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Lithium‐rich transition metal oxides with a cation‐disordered rocksalt structure (disordered rocksalt oxides or DRX) are promising candidates for sustainable, next‐generation Li‐ion cathodes due to their high energy densities and compositional flexibility, enabling Co‐ and Ni‐free battery chemistries. However, current methods to synthesize DRX compounds require either high temperature (≈1000 °C) sintering for several hours, or high energy ball milling for several days in an inert atmosphere. Both methods are time‐ and energy‐intensive, limiting the scale up of DRX production. The present study reports the rapid synthesis of various DRX compositions in ambient air via a microwave‐assisted solid‐state technique resulting in reaction times as short as 5 min, which are more than two orders of magnitude faster than current synthesis methods. The DRX compounds synthesized via microwave are phase‐pure and have a similar short‐ and long‐range structure as compared to DRX materials synthesized via a standard solid‐state route, resulting in nearly identical electrochemical performance. In some cases, microwave heating allows for better particle size and morphology control. Overall, the rapid and energy‐efficient microwave technique provides a more sustainable route to produce DRX materials, further incentivizes the development of next‐generation DRX cathodes, and is key to accelerating their optimization via high‐throughput studies.

Weberite-type sodium transition metal fluorides (Na2M2+M'3+F7) have emerged as potential high-performance sodium intercalation cathodes, with predicted energy densities in the 600-800 W h/kg range and fast Na-ion transport. One of the few weberites that have been electrochemically tested is Na2Fe2F7, yet inconsistencies in its reported structure and electrochemical properties have hampered the establishment of clear structure-property relationships. In this study, we reconcile structural characteristics and electrochemical behavior using a combined experimental-computational approach. First-principles calculations reveal the inherent metastability of weberite-type phases, the close energetics of several Na2Fe2F7 weberite polymorphs, and their predicted (de)intercalation behavior. We find that the as-prepared Na2Fe2F7 samples inevitably contain a mixture of polymorphs, with local probes such as solid-state nuclear magnetic resonance (NMR) and Mössbauer spectroscopy providing unique insights into the distribution of Na and Fe local environments. Polymorphic Na2Fe2F7 exhibits a respectable initial capacity yet steady capacity fade, a consequence of the transformation of the Na2Fe2F7 weberite phases to the more stable perovskite-type NaFeF3 phase upon cycling, as revealed by ex situ synchrotron X-ray diffraction and solid-state NMR. Overall, these findings highlight the need for greater control over weberite polymorphism and phase stability through compositional tuning and synthesis optimization.

Penelitian ini bertujuan untuk menganalisa faktor-faktor yang dapat mempengaruhi loyalitas karyawan di perusahaam sparepart alat berat yang ada di Kota Batam. Adapun variabel indpenden yang digunakan yaitu compensation, work-life balance, career development, workload dan work environment. Sementara variabel dependen yang digunakan yaitu employee loyalty dengan job satisfaction sebagai variabel mediasi. Jenis penelitian yang digunakan yaitu dengan menggunakan pendekatan kuantitatif dengan menggunakan data primer yang didapatkan melalui penyebaran kuesioner melalui google form. Teknik purposive sampling merupakan metode yang digunakan pada penelitian ini dengan jumlah responden sebanyak 311 orang dimana merupakan karyawan di 9 perusahaan sparepart alat bereat dan lori yang ada di Kota Batam. Data kemudian dianalisis dengan menggunakan metode Structural Equation Modeling (SEM) PLS dengan aplikasi SmartPLS versi 3.0 untuk dapat menjelaskan keterkaitan yang terjadi diantara variable yang berhubungan. Hasil penelitian menunjukkan bahwa compensation, workload dan work environment memiliki pengaruh yang signifikan terhadap employee loyalty. Work-life balance dan career development tidak memiliki pengaruh yang signifikan terhadap employee loyalty. Job satisfaction diketahui mampu memediasi hubungan antara compensation terhadap employee loyalty dan work environment terhadap employee loyalty. Job satisfaction juga memiliki hubungan dan pengaruh yang signifikan terhadap employee loyalty.

Penelitian ini bertujuan untuk menganalisa faktor-faktor yang dapat mempengaruhi loyalitas karyawan di perusahaam sparepart alat berat yang ada di Kota Batam. Adapun variabel indpenden yang digunakan yaitu compensation, work-life balance, career development, workload dan work environment. Sementara variabel dependen yang digunakan yaitu employee loyalty dengan job satisfaction sebagai variabel mediasi. Jenis penelitian yang digunakan yaitu dengan menggunakan pendekatan kuantitatif dengan menggunakan data primer yang didapatkan melalui penyebaran kuesioner melalui google form. Teknik purposive sampling merupakan metode yang digunakan pada penelitian ini dengan jumlah responden sebanyak 311 orang dimana merupakan karyawan di 9 perusahaan sparepart alat bereat dan lori yang ada di Kota Batam. Data kemudian dianalisis dengan menggunakan metode Structural Equation Modeling (SEM) PLS dengan aplikasi SmartPLS versi 3.0 untuk dapat menjelaskan keterkaitan yang terjadi diantara variable yang berhubungan. Hasil penelitian menunjukkan bahwa compensation, workload dan work environment memiliki pengaruh yang signifikan terhadap employee loyalty. Work-life balance dan career development tidak memiliki pengaruh yang signifikan terhadap employee loyalty. Job satisfaction diketahui mampu memediasi hubungan antara compensation terhadap employee loyalty dan work environment terhadap employee loyalty. Job satisfaction juga memiliki hubungan dan pengaruh yang signifikan terhadap employee loyalty.

Filamentous fungi, such as Neurospora crassa, are very efficient in deconstructing plant biomass by the secretion of an arsenal of plant cell wall-degrading enzymes, by remodeling metabolism to accommodate production of secreted enzymes, and by enabling transport and intracellular utilization of plant biomass components. Although a number of enzymes and transcriptional regulators involved in plant biomass utilization have been identified, how filamentous fungi sense and integrate nutritional information encoded in the plant cell wall into a regulatory hierarchy for optimal utilization of complex carbon sources is not understood. Here, we performed transcriptional profiling of N. crassa on 40 different carbon sources, including plant biomass, to provide data on how fungi sense simple to complex carbohydrates. From these data, we identified regulatory factors in N. crassa and characterized one (PDR-2) associated with pectin utilization and one with pectin/hemicellulose utilization (ARA-1). Using in vitro DNA affinity purification sequencing (DAP-seq), we identified direct targets of transcription factors involved in regulating genes encoding plant cell wall-degrading enzymes. In particular, our data clarified the role of the transcription factor VIB-1 in the regulation of genes encoding plant cell wall-degrading enzymes and nutrient scavenging and revealed a major role of the carbon catabolite repressor CRE-1 in regulating the expression of major facilitator transporter genes. These data contribute to a more complete understanding of cross talk between transcription factors and their target genes, which are involved in regulating nutrient sensing and plant biomass utilization on a global level.

Filamentous fungi, such as Neurospora crassa, are very efficient in deconstructing plant biomass by the secretion of an arsenal of plant cell wall-degrading enzymes, by remodeling metabolism to accommodate production of secreted enzymes, and by enabling transport and intracellular utilization of plant biomass components. Although a number of enzymes and transcriptional regulators involved in plant biomass utilization have been identified, how filamentous fungi sense and integrate nutritional information encoded in the plant cell wall into a regulatory hierarchy for optimal utilization of complex carbon sources is not understood. Here, we performed transcriptional profiling of N. crassa on 40 different carbon sources, including plant biomass, to provide data on how fungi sense simple to complex carbohydrates. From these data, we identified regulatory factors in N. crassa and characterized one (PDR-2) associated with pectin utilization and one with pectin/hemicellulose utilization (ARA-1). Using in vitro DNA affinity purification sequencing (DAP-seq), we identified direct targets of transcription factors involved in regulating genes encoding plant cell wall-degrading enzymes. In particular, our data clarified the role of the transcription factor VIB-1 in the regulation of genes encoding plant cell wall-degrading enzymes and nutrient scavenging and revealed a major role of the carbon catabolite repressor CRE-1 in regulating the expression of major facilitator transporter genes. These data contribute to a more complete understanding of cross talk between transcription factors and their target genes, which are involved in regulating nutrient sensing and plant biomass utilization on a global level.

Since its initial documentation in 1981, the acquired immunodeficiency syndrome (AIDS) epidemic has impacted more than 70 million people. As the causative agent of AIDS, human immunodeficiency virus (HIV) is a retrovirus that integrates a copy of its viral genome (termed provirus) into the genome of target cells known as CD4+ T cells. With antiretroviral therapy (ART), persons living with HIV (PLWH) can suppress viral replication and prevent progression to AIDS. However, ART is usually a lifelong requirement as interruption can lead to viral rebound via transcription at the integrated viral genome. During early HIV infection, some infected cells can enter a quiescent state that results in a state of infection known as latency. Persistent infection and reversible latency are characteristics of the HIV reservoir. As a result, there is no universally accessible cure for HIV. A functional cure of HIV – which is to suppress viral replication without ART – is a primary goal of HIV cure research. Such a strategy requires an in-depth understanding of the HIV reservoir but many CD4+ T cells, each with different function and phenotype, contribute to the collective HIV reservoir that is located throughout different tissue environments. To address this challenge, this study deciphers the reservoir through the lens of tissue versus blood compartmentalization at a subset resolution to address the impact of residency and circulation as well as with a novel single-cell approach to enable a comprehensive analysis of the HIV reservoir during ART. While the characteristics of integration were generally the same across compartments, there was a lack of overlap between tissues and blood to suggest the existence of sub-reservoirs where there may be unique reservoirs based on residency status. Furthermore, the single-cell profiling revealed extensive heterogeneity between and within individuals with some infected cells displaying a poised signature for reactivation. These results provided

This study introduces a flexible, adhesive-integrated electrode array that was developed to enable non-invasive monitoring of cervical nerve activity. The device uses silver-silver chloride as the electrode material of choice and combines it with an electrode array consisting of a customized biopotential data acquisition unit and integrated graphical user interface (GUI) for visualization of real-time monitoring. Preliminary testing demonstrated this electrode design can achieve a high signal to noise ratio during cervical neural recordings. To demonstrate the capability of the surface electrodes to detect changes in cervical neuronal activity, the cold-pressor test (CPT) and a timed respiratory challenge were employed as stressors to the autonomic nervous system. This sensor system recording, a new technique, was termed Cervical Electroneurography (CEN). By applying a custom spike sorting algorithm to the electrode measurements, neural activity was classified in two ways: (1) pre-to-post CPT, and (2) during a timed respiratory challenge. Unique to this work: (1) rostral to caudal channel position-specific (cephalad to caudal) firing patterns and (2) cross challenge biotype-specific change in average CEN firing, were observed with both CPT and the timed respiratory challenge. Future work is planned to develop an ambulatory CEN recording device that could provide immediate notification of autonomic nervous system activity changes that might indicate autonomic dysregulation in healthy subjects and clinical disease states.

Glaucoma, the leading cause of irreversible blindness, affects more than 70 million people worldwide. Lowering intraocular pressure, the most common method of delaying both the development and progression of glaucoma, is typically done with daily topical administration of therapeutic eye drops, which has notoriously high non-adherence rates ranging from 30% to 80%. The advent of smart phone enabled technologies creates an opportunity to address the non-adherence problem. However, previous eyedrop electronic monitoring solutions had awkward medication bottle adjuncts and crude software design for monitoring the administration of a drop which adversely affected their ability to foster significant, sustainable improvements in adherence. Here we present our prototype, “smart drop” bottle, that is capable to detect each eyedrop medication administered while maintaining the shape or size of the eyedrop bottle. This is achieved by developing a smart-phone application that interact with small rigid-flex electronics beneath the bottle and bottle label respectively. We have shown we can achieve wireless communication close to 100 feet with 0% of false positive results thus providing a potential solution for adherence monitoring of glaucoma patients.

Glaucoma, the leading cause of irreversible blindness, affects more than 70 million people worldwide. Lowering intraocular pressure, the most common method of delaying both the development and progression of glaucoma, is typically done with daily topical administration of therapeutic eye drops, which has notoriously high non-adherence rates ranging from 30% to 80%. The advent of smart phone enabled technologies creates an opportunity to address the non-adherence problem. However, previous eyedrop electronic monitoring solutions had awkward medication bottle adjuncts and crude software design for monitoring the administration of a drop which adversely affected their ability to foster significant, sustainable improvements in adherence. Here we present our prototype, “smart drop” bottle, that is capable to detect each eyedrop medication administered while maintaining the shape or size of the eyedrop bottle. This is achieved by developing a smart-phone application that interact with small rigid-flex electronics beneath the bottle and bottle label respectively. We have shown we can achieve wireless communication close to 100 feet with 0% of false positive results thus providing a potential solution for adherence monitoring of glaucoma patients.

Sensing available nutrients and efficiently utilizing them is a challenge common to all organisms. The model filamentous fungus Neurospora crassa is capable of utilizing a variety of inorganic and organic nitrogen sources. Nitrogen utilization in N. crassa is regulated by a network of pathway-specific transcription factors that activate genes necessary to utilize specific nitrogen sources in combination with nitrogen catabolite repression regulatory proteins. We identified an uncharacterized pathway-specific transcription factor, amn-1, that is required for utilization of the nonpreferred nitrogen sources proline, branched-chain amino acids, and aromatic amino acids. AMN-1 also plays a role in regulating genes involved in responding to the simple sugar mannose, suggesting an integration of nitrogen and carbon metabolism. The utilization of nonpreferred nitrogen sources, which require metabolic processing before being used as a nitrogen source, is also regulated by the nitrogen catabolite regulator NIT-2. Using RNA sequencing combined with DNA affinity purification sequencing, we performed a survey of the role of NIT-2 and the pathway-specific transcription factors NIT-4 and AMN-1 in directly regulating genes involved in nitrogen utilization. Although previous studies suggested promoter binding by both a pathway-specific transcription factor and NIT-2 may be necessary for activation of nitrogen-responsive genes, our data show that pathway-specific transcription factors regulate genes involved in the catabolism of specific nitrogen sources, while NIT-2 regulates genes involved in utilization of all nonpreferred nitrogen sources, such as nitrogen transporters. Together, these transcription factors form a nutrient sensing network that allows N. crassa cells to regulate nitrogen utilization.

Accurate nutrient sensing is important for rapid fungal growth and exploitation of available resources. Sulfur is an important nutrient source found in a number of biological macromolecules, including proteins and lipids. The model filamentous fungus Neurospora crassa is capable of utilizing sulfur found in a variety of sources from amino acids to sulfate. During sulfur starvation, the transcription factor CYS-3 is responsible for upregulation of genes involved in sulfur uptake and assimilation. Using a combination of RNA sequencing and DNA affinity purification sequencing, we performed a global survey of the N. crassa sulfur starvation response and the role of CYS-3 in regulating sulfur-responsive genes. The CYS-3 transcription factor bound the promoters and regulated genes involved in sulfur metabolism. Additionally, CYS-3 directly activated the expression of a number of uncharacterized transporter genes, suggesting that regulation of sulfur import is an important aspect of regulation by CYS-3. CYS-3 also directly regulated the expression of genes involved in mitochondrial electron transfer. During sulfur starvation, genes involved in nitrogen metabolism, such as amino acid and nucleic acid metabolic pathways, along with genes encoding proteases and nucleases that are necessary for scavenging nitrogen, were activated. Sulfur starvation also caused changes in the expression of genes involved in carbohydrate metabolism, such as those encoding glycosyl hydrolases. Thus, our data suggest a connection between sulfur metabolism and other aspects of cellular metabolism. IMPORTANCE Identification of nutrients present in the environment is a challenge common to all organisms. Sulfur is an important nutrient source found in proteins, lipids, and electron carriers that are required for the survival of filamentous fungi such as Neurospora crassa. Here, we transcriptionally profiled the response of N. crassa to characterize the global response to sulfur starvation. We also use

Sensing available nutrients and efficiently utilizing them is a challenge common to all organisms. The model filamentous fungus Neurospora crassa is capable of utilizing a variety of inorganic and organic nitrogen sources. Nitrogen utilization in N. crassa is regulated by a network of pathway-specific transcription factors that activate genes necessary to utilize specific nitrogen sources in combination with nitrogen catabolite repression regulatory proteins. We identified an uncharacterized pathway-specific transcription factor, amn-1, that is required for utilization of the nonpreferred nitrogen sources proline, branched-chain amino acids, and aromatic amino acids. AMN-1 also plays a role in regulating genes involved in responding to the simple sugar mannose, suggesting an integration of nitrogen and carbon metabolism. The utilization of nonpreferred nitrogen sources, which require metabolic processing before being used as a nitrogen source, is also regulated by the nitrogen catabolite regulator NIT-2. Using RNA sequencing combined with DNA affinity purification sequencing, we performed a survey of the role of NIT-2 and the pathway-specific transcription factors NIT-4 and AMN-1 in directly regulating genes involved in nitrogen utilization. Although previous studies suggested promoter binding by both a pathway-specific transcription factor and NIT-2 may be necessary for activation of nitrogen-responsive genes, our data show that pathway-specific transcription factors regulate genes involved in the catabolism of specific nitrogen sources, while NIT-2 regulates genes involved in utilization of all nonpreferred nitrogen sources, such as nitrogen transporters. Together, these transcription factors form a nutrient sensing network that allows N. crassa cells to regulate nitrogen utilization.

Accurate nutrient sensing is important for rapid fungal growth and exploitation of available resources. Sulfur is an important nutrient source found in a number of biological macromolecules, including proteins and lipids. The model filamentous fungus Neurospora crassa is capable of utilizing sulfur found in a variety of sources from amino acids to sulfate. During sulfur starvation, the transcription factor CYS-3 is responsible for upregulation of genes involved in sulfur uptake and assimilation. Using a combination of RNA sequencing and DNA affinity purification sequencing, we performed a global survey of the N. crassa sulfur starvation response and the role of CYS-3 in regulating sulfur-responsive genes. The CYS-3 transcription factor bound the promoters and regulated genes involved in sulfur metabolism. Additionally, CYS-3 directly activated the expression of a number of uncharacterized transporter genes, suggesting that regulation of sulfur import is an important aspect of regulation by CYS-3. CYS-3 also directly regulated the expression of genes involved in mitochondrial electron transfer. During sulfur starvation, genes involved in nitrogen metabolism, such as amino acid and nucleic acid metabolic pathways, along with genes encoding proteases and nucleases that are necessary for scavenging nitrogen, were activated. Sulfur starvation also caused changes in the expression of genes involved in carbohydrate metabolism, such as those encoding glycosyl hydrolases. Thus, our data suggest a connection between sulfur metabolism and other aspects of cellular metabolism. IMPORTANCE Identification of nutrients present in the environment is a challenge common to all organisms. Sulfur is an important nutrient source found in proteins, lipids, and electron carriers that are required for the survival of filamentous fungi such as Neurospora crassa. Here, we transcriptionally profiled the response of N. crassa to characterize the global response to sulfur starvation. We also use

CD8(+) T cell exhaustion is a hallmark of many cancers and chronic infections. In mice, T cell factor 1 (TCF-1) maintains exhausted CD8(+) T cell responses, whereas thymocyte selection-associated HMG box (TOX) is required for the epigenetic remodeling and survival of exhausted CD8(+) T cells. However, it has remained unclear to what extent these transcription factors play analogous roles in humans. In this study, we mapped the expression of TOX and TCF-1 as a function of differentiation and specificity in the human CD8(+) T cell landscape. Here, we demonstrate that circulating TOX+ CD8(+) T cells exist in most humans, but that TOX is not exclusively associated with exhaustion. Effector memory CD8(+) T cells generally expressed TOX, whereas naive and early-differentiated memory CD8(+) T cells generally expressed TCF-1. Cytolytic gene and protein expression signatures were also defined by the expression of TOX. In the context of a relentless immune challenge, exhausted HIV-specific CD8(+) T cells commonly expressed TOX, often in clusters with various activation markers and inhibitory receptors, and expressed less TCF-1. However, polyfunctional memory CD8(+) T cells specific for cytomegalovirus (CMV) or Epstein-Barr virus (EBV) also expressed TOX, either with or without TCF-1. A similar phenotype was observed among HIV-specific CD8(+) T cells from individuals who maintained exceptional immune control of viral replication. Collectively, these data demonstrate that TOX is expressed by most circulating effector memory CD8(+) T cell subsets and not exclusively linked to exhaustion., QC 20200728

Glaucoma, the leading cause of irreversible blindness, affects >70 million people worldwide. Lowering intraocular pressure via topical administration of eye drops is the most common first-line therapy for glaucoma. This treatment paradigm has notoriously high non-adherence rates: ranging from 30% to 80%. The advent of smart phone enabled technologies creates promise for improving eyedrop adherence. However, previous eyedrop electronic monitoring solutions had awkward medication bottle adjuncts and crude software for monitoring the administration of a drop that adversely affected their ability to foster sustainable improvements in adherence. The current work begins to address this unmet need for wireless technology by creating a "smart drop" bottle. This medication bottle is instrumented with sensing electronics that enable detection of each eyedrop administered while maintaining the shape and size of the bottle. This is achieved by a thin electronic force sensor wrapped around the bottle and underneath the label, interfaced with a thin electronic circuit underneath the bottle that allows for detection and wireless transmission to a smart-phone application. We demonstrate 100% success rate of wireless communication over 75 feet with <1% false positive and false negative rates of single drop deliveries, thus providing a viable solution for eyedrop monitoring for glaucoma patients.

Glaucoma, the leading cause of irreversible blindness, affects >70 million people worldwide. Lowering intraocular pressure via topical administration of eye drops is the most common first-line therapy for glaucoma. This treatment paradigm has notoriously high non-adherence rates: ranging from 30% to 80%. The advent of smart phone enabled technologies creates promise for improving eyedrop adherence. However, previous eyedrop electronic monitoring solutions had awkward medication bottle adjuncts and crude software for monitoring the administration of a drop that adversely affected their ability to foster sustainable improvements in adherence. The current work begins to address this unmet need for wireless technology by creating a "smart drop" bottle. This medication bottle is instrumented with sensing electronics that enable detection of each eyedrop administered while maintaining the shape and size of the bottle. This is achieved by a thin electronic force sensor wrapped around the bottle and underneath the label, interfaced with a thin electronic circuit underneath the bottle that allows for detection and wireless transmission to a smart-phone application. We demonstrate 100% success rate of wireless communication over 75 feet with <1% false positive and false negative rates of single drop deliveries, thus providing a viable solution for eyedrop monitoring for glaucoma patients.

Lessons in Plant Cell Wall Degradation by the Ascomycete Model Fungus, Neurospora crassabyVincent W. WuDoctor of Philosophy in MicrobiologyUniversity of California, BerkeleyProfessor N. Louise Glass, Chair In order to mitigate the negative environmental effects of anthropogenic global climate change, renewable resources must be developed to replace fossil fuels. One potential fossil fuel replacement platform utilizes plant biomass from agricultural waste as a feedstock to generate fuels and high value products. Plant cell wall polysaccharides including cellulose, hemicellulose and pectin are broken down into fermentable sugars that can be converted to compounds of interest by microorganisms. Filamentous fungi are capable of both depolymerizing polysaccharides and converting fermentable sugars to high value products. However, further engineering is required to generate strains of filamentous fungi that can efficiently secrete both polysaccharide-degrading enzymes and/or synthesized compounds of interest. This engineering is only possible through a better understanding of the metabolic biology of these organisms. This dissertation describes the mechanisms by which the model filamentous fungus Neurospora crassa regulates and produces the enzymes required for depolymerization of plant cell wall polysaccharides. We investigated pyroglutamate, a unique post-translational modification, and have determined the mechanism of catalysis, as well as its role in the secretion of plant cell wall degrading enzymes. Pyroglutamate modification is catalyzed by two glutaminyl cyclases localized in the endoplasmic reticulum. Secreted proteins with either an N-terminal glutamine or glutamate are likely to have this terminal amino acid converted to a pyroglutamate. Pyroglutamate provides proteins with protection from native N. crassa proteases. The modification also provides structural stability for enzymes within glycosyl hydrolase family 7, including CBH-1 and other important cellulases

Lessons in Plant Cell Wall Degradation by the Ascomycete Model Fungus, Neurospora crassabyVincent W. WuDoctor of Philosophy in MicrobiologyUniversity of California, BerkeleyProfessor N. Louise Glass, Chair In order to mitigate the negative environmental effects of anthropogenic global climate change, renewable resources must be developed to replace fossil fuels. One potential fossil fuel replacement platform utilizes plant biomass from agricultural waste as a feedstock to generate fuels and high value products. Plant cell wall polysaccharides including cellulose, hemicellulose and pectin are broken down into fermentable sugars that can be converted to compounds of interest by microorganisms. Filamentous fungi are capable of both depolymerizing polysaccharides and converting fermentable sugars to high value products. However, further engineering is required to generate strains of filamentous fungi that can efficiently secrete both polysaccharide-degrading enzymes and/or synthesized compounds of interest. This engineering is only possible through a better understanding of the metabolic biology of these organisms. This dissertation describes the mechanisms by which the model filamentous fungus Neurospora crassa regulates and produces the enzymes required for depolymerization of plant cell wall polysaccharides. We investigated pyroglutamate, a unique post-translational modification, and have determined the mechanism of catalysis, as well as its role in the secretion of plant cell wall degrading enzymes. Pyroglutamate modification is catalyzed by two glutaminyl cyclases localized in the endoplasmic reticulum. Secreted proteins with either an N-terminal glutamine or glutamate are likely to have this terminal amino acid converted to a pyroglutamate. Pyroglutamate provides proteins with protection from native N. crassa proteases. The modification also provides structural stability for enzymes within glycosyl hydrolase family 7, including CBH-1 and other important cellulases

The functional properties of circulating CD8+ T cells have been associated with immune control of HIV. However, viral replication occurs predominantly in secondary lymphoid tissues, such as lymph nodes (LNs). We used an integrated single-cell approach to characterize effective HIV-specific CD8+ T cell responses in the LNs of elite controllers (ECs), defined as individuals who suppress viral replication in the absence of antiretroviral therapy (ART). Higher frequencies of total memory and follicle-homing HIV-specific CD8+ T cells were detected in the LNs of ECs compared with the LNs of chronic progressors (CPs) who were not receiving ART. Moreover, HIV-specific CD8+ T cells potently suppressed viral replication without demonstrable cytolytic activity in the LNs of ECs, which harbored substantially lower amounts of CD4+ T cell-associated HIV DNA and RNA compared with the LNs of CPs. Single-cell RNA sequencing analyses further revealed a distinct transcriptional signature among HIV-specific CD8+ T cells from the LNs of ECs, typified by the down-regulation of inhibitory receptors and cytolytic molecules and the up-regulation of multiple cytokines, predicted secreted factors, and components of the protein translation machinery. Collectively, these results provide a mechanistic framework to expedite the identification of novel antiviral factors, highlighting a potential role for the localized deployment of noncytolytic functions as a determinant of immune efficacy against HIV.

Alloying different semiconductors is a powerful approach to tuning the optical and electronic properties of semiconductor materials. In halide perovskites (ABX3), alloys with different anions have been widely studied, and great band gap tunability in the visible range has been achieved. However, perovskite alloys with different cations at the "B" site are less understood due to the synthetic challenges. Herein, we first have developed the synthesis of single-crystalline CsPb xSn1- xI3 nanowires (NWs). The electronic band gaps of CsPb xSn1- xI3 NWs can be tuned from 1.3 to 1.78 eV by varying the Pb/Sn ratio, which leads to the tunable photoluminescence (PL) in the near-infrared range. More importantly, we found that the electrical conductivity increases as more Sn2+ is alloyed with Pb2+, possibly due to the increase of charge carrier concentration when more Sn2+ is introduced. The wide tunability of the optical and electronic properties makes CsPb xSn1- xI3 alloy NWs promising candidates for future optoelectronic device applications.

Alloying different semiconductors is a powerful approach to tuning the optical and electronic properties of semiconductor materials. In halide perovskites (ABX3), alloys with different anions have been widely studied, and great band gap tunability in the visible range has been achieved. However, perovskite alloys with different cations at the "B" site are less understood due to the synthetic challenges. Herein, we first have developed the synthesis of single-crystalline CsPb xSn1- xI3 nanowires (NWs). The electronic band gaps of CsPb xSn1- xI3 NWs can be tuned from 1.3 to 1.78 eV by varying the Pb/Sn ratio, which leads to the tunable photoluminescence (PL) in the near-infrared range. More importantly, we found that the electrical conductivity increases as more Sn2+ is alloyed with Pb2+, possibly due to the increase of charge carrier concentration when more Sn2+ is introduced. The wide tunability of the optical and electronic properties makes CsPb xSn1- xI3 alloy NWs promising candidates for future optoelectronic device applications.

Sterol regulatory element binding proteins (SREBPs) are conserved from yeast to mammalian cells and function in the regulation of sterol homeostasis. In fungi, the SREBP pathway has been implicated in the adaptation to hypoxia and in virulence. In Neurospora crassa and Trichoderma reesei, the SREBP pathway also negatively regulates protein secretion under lignocellulolytic conditions. Here we utilized global transcriptional profiling combined with genetic and physiological analyses to address the regulatory link between the SREBP pathway and protein secretion in N. crassa Our results demonstrated that the function of the SREBP pathway in ergosterol biosynthesis and adaptation to hypoxia was conserved in N. crassa Under lignocellulolytic conditions, the SREBP pathway was highly activated, resulting in the reduced expression of lytic polysaccharide monooxygenases, which require molecular oxygen for catalytic activity. Additionally, activation of the SREBP pathway under lignocellulolytic conditions repressed a set of genes predicted to be involved in the endoplasmic reticulum stress response. Here we show that the inability of a hac-1 mutant, which bears a deletion of the major regulator of the unfolded protein response (UPR), to efficiently produce cellulases and utilize cellulose was suppressed by mutations in the SREBP pathway. The analyses presented here demonstrated new SREBP pathway functions, including linkages to the UPR, and provide new clues for genetic engineering of filamentous fungi to improve their production of extracellular proteins.IMPORTANCE The role of SREBP transcription factors in the regulation of sterol biosynthesis is conserved from humans to yeast. In filamentous fungi, this pathway regulates the secretion of lignocellulolytic enzymes during plant biomass deconstruction. Here we show that the SREBP pathway in Neurospora crassa regulates the production of specific cellulases, lytic polysaccharide monooxygenases that utilize molecular oxygen. Via

Sterol regulatory element binding proteins (SREBPs) are conserved from yeast to mammalian cells and function in the regulation of sterol homeostasis. In fungi, the SREBP pathway has been implicated in the adaptation to hypoxia and in virulence. In Neurospora crassa and Trichoderma reesei, the SREBP pathway also negatively regulates protein secretion under lignocellulolytic conditions. Here we utilized global transcriptional profiling combined with genetic and physiological analyses to address the regulatory link between the SREBP pathway and protein secretion in N. crassa Our results demonstrated that the function of the SREBP pathway in ergosterol biosynthesis and adaptation to hypoxia was conserved in N. crassa Under lignocellulolytic conditions, the SREBP pathway was highly activated, resulting in the reduced expression of lytic polysaccharide monooxygenases, which require molecular oxygen for catalytic activity. Additionally, activation of the SREBP pathway under lignocellulolytic conditions repressed a set of genes predicted to be involved in the endoplasmic reticulum stress response. Here we show that the inability of a hac-1 mutant, which bears a deletion of the major regulator of the unfolded protein response (UPR), to efficiently produce cellulases and utilize cellulose was suppressed by mutations in the SREBP pathway. The analyses presented here demonstrated new SREBP pathway functions, including linkages to the UPR, and provide new clues for genetic engineering of filamentous fungi to improve their production of extracellular proteins.IMPORTANCE The role of SREBP transcription factors in the regulation of sterol biosynthesis is conserved from humans to yeast. In filamentous fungi, this pathway regulates the secretion of lignocellulolytic enzymes during plant biomass deconstruction. Here we show that the SREBP pathway in Neurospora crassa regulates the production of specific cellulases, lytic polysaccharide monooxygenases that utilize molecular o

The bitter taste receptor TAS2R38 is a G protein coupled receptor (GPCR) that has been found in many extra-oral locations like the gastrointestinal (GI) system, respiratory system, and brain, though its function at these locations is only beginning to be understood. To probe the receptor's potential metabolic role, immunohistochemistry of human ileum tissues was performed, which showed that the receptor was co-localized with glucagon-like peptide 1 (GLP-1) in L-cells. In a previous study, we had modeled the structure of this receptor for its many taste-variant haplotypes (Tan et al. 2011), including the taster haplotype PAV. The structure of this haplotype was then used in a virtual ligand screening pipeline using a collection of ∼2.5 million purchasable molecules from the ZINC database. Three compounds (Z7, Z3, Z1) were purchased from the top hits and tested along with PTU (known TAS2R38 agonist) in in vitro and in vivo assays. The dose-response study of the effect of PTU and Z7 on GLP-1 release using wild-type and TAS2R38 knockout HuTu-80 cells showed that the receptor TAS2R38 plays a major role in GLP-1 release due to these molecules. In vivo studies of PTU and the three compounds showed that they each increase GLP-1 release. PTU was also chemical linked to cellulose to slow its absorption and when tested in vivo, it showed an enhanced and prolonged GLP-1 release. These results suggest that the GI lumen location of TAS2R38 on the L-cell makes it a relatively safe drug target as systemic absorption is not needed for a TAS2R38 agonist drug to effect GLP-1 release.

BackgroundRecent evidence from rat neuron-free mucosa study suggests that the membrane bile acid receptor TGR5 decreases colonic secretion under basal and stimulated conditions. As submucosal neurons are key players in secretory processes and highly express TGR5, we investigated their role in TGR5 agonist-induced inhibition of secretion and the pathways recruited.MethodsTGR5 expression and localization were assessed in rat proximal (pC) and distal (dC) colon by qPCR and immunohistochemistry with double labeling for cholinergic neurons in whole-mount preparations. The influence of a selective (INT-777) or weak (ursodeoxycholic acid, UDCA) TGR5 agonist on colonic secretion was assessed in Ussing chambers, in dC preparation removing seromuscular ± submucosal tissues, in the presence of different inhibitors of secretion pathways.Key resultsTGR5 mRNA is expressed in full thickness dC and pC and immunoreactivity is located in colonocytes and pChAT-positive neurons. Addition of INT-777, and less potently UDCA, decreased colonic secretion in seromuscular stripped dC by -58.17± 2.6%. INT-777 effect on basal secretion was reduced in neuron-free and TTX-treated mucosal-submucosal preparations. Atropine, hexamethonium, indomethacin, and L-NAME all reduced significantly INT-777's inhibitory effect while the 5-HT4 antagonist, RS-39604, and lidocaine abolished it. INT-777 inhibited stimulated colonic secretion induced by nicotine, but not cisapride, carbachol or PGE2.Conclusions & inferencesTGR5 activation inhibits basal and stimulated distal colonic secretion in rats by acting directly on epithelial cells and also inhibiting submucosal neurons. This could represent a counter-regulatory mechanism, at the submucosal level, of the known prosecretory effect of bile acids in the colon.