that ~90% tumor growth inhibition was achieved by Dox-loaded HER2 receptor targeted nanoparticles, TNPHER2pep, over control while all nanoparticle formulations minimized overall systemic toxicity relative to free Dox. This study highlights the significance of understanding and optimizing the effects of liposomal nanoparticle design parameters for enhancement of tumor selectivity to achieve improved in vivo therapeutic outcomes.

Death effector domain-containing protein induces vulnerability to cell cycle inhibition in triple-negative breast cancer.

Lacking targetable molecular drivers, triple-negative breast cancer (TNBC) is the most clinically challenging subtype of breast cancer. In this study, we reveal that Death Effector Domain-containing DNA-binding protein (DEDD), which is overexpressed in > 60% of TNBCs, drives a mitogen-independent G1/S cell cycle transition through cytoplasm localization. The gain of cytosolic DEDD enhances cyclin D1 expression by interacting with heat shock 71 kDa protein 8 (HSC70). Concurrently, DEDD interacts with Rb family proteins and promotes their proteasome-mediated degradation. DEDD overexpression renders TNBCs vulnerable to cell cycle inhibition. Patients with TNBC have been excluded from CDK 4/6 inhibitor clinical trials due to the perceived high frequency of Rb-loss in TNBCs. Interestingly, our study demonstrated that, irrespective of Rb status, TNBCs with DEDD overexpression exhibit a DEDD-dependent vulnerability to combinatorial treatment with CDK4/6 inhibitor and EGFR inhibitor in vitro and in vivo. Thus, our study provided a rationale for the clinical application of CDK4/6 inhibitor combinatorial regimens for patients with TNBC.

Phosphorylation of TIP60 Suppresses 53BP1 Localization at DNA Damage Sites.

A proper balance between the repair of DNA double-strand breaks (DSBs) by homologous recombination and nonhomologous end joining is critical for maintaining genome integrity and preventing tumorigenesis. This balance is regulated and fine-tuned by a variety of factors, including cell cycle and the chromatin environment. The histone acetyltransferase TIP60 was previously shown to suppress pathological end joining and promote homologous recombination. However, it is unknown how regulatory posttranslational modifications impact TIP60 acetyltransferase activity to influence the outcome of DSB responses. In this study, we report that phosphorylation of TIP60 on serines 90 and 86 is important for limiting the accumulation of the pro-end joining factor 53BP1 at DSBs in S and G2 cell cycle phases. Mutation of these sites disrupts histone acetylation changes in response to DNA damage, BRCA1 localization to DSBs, and poly(ADP-ribose) polymerase (PARP) inhibitor resistance. These findings reveal that phosphorylation directs TIP60-dependent acetylation to promote homologous recombination and maintain genome stability.

Versatility of CoPcS in CoPcS/TiO2 for MB degradation: photosensitization, charge separation and oxygen activation.

In this report, a composite photocatalyst consisting of cobalt phthalocyanine sulfate (CoPcS) and TiO2 was prepared by a facile synthesis. Careful characterizations and measurements indicate a covalent grafting of CoPcS onto TiO2 through Ti-O-S linkages, acquiring an intimate heterojunction between TiO2 and CoPcS. The obtained composite was evaluated for its photocatalytic activity toward the degradation of methyl blue (MB) under visible light irradiation. The evaluation showed a significantly enhanced degradation rate of MB by CoPcS/TiO2. The improved photocatalytic performance of CoPcS/TiO2 was attributed to the photosensitization of TiO2 by CoPcS, charge separation by electron transfer at the interface of the heterojunction formed between CoPcS and TiO2, and oxygen activation via CoPcS. A synergetic mechanism in improving the photocatalytic performance of TiO2 by CoPcS was investigated.

The AKT inhibitor triciribine in combination with paclitaxel has order-specific efficacy against Zfp217-induced breast cancer chemoresistance.

We previously identified the transcription factor ZNF217 (human) / Zfp217 (mouse) as an oncogene and prognostic indicator of reduced survival, increased metastasis, and reduced response to therapy in breast cancer patients. Here we investigated the role of Zfp217 in chemotherapy resistance. Preclinical animal models of Zfp217 overexpression were treated with a combination therapy of the microtubule inhibitor epothilone B, doxorubicin (Adriamycin), and cyclophosphamide (EAC). Tumors overexpressing Zfp217 increased their tumor burden compared to control tumors after treatment and accumulated a mammary gland progenitor cell population (K8+K14+). To overcome this chemoresistance after ZNF217 overexpression, we treated tumors ± Zfp217 overexpression with paclitaxel and triciribine, a nucleoside analog and AKT inhibitor that kills cells that overexpress ZNF217. Treatment order critically impacted the efficacy of the therapy. Combination treatment of triciribine followed by paclitaxel (TCN→PAC) inhibited tumor burden and increased survival in tumors that overexpressed Zfp217, whereas single agent or combination treatment in the reverse order (PAC→TCN) did not improve response. Analysis of these tumors and patient-derived tumor xenograft tumors treated with the same therapies suggested that Zfp217 overexpression in tumors contributes both to decreased microvessel density and vessel maturity, while TCN→PAC tumors overexpressing Zfp217 showed improved vessel maturity.

MERIT40 cooperates with BRCA2 to resolve DNA interstrand cross-links.

MERIT40 is an essential component of the RAP80 ubiquitin recognition complex that targets BRCA1 to DNA damage sites. Although this complex is required for BRCA1 foci formation, its physiologic role in DNA repair has remained enigmatic, as has its relationship to canonical DNA repair mechanisms. Surprisingly, we found that Merit40(-/-) mice displayed marked hypersensitivity to DNA interstrand cross-links (ICLs) but not whole-body irradiation. MERIT40 was rapidly recruited to ICL lesions prior to FANCD2, and Merit40-null cells exhibited delayed ICL unhooking coupled with reduced end resection and homologous recombination at ICL damage. Interestingly, Merit40 mutation exacerbated ICL-induced chromosome instability in the context of concomitant Brca2 deficiency but not in conjunction with Fancd2 mutation. These findings implicate MERIT40 in the earliest stages of ICL repair and define specific functional interactions between RAP80 complex-dependent ubiquitin recognition and the Fanconi anemia (FA)-BRCA ICL repair network.

Analysis of a lung defect in autophagy-deficient mouse strains.

Yeast Atg1 initiates autophagy in response to nutrient limitation. The Ulk gene family encompasses the mammalian orthologs of yeast ATG1. We created mice deficient for both Ulk1 and Ulk2 and found that the mice die within 24 h of birth. When found alive, pups exhibited signs of respiratory distress. Histological sections of lungs of the Ulk1/2 DKO pups showed reduced airspaces with thickened septae. A similar defect was seen in Atg5-deficient pups as both Ulk1/2 DKO and Atg5 KO lungs show numerous glycogen-laden alveolar type II cells by electron microscopy, PAS staining, and increased levels of glycogen in lung homogenates. No abnormalities were noted in expression of genes encoding surfactant proteins but the ability to incorporate exogenous choline into phosphatidylcholine, the major phospholipid component of surfactant, was increased in comparison to controls. Despite this, there was a trend for total phospholipid levels in lung tissue to be lower in Ulk1/2 DKO and Atg5 KO compared with controls. Autophagy was abundant in lung epithelial cells from wild-type mice, but lacking in Atg5 KO and Ulk1/2 DKO mice at P1. Analysis of the autophagy signaling pathway showed the existence of a negative feedback loop between the ULK1 and 2 and MTORC1 and 2, in lung tissue. In the absence of autophagy, alveolar epithelial cells are unable to mobilize internal glycogen stores independently of surfactant maturation. Together, the data suggested that autophagy plays a vital role in lung structural maturation in support of perinatal adaptation to air breathing.

A BRISC-SHMT complex deubiquitinates IFNAR1 and regulates interferon responses.

Lysine63-linked ubiquitin (K63-Ub) chains represent a particular ubiquitin topology that mediates proteasome-independent signaling events. The deubiquitinating enzyme (DUB) BRCC36 segregates into distinct nuclear and cytoplasmic complexes that are specific for K63-Ub hydrolysis. RAP80 targets the five-member nuclear BRCC36 complex to K63-Ub chains at DNA double-strand breaks. The alternative four-member BRCC36 containing complex (BRISC) lacks a known targeting moiety. Here, we identify serine hydroxymethyltransferase (SHMT) as a previously unappreciated component that fulfills this function. SHMT directs BRISC activity at K63-Ub chains conjugated to the type 1 interferon (IFN) receptor chain 1 (IFNAR1). BRISC-SHMT2 complexes localize to and deubiquitinate actively engaged IFNAR1, thus limiting its K63-Ub-mediated internalization and lysosomal degradation. BRISC-deficient cells and mice exhibit attenuated responses to IFN and are protected from IFN-associated immunopathology. These studies reveal a mechanism of DUB regulation and suggest a therapeutic use of BRISC inhibitors for treating pathophysiological processes driven by elevated IFN responses.

The transcription factors T-bet and Eomes control key checkpoints of natural killer cell maturation.

Natural killer (NK) cells play critical roles defending against tumors and pathogens. We show that mice lacking both transcription factors Eomesodermin (Eomes) and T-bet failed to develop NK cells. Developmental stability of immature NK cells constitutively expressing the death ligand TRAIL depended on T-bet. Conversely, maturation characterized by loss of constitutive TRAIL expression and induction of Ly49 receptor diversity and integrin CD49b (DX5(+)) required Eomes. Mature NK cells from which Eomes was deleted reverted to phenotypic immaturity if T-bet was present or downregulated NK lineage antigens if T-bet was absent, despite retaining expression of Ly49 receptors. Fetal and adult hepatic hematopoiesis restricted Eomes expression and limited NK development to the T-bet-dependent, immature stage, whereas medullary hematopoiesis permitted Eomes-dependent NK maturation in adult mice. These findings reveal two sequential, genetically separable checkpoints of NK cell maturation, the progression of which is metered largely by the anatomic localization of hematopoiesis.