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This article reports changes to virus taxonomy and taxon nomenclature that were approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in April 2024. The entire ICTV membership was invited to vote on 203 taxonomic proposals that had been approved by the ICTV Executive Committee (EC) in July 2023 at the 55th EC meeting in Jena, Germany, or in the second EC vote in November 2023. All proposals were ratified by online vote. Taxonomic additions include one new phylum (Ambiviricota), one new class, nine new orders, three new suborders, 51 new families, 18 new subfamilies, 820 new genera, and 3547 new species (excluding taxa that have been abolished). Proposals to complete the process of species name replacement to the binomial (genus + species epithet) format were ratified. Currently, a total of 14,690 virus species have been established., (© 2024. The Author(s).)

Objective: Veterans with a history of blast-related mild traumatic brain injury (mTBI) and posttraumatic stress disorder (PTSD) may be at risk for greater cognitive concerns and worse functional outcomes compared to those with either condition in isolation. However, traditional neuropsychological assessment approaches have yielded equivocal results in these populations. The present study examined an alternative method for detecting subtle cognitive inefficiencies: neurocognitive intraindividual variability (IIV), a measure of within-person performance consistency., Method: Participants were 79 male Veterans with a history of blast-related mTBI and current PTSD (mTBI/PTSD group; n = 54) or neither diagnosis (controls; n = 25). Mean T-scores and IIV scores were calculated from neuropsychological measures of attention and speed of information processing (A/SoP) as well as executive functioning (EF)., Results: Global IIV was significantly higher in the mTBI/PTSD group compared to controls (p = .047, Cohen's d = 0.49). At the domain level, larger effect sizes were observed for EF IIV (Cohen's d = 0.46) compared to A/SoP IIV (d = 0.32), although neither were statistically significant. Within the mTBI/PTSD group, higher Global IIV was associated with worse self-reported executive dysfunction, psychological quality of life, and cognitive post-concussive symptoms; at the domain level, these clinical outcomes were generally associated with greater A/SoP IIV (but not EF IIV)., Conclusion: Findings extend previous investigations of neurocognitive IIV in individuals with a history of mTBI across PTSD status. Among Veterans with a history of mTBI and comorbid PTSD, neurocognitive variability may be a better indicator of self-reported cognitive inefficiencies and Veteran experience of daily cognitive functioning than mean neuropsychological performances., (Published by Oxford University Press 2024.)

Acinar cell carcinoma (ACC) and pancreatoblastoma (PBL) are rare pancreatic malignancies with acinar differentiation. Proteogenomic profiling of ACC and PBL revealed distinct protein expression patterns compared to pancreatic ductal adenocarcinoma (PDAC) and benign pancreas. ACC and PBL exhibited similarities, with enrichment in proteins related to RNA processing, chromosome organization, and the mitoribosome, while PDACs overexpressed proteins associated with actin-based processes, extracellular matrix, and immune-active stroma. Pathway activity differences in metabolic adaptation, epithelial-to-mesenchymal transition, and DNA repair were characterized between these diseases. PBL showed upregulation of Wnt-CTNNB1 and IGF2 pathways. Seventeen ACC-specific proteins suggested connections to metabolic diseases with mitochondrial dysfunction, while 34 PBL-specific proteins marked this pediatric cancer with an embryonic stem cell phenotype and alterations in chromosomal proteins and the cell cycle. This study provides novel insights into the proteomic landscapes of ACC and PBL, offering potential targets for diagnostic and therapeutic development., (© 2024. The Author(s).)

Background: Traumatic brain injury (TBI), posttraumatic stress disorder (PTSD), and alcohol use are highly prevalent among military Veterans and independently associated with cognitive difficulties; less is known about the combined effects. This study aimed to investigate the association between alcohol use patterns and cognitive diagnoses in Veterans with TBI and/or PTSD., Methods: Using electronic health record data,193,663 Veterans were classified into three alcohol use trajectory groups (consistently low, initially high transitioning to low, initially moderate transitioning to high) based on self-reported Alcohol Use Disorders Identification Test-C (AUDIT-C) scores. Cox proportional hazards models were used to examine the association between alcohol use patterns, TBI, PTSD, and the risk of cognitive diagnosis, while adjusting for demographic factors and comorbidities., Results: Veterans with initially high transitioning to low (HR = 1.21, 95 % CI: 1.11-1.31) and initially moderate transitioning to high (HR = 1.42, 95 % CI: 1.33-1.51) alcohol use patterns had a significantly greater risk of cognitive diagnosis compared to those with consistently low alcohol use when accounting for TBI, PTSD, and comorbidities. TBI (HR = 5.40, 95 % CI: 5.06-5.76) and PTSD (HR = 2.42, 95 % CI: 2.25-2.61) were also independently associated with an elevated risk of cognitive diagnosis., Conclusions: Findings suggest that Higher levels of alcohol consumption, even if decreasing over time, may confer an increased risk of cognitive diagnosis for Veterans with TBI and/or PTSD. Long-term alcohol use patterns should be considered in clinical assessments and interventions to identify individuals at greater risk for experiencing cognitive difficulties., Competing Interests: Declaration of Competing Interest The authors have no declarations of interest to report., (Published by Elsevier B.V.)

Phagocytosis is an intensely physical process that depends on the mechanical properties of both the phagocytic cell and its chosen target. Here, we employed differentially deformable hydrogel microparticles to examine the role of cargo rigidity in the regulation of phagocytosis by macrophages. Whereas stiff cargos elicited canonical phagocytic cup formation and rapid engulfment, soft cargos induced an architecturally distinct response, characterized by filamentous actin protrusions at the center of the contact site, slower cup advancement, and frequent phagocytic stalling. Using phosphoproteomics, we identified β2 integrins as critical mediators of this mechanically regulated phagocytic switch. Macrophages lacking β2 integrins or their downstream effectors, Talin1 and Vinculin, exhibited specific defects in phagocytic cup architecture and selective suppression of stiff cargo uptake. We conclude that integrin signaling serves as a mechanical checkpoint during phagocytosis to pair cargo rigidity to the appropriate mode of engulfment., (© 2024. The Author(s).)

Pancreatic neuroendocrine tumors (PanNETs) represent well-differentiated endocrine neoplasms with variable clinical outcomes. Predicting patient outcomes using the current tumor grading system is challenging. In addition, traditional systemic treatment options for PanNETs, such as somatostatin analogs or cytotoxic chemotherapies, are very limited. To address these issues, we characterized PanNETs using integrated proteogenomics and identified four subtypes. Two proteomic subtypes showed high recurrence rates, suggesting clinical aggressiveness that was missed by current classification. Hypoxia and inflammatory pathways were significantly enriched in the clinically aggressive subtypes. Detailed analyses revealed metabolic adaptation via glycolysis upregulation and oxidative phosphorylation downregulation under hypoxic conditions. Inflammatory signature analysis revealed that immunosuppressive molecules were enriched in immune hot tumors and might be immunotherapy targets. In this study, we characterized clinically aggressive proteomic subtypes of well-differentiated PanNETs and identified candidate therapeutic targets., Competing Interests: D.S.K. is now an employee of Paige.AI. M.H.R. is a member of the Scientific Advisory Boards of Azenta Life Sciences and Universal Diagnostics (UDX). None of these companies had any role in the support, design, execution, data analysis, or any other aspect of this study., (© 2024 Published by Elsevier Inc.)

Ubiquitin-like protein ISG15 plays an important role in an array of cellular functions via its covalent attachment to target proteins (ISGylation). Here, we present a protocol for the identification of ISGylated proteins that avoids the caveats associated with ISG15 overexpression and minimizes the likelihood of false positives. We describe steps for the tagging of endogenous ISG15, followed by genotyping and clone selection. We then detail steps for ISGylation induction, the isolation of ISGylated proteins, and their identification via quantitative mass spectrometry. For complete details on the use and execution of this protocol, please refer to Wardlaw and Petrini. 1 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Outer membrane proteins perform essential functions in uptake and secretion processes in bacteria. MspA is an octameric channel protein in the outer membrane of Mycobacterium smegmatis and is structurally distinct from any other known outer membrane protein. MspA is the founding member of a family with more than 3000 homologs and is one of the most widely used proteins in nanotechnological applications due to its advantageous pore structure and extraordinary stability. While a conserved C-terminal signal sequence is essential for folding and protein assembly in the outer membrane of Gram-negative bacteria, the molecular determinants of these processes are unknown for MspA. In this study, we show that mutation and deletion of methionine 183 in the highly conserved C-terminus of MspA and mutation of the conserved tryptophan 40 lead to a complete loss of protein in heat extracts of M. smegmatis. Swapping these residues partially restores the heat stability of MspA indicating that methionine 183 and tryptophan 40 form a conserved sulfur-π electron interaction, which stabilizes the MspA monomer. Flow cytometry showed that all MspA mutants are surface-accessible demonstrating that oligomerization and membrane integration in M. smegmatis are not affected. Thus, the conserved C-terminus of MspA is essential for its thermal stability, but it is not required for protein assembly in its native membrane, indicating that this process is mediated by a mechanism distinct from that in Gram-negative bacteria. These findings will benefit the rational design of MspA-like pores to tailor their properties in current and future applications., (© 2024 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)

Metastatic progression of colorectal adenocarcinoma (CRC) remains poorly understood and poses significant challenges for treatment. To overcome these challenges, we performed multiomics analyses of primary CRC and liver metastases. Genomic alterations, such as structural variants or copy number alterations, were enriched in oncogenes and tumor suppressor genes and increased in metastases. Unsupervised mass spectrometry-based proteomics of 135 primary and 123 metastatic CRCs uncovered distinct proteomic subtypes, three each for primary and metastatic CRCs, respectively. Integrated analyses revealed that hypoxia, stemness, and immune signatures characterize these 6 subtypes. Hypoxic CRC harbors high epithelial-to-mesenchymal transition features and metabolic adaptation. CRC with a stemness signature shows high oncogenic pathway activation and alternative telomere lengthening (ALT) phenotype, especially in metastatic lesions. Tumor microenvironment analysis shows immune evasion via modulation of major histocompatibility complex (MHC) class I/II and antigen processing pathways. This study characterizes both primary and metastatic CRCs and provides a large proteogenomics dataset of metastatic progression., Competing Interests: Declaration of interests D.S.K. was a consultant for and is now employed by Paige.AI. P.G.B. and J.G. are employees of and hold equity interests in Genuity Science. J.Y.W. is the founder of Curandis. M.H.A.R. is a member of the scientific advisory boards of Azenta Life Sciences and Universal DX. None of these companies had any role in design, execution, data analysis, or any other aspect of this study., (Copyright © 2024. Published by Elsevier Inc.)

Efficient treatment of acute myeloid leukemia (AML) patients remains a challenge despite recent therapeutic advances. Here, using a CRISPRi screen targeting chromatin factors, we identified the nucleosome-remodeling factor (NURF) subunit BPTF as an essential regulator of AML cell survival. We demonstrate that BPTF forms an alternative NURF chromatin remodeling complex with SMARCA5 and BAP18, which regulates the accessibility of a large set of insulator regions in leukemic cells. This ensures efficient CTCF binding and boundary formation between topologically associated domains that is essential for maintaining the leukemic transcriptional programs. We also demonstrate that the well-studied PHD2-BROMO chromatin reader domains of BPTF, while contributing to complex recruitment to chromatin, are dispensable for leukemic cell growth. Taken together, our results uncover how the alternative NURF complex contributes to leukemia and provide a rationale for its targeting in AML., (© 2023 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

Objective/Background: Healthcare workers have experienced high rates of psychiatric symptom burden and occupational attrition during the COVID-19 pandemic. Identifying contributory factors can inform prevention and mitigation measures. Here, we explore the potential contributions of occupational stressors vs COVID-19 infection to insomnia symptoms in US healthcare workers.Patients/Methods: An online self-report survey was collected between September 2020 and July 2022 from N = 594 US healthcare workers, with longitudinal follow-up up to 9 months. Assessments included the Insomnia Severity Index (ISI), the PTSD Checklist for DSM-5 (PCL-5), and a 13-item scale assessing COVID-19 related occupational stressors., Results: Insomnia was common (45% of participants reported at least moderate and 9.2% reported severe symptoms at one or more timepoint) and significantly associated with difficulty completing work-related tasks, increased likelihood of occupational attrition, and thoughts of suicide or self-harm (all p<.0001). In multivariable regression with age, gender, and family COVID-19 history as covariates, past two-week COVID-related occupational stressors, peak COVID-related occupational stressors, and personal history of COVID-19 infection were all significantly related to past two-week ISI scores (β = 1.7 ± 0.14SE, β = 0.08 ± 0.03, and β = 0.69 ± 0.22 respectively). Although similar results were found for the PCL-5, when ISI and PCL-5 items were separated by factor, COVID-19 infection was significantly related only to the factor consisting of sleep-related items., Conclusions: Both recent occupational stress and personal history of COVID-19 infection were significantly associated with insomnia in healthcare workers. These results suggest that both addressing occupational stressors and reducing rate of COVID-19 infection are important to protect healthcare workers and the healthcare workforce., Competing Interests: The views expressed are those of the authors and do not reflect the official policy of the Department of Veterans Affairs or the U.S. Government. There are no other conflicts of interests for any authors.

The FOXA1 pioneer factor is an essential mediator of steroid receptor function in multiple hormone-dependent cancers, including breast and prostate cancers, enabling nuclear receptors such as estrogen receptor (ER) and androgen receptor (AR) to activate lineage-specific growth programs. FOXA1 is also highly expressed in non-small cell lung cancer (NSCLC), but whether and how it regulates tumor growth in this context is not known. Analyzing data from loss-of-function screens, we identified a subset of NSCLC tumor lines where proliferation is FOXA1 dependent. Using rapid immunoprecipitation and mass spectrometry of endogenous protein, we identified chromatin-localized interactions between FOXA1 and glucocorticoid receptor (GR) in these tumor cells. Knockdown of GR inhibited proliferation of FOXA1-dependent, but not FOXA1-independent NSCLC cells. In these FOXA1-dependent models, FOXA1 and GR cooperate to regulate gene targets involved in EGF signaling and G 1 -S cell-cycle progression. To investigate the therapeutic potential for targeting this complex, we examined the effects of highly selective inhibitors of the GR ligand-binding pocket and found that GR antagonism with ORIC-101 suppressed FOXA1/GR target expression, activation of EGF signaling, entry into the S-phase, and attendant proliferation in vitro and in vivo . Taken together, our findings point to a subset of NSCLCs harboring a dependence on the FOXA1/GR growth program and provide rationale for its therapeutic targeting., Significance: NSCLC is the leading cause of cancer deaths worldwide. There is a need to identify novel druggable dependencies. We identify a subset of NSCLCs dependent on FOXA1-GR and sensitive to GR antagonism., (© 2023 The Authors; Published by the American Association for Cancer Research.)

This article reports changes to virus taxonomy and taxon nomenclature that were approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in April 2023. The entire ICTV membership was invited to vote on 174 taxonomic proposals that had been approved by the ICTV Executive Committee in July 2022, as well as a proposed revision of the ICTV Statutes. All proposals and the revised ICTV Statutes were approved by a majority of the voting membership. Of note, the ICTV continued the process of renaming existing species in accordance with the recently mandated binomial format and included gene transfer agents (GTAs) in the classification framework by classifying them as viriforms. In total, one class, seven orders, 31 families, 214 genera, and 858 species were created., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Objective: Prospective memory (PM) or "remembering to remember" has been shown to be reduced in Veterans with histories of mild traumatic brain injury (mTBI), particularly on tasks with high strategic demands such as recalling time-based information in the absence of external cues. This study examined whether time monitoring during a PM task was reduced in Veterans with a history of mTBI and was associated with time-based PM performance. Method: Veterans with a history of mTBI (n = 49) and Veterans without a history of TBI (n = 16) completed the Memory for Intentions Screening Test (MIST) as a measure of PM during which their time monitoring (i.e. number of clock checks) was recorded. Results: Adjusting for age, education, depression, and PTSD symptoms, negative binomial regression revealed that the mTBI group checked the clock less frequently compared to the control group (Cohen's d  = 0.84, p  = 0.005). Within the mTBI group, less frequent time monitoring across the entire MIST task was associated with poorer time-based MIST performance ( r s = .57, p  < 0.001), but not with event-based MIST ( r s = .04, p  = 0.768). Conclusions: Veterans with a history of mTBI evidenced significantly reduced time monitoring during a PM task compared to Veterans without a history mTBI, which was associated with strategically-demanding PM. Current findings provide that mTBI-associated difficulties with strategic aspects of PM may be due to reduced time monitoring. Future studies are needed to determine if reduced time monitoring also contributes to mTBI-associated PM difficulties in the real-world (e.g. medication non-adherence).

Trimethylation of histone H3 lysine 4 (H3K4me3) is associated with transcriptional start sites and has been proposed to regulate transcription initiation 1,2 . However, redundant functions of the H3K4 SET1/COMPASS methyltransferase complexes complicate the elucidation of the specific role of H3K4me3 in transcriptional regulation 3,4 . Here, using mouse embryonic stem cells as a model system, we show that acute ablation of shared subunits of the SET1/COMPASS complexes leads to a complete loss of all H3K4 methylation. Turnover of H3K4me3 occurs more rapidly than that of H3K4me1 and H3K4me2 and is dependent on KDM5 demethylases. Notably, acute loss of H3K4me3 does not have detectable effects on transcriptional initiation but leads to a widespread decrease in transcriptional output, an increase in RNA polymerase II (RNAPII) pausing and slower elongation. We show that H3K4me3 is required for the recruitment of the integrator complex subunit 11 (INTS11), which is essential for the eviction of paused RNAPII and transcriptional elongation. Thus, our study demonstrates a distinct role for H3K4me3 in transcriptional pause-release and elongation rather than transcriptional initiation., (© 2023. The Author(s).)

Metastatic prostate cancer (PCa) inevitably acquires resistance to standard therapy preceding lethality. Here, we unveil a chromosomal instability (CIN) tolerance mechanism as a therapeutic vulnerability of therapy-refractory lethal PCa. Through genomic and transcriptomic analysis of patient datasets, we find that castration and chemotherapy-resistant tumors display the highest CIN and mitotic kinase levels. Functional genomics screening coupled with quantitative phosphoproteomics identify MASTL kinase as a survival vulnerability specific of chemotherapy-resistant PCa cells. Mechanistically, MASTL upregulation is driven by transcriptional rewiring mechanisms involving the non-canonical transcription factors androgen receptor splice variant 7 and E2F7 in a circuitry that restrains deleterious CIN and prevents cell death selectively in metastatic therapy-resistant PCa cells. Notably, MASTL pharmacological inhibition re-sensitizes tumors to standard therapy and improves survival of pre-clinical models. These results uncover a targetable mechanism promoting high CIN adaptation and survival of lethal PCa., Competing Interests: Declaration of interests The authors declare no competing financial interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Long noncoding RNAs (lncRNAs) play essential roles in the development and progression of many cancers. However, the contributions of lncRNAs to medulloblastoma (MB) remain poorly understood. Here, we identify Miat as an lncRNA enriched in the sonic hedgehog group of MB that is required for maintenance of a treatment-resistant stem-like phenotype in the disease. Loss of Miat results in the differentiation of tumor-initiating, stem-like MB cells and enforces the differentiation of tumorigenic stem-like MB cells into a nontumorigenic state. Miat expression in stem-like MB cells also facilitates treatment resistance by down-regulating p53 signaling and impairing radiation-induced cell death, which can be reversed by therapeutic inhibition of Miat using antisense oligonucleotides. Mechanistically, the RNA binding protein Metadherin (Mtdh), previously linked to resistance to cytotoxic therapy in cancer, binds to Miat in stem-like MB cells. Like the loss of Miat , the loss of Mtdh reduces tumorigenicity and increases sensitivity to radiation-induced death in stem-like MB cells. Moreover, Miat and Mtdh function to regulate the biogenesis of several microRNAs and facilitate tumorigenesis and treatment resistance. Taken together, these data reveal an essential role for the lncRNA Miat in sustaining a treatment-resistant pool of tumorigenic stem-like MB cells.

Estrogen receptor alpha (ERα) drives mammary gland development and breast cancer (BC) growth through an evolutionarily conserved linkage of DNA binding and hormone activation functions. Therapeutic targeting of the hormone binding pocket is a widely utilized and successful strategy for breast cancer prevention and treatment. However, resistance to this endocrine therapy is frequently encountered and may occur through bypass or reactivation of ER-regulated transcriptional programs. We now identify the induction of an ERα isoform, ERα-LBD, that is encoded by an alternative ESR1 transcript and lacks the activation function and DNA binding domains. Despite lacking the transcriptional activity, ERα-LBD is found to promote breast cancer growth and resistance to the ERα antagonist fulvestrant. ERα-LBD is predominantly localized to the cytoplasm and mitochondria of BC cells and leads to enhanced glycolysis, respiration and stem-like features. Intriguingly, ERα-LBD expression and function does not appear to be restricted to cancers that express full length ERα but also promotes growth of triple-negative breast cancers and ERα-LBD transcript (ESR1-LBD) is also present in BC samples from both ERα(+) and ERα(-) human tumors. These findings point to ERα-LBD as a potential mediator of breast cancer progression and therapy resistance., (© 2022. The Author(s).)

Here, we show that the capacity to manufacture IL-2 identifies constituents of the expanded CD8 T cell effector pool that display stem-like features, preferentially survive, rapidly attain memory traits, resist exhaustion, and control chronic viral challenges. The cell-intrinsic synthesis of IL-2 by CD8 T cells attenuates the ability to receive IL-2-dependent STAT5 signals, thereby limiting terminal effector formation, endowing the IL-2-producing effector subset with superior protective powers. In contrast, the non-IL-2-producing effector cells respond to IL-2 signals and gain effector traits at the expense of memory formation. Despite having distinct properties during the effector phase, IL-2-producing and nonproducing CD8 T cells appear to converge transcriptionally as memory matures to form populations with equal recall abilities. Therefore, the potential to produce IL-2 during the effector, but not memory stage, is a consequential feature that dictates the protective capabilities of the response.

Chimeric antigen receptors (CARs) are receptors for antigen that direct potent immune responses. Tumor escape associated with low target antigen expression is emerging as one potential limitation of their efficacy. Here we edit the TRAC locus in human peripheral blood T cells to engage cell-surface targets through their T cell receptor-CD3 complex reconfigured to utilize the same immunoglobulin heavy and light chains as a matched CAR. We demonstrate that these HLA-independent T cell receptors (HIT receptors) consistently afford high antigen sensitivity and mediate tumor recognition beyond what CD28-based CARs, the most sensitive design to date, can provide. We demonstrate that the functional persistence of HIT T cells can be augmented by constitutive coexpression of CD80 and 4-1BBL. Finally, we validate the increased antigen sensitivity afforded by HIT receptors in xenograft mouse models of B cell leukemia and acute myeloid leukemia, targeting CD19 and CD70, respectively. Overall, HIT receptors are well suited for targeting cell surface antigens of low abundance., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: The COVID-19 pandemic has greatly affected front-line health care workers (HCW) and first responders (FR). The specific components of COVID-19 related occupational stressors (CROS) associated with psychiatric symptoms and reduced occupational functioning or retention remain poorly understood., Objectives: Examine the relationships between total and factored CROS, psychiatric symptoms, and occupational outcomes., Design: Observational, self-report, single time-point online assessment., Participants: A total of 510 US HCW (N = 301) and FR (N = 200) with occupational duties affected by the COVID-19 pandemic., Main Outcomes and Measures: CROS were assessed using a custom 17-item questionnaire. Post-traumatic stress disorder (PTSD), depression, insomnia, and generalized anxiety symptoms were assessed using the PTSD Checklist-5 (PCL5), Patient Health Questionnaire-9 (PHQ9), Insomnia Severity Index (ISI), and General Anxiety Disorder-7 (GAD7). Respondents' likelihood of leaving current field and occupational functioning were assessed with 2-item PROMIS subscales. Relationships were modeled using multivariable regression. Open-ended responses were coded using rapid template analysis., Results: CROS total scores correlated significantly with all four psychiatric symptom domains (R's = .42-.53), likelihood of leaving one's current occupation (R = .18), and trouble doing usual work (R = .28), all p's < .001. Half of HCW indicated a decreased likelihood of staying in their current occupation as a result of the pandemic. CROS were fit to a 3-factor model consisting of risk, demoralization, and volume factors. All CROS factors were associated with psychiatric symptom burden, but demoralization was most prominently associated with psychiatric symptoms and negative occupational outcomes. Among psychiatric symptoms, PTSD symptoms were most strongly associated with negative occupational outcomes. Open-ended statements emphasized lack of protection and support, increased occupational demands, and emotional impact of work duties., Conclusions and Relevance: These results demonstrate potentially treatable psychiatric symptoms in HCW and FR experiencing CROS, impacting both wellbeing and the health care system. Mitigating CROS, particularly by addressing factors driving demoralization, may improve HCW and FR mental health, occupational functioning, and retention., (© 2021. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

Blast exposure ( via detonation of high explosives) represents a major potential trauma source for Servicemembers and Veterans, often resulting in mild traumatic brain injury (mTBI). Executive dysfunction (e.g., alterations in memory, deficits in mental flexibility, difficulty with adaptability) is commonly reported by Veterans with a history of blast-related mTBI, leading to impaired daily functioning and decreased quality of life, but underlying mechanisms are not fully understood and have not been well studied in animal models of blast. To investigate potential underlying behavioral mechanisms contributing to deficits in executive functioning post-blast mTBI, here we examined how a history of repetitive blast exposure in male mice affects anxiety/compulsivity-like outcomes and appetitive goal-directed behavior using an established mouse model of blast mTBI. We hypothesized that repetitive blast exposure in male mice would result in anxiety/compulsivity-like outcomes and corresponding performance deficits in operant-based reward learning and behavioral flexibility paradigms. Instead, results demonstrate an increase in reward-seeking and goal-directed behavior and a congruent decrease in behavioral flexibility. We also report chronic adverse behavioral changes related to anxiety, compulsivity, and hyperarousal. In combination, these data suggest that potential deficits in executive function following blast mTBI are at least in part related to enhanced compulsivity/hyperreactivity and behavioral inflexibility and not simply due to a lack of motivation or inability to acquire task parameters, with important implications for subsequent diagnosis and treatment management., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Baskin, Lee, Skillen, Wong, Rau, Hendrickson, Pagulayan, Raskind, Peskind, Phillips, Cook and Schindler.)

Copper serves as a co-factor for a host of metalloenzymes that contribute to malignant progression. The orally bioavailable copper chelating agent tetrathiomolybdate (TM) has been associated with a significant survival benefit in high-risk triple negative breast cancer (TNBC) patients. Despite these promising data, the mechanisms by which copper depletion impacts metastasis are poorly understood and this remains a major barrier to advancing TM to a randomized phase II trial. Here, using two independent TNBC models, we report a discrete subpopulation of highly metastatic SOX2/OCT4+ cells within primary tumors that exhibit elevated intracellular copper levels and a marked sensitivity to TM. Global proteomic and metabolomic profiling identifies TM-mediated inactivation of Complex IV as the primary metabolic defect in the SOX2/OCT4+ cell population. We also identify AMPK/mTORC1 energy sensor as an important downstream pathway and show that AMPK inhibition rescues TM-mediated loss of invasion. Furthermore, loss of the mitochondria-specific copper chaperone, COX17, restricts copper deficiency to mitochondria and phenocopies TM-mediated alterations. These findings identify a copper-metabolism-metastasis axis with potential to enrich patient populations in next-generation therapeutic trials., (© 2021. The Author(s).)

This article reports the changes to virus taxonomy approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in March 2021. The entire ICTV was invited to vote on 290 taxonomic proposals approved by the ICTV Executive Committee at its meeting in October 2020, as well as on the proposed revision of the International Code of Virus Classification and Nomenclature (ICVCN). All proposals and the revision were ratified by an absolute majority of the ICTV members. Of note, ICTV mandated a uniform rule for virus species naming, which will follow the binomial 'genus-species' format with or without Latinized species epithets. The Study Groups are requested to convert all previously established species names to the new format. ICTV has also abolished the notion of a type species, i.e., a species chosen to serve as a name-bearing type of a virus genus. The remit of ICTV has been clarified through an official definition of 'virus' and several other types of mobile genetic elements. The ICVCN and ICTV Statutes have been amended to reflect these changes., (© 2021. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

ID proteins are helix-loop-helix (HLH) transcriptional regulators frequently overexpressed in cancer. ID proteins inhibit basic-HLH transcription factors often blocking differentiation and sustaining proliferation. A small-molecule, AGX51, targets ID proteins for degradation and impairs ocular neovascularization in mouse models. Here we show that AGX51 treatment of cancer cell lines impairs cell growth and viability that results from an increase in reactive oxygen species (ROS) production upon ID degradation. In mouse models, AGX51 treatment suppresses breast cancer colonization in the lung, regresses the growth of paclitaxel-resistant breast tumors when combined with paclitaxel and reduces tumor burden in sporadic colorectal neoplasia. Furthermore, in cells and mice, we fail to observe acquired resistance to AGX51 likely the result of the inability to mutate the binding pocket without loss of ID function and efficient degradation of the ID proteins. Thus, AGX51 is a first-in-class compound that antagonizes ID proteins, shows strong anti-tumor effects and may be further developed for the management of multiple cancers.

Acetylation of lysine 16 on histone H4 (H4K16ac) is catalyzed by histone acetyltransferase KAT8 and can prevent chromatin compaction in vitro. Although extensively studied in Drosophila, the functions of H4K16ac and two KAT8-containing protein complexes (NSL and MSL) are not well understood in mammals. Here, we demonstrate a surprising complex-dependent activity of KAT8: it catalyzes H4K5ac and H4K8ac as part of the NSL complex, whereas it catalyzes the bulk of H4K16ac as part of the MSL complex. Furthermore, we show that MSL complex proteins and H4K16ac are not required for cell proliferation and chromatin accessibility, whereas the NSL complex is essential for cell survival, as it stimulates transcription initiation at the promoters of housekeeping genes. In summary, we show that KAT8 switches catalytic activity and function depending on its associated proteins and that, when in the NSL complex, it catalyzes H4K5ac and H4K8ac required for the expression of essential genes., Competing Interests: Declaration of interests EpiCypher is a commercial developer and supplier of reagents (recombinant semi-synthetic modified nucleosomes) and the antibody characterization platforms used in this study. K.H. is a consultant for Inthera Bioscience AG and a scientific advisor for Hannibal Health Innovation., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Increasing evidence supports the role of brain and systemic inflammation in the etiology of Parkinson disease (PD). We used gene expression profiling to examine the activation state of peripheral blood monocytes in 18 patients with early, untreated PD and 16 healthy control (HC) subjects. Monocytes were isolated by negative selection, and gene expression studied by RNA-seq and gene set enrichment analysis. A computational model that incorporated case/control status, sex, and the interaction between case/control status and sex was utilized. We found that there was a striking effect of sex on monocyte gene expression. There was inflammatory activation of monocytes in females with PD, with enrichment of gene sets associated with interferon gamma stimulation. In males, the activation patterns were more heterogeneous. These data point to the importance of systemic monocyte activation in PD, and the importance of studies which examine the differential effects of sex on pathophysiology of the disease.

Posttraumatic stress disorder (PTSD) is linked to both altered physiological functioning and poorer cardiovascular health outcomes, including an increased risk for cardiovascular disease and cardiovascular-related mortality. An important question is whether interventions for PTSD might ameliorate the risk for poorer health by improving cardiovascular physiological intermediaries. To begin to characterize the literature addressing this question, we conducted a systematic review of empirical studies examining the impact of PTSD interventions on cardiovascular physiological intermediaries, including blood pressure (BP), heart rate (HR), cardiac impedance, and subclinical atherosclerosis. Outcomes included both tonic (i.e., resting) cardiovascular functioning and cardiovascular reactivity (CVR). A total of 44 studies met the inclusion criteria. There was mixed evidence regarding whether PTSD treatment improved tonic cardiovascular functioning. There was stronger evidence that PTSD treatments reduced CVR to trauma-related stressors, particularly for higher-quality studies of cognitive behavioral interventions. No studies examined cardiac impedance or subclinical atherosclerosis. The studies had a high degree of heterogeneity in the populations sampled and interventions tested. Moreover, they generally included small sample sizes and lacked control conditions. Interventions for PTSD may improve cardiovascular physiological outcomes, particularly CVR to trauma cues, although additional methodologically rigorous studies are needed. We outline changes to future research that would improve the literature regarding this important question, including the more frequent use of control groups and larger sample sizes., (© 2020 International Society for Traumatic Stress Studies.)

Chaperones aid in protein folding and maintenance of protein integrity. In doing so, they have the unique ability to directly stabilize resistance-conferring amino acid substitutions in drug targets and to counter the stress imparted by these substitutions, thus supporting heritable antimicrobial resistance (AMR). We asked whether chaperones support AMR in Mycobacterium smegmatis , a saprophytic model of Mycobacterium tuberculosis , the causative agent of tuberculosis (TB). We show that DnaK associates with many drug targets and that DnaK associates more with AMR-conferring mutant RNA polymerase (RNAP) than with wild-type RNAP. In addition, frequency-of-resistance (FOR) and fitness studies reveal that the DnaK system of chaperones supports AMR in antimicrobial targets in mycobacteria, including RNAP and the ribosome. These findings highlight chaperones as potential targets for drugs to overcome AMR in mycobacteria, including M. tuberculosis , as well as in other pathogens. IMPORTANCE AMR is a global problem, especially for TB. Here, we show that mycobacterial chaperones support AMR in M. smegmatis , a nonpathogenic model of M. tuberculosis , the causative agent of TB. In particular, the mycobacterial DnaK system of chaperones supports AMR in the antimicrobial targets RNA polymerase and the ribosome. This is the first report showing a role for protein chaperones in mediating AMR in mycobacteria. Given the widespread role of protein chaperones in enabling genomic diversity, we anticipate that our findings can be extended to other microbes., (Copyright © 2021 Fay et al.)

Background: The α 1 -adrenoreceptor antagonist prazosin has in many but not all studies been found to be effective for PTSD associated nightmares, hyperarousal symptoms, and total symptom severity. The particular efficacy of prazosin for nightmares and hyperarousal symptoms suggests there may be a subset of PTSD symptoms that are more tightly associated with an α 1 -adrenoreceptor mediated noradrenergic mechanism, but cross traditional diagnostic symptom clusters. However, the efficacy of prazosin for individual symptoms other than nightmares and sleep disruption has not previously been examined., Methods: In a post hoc reanalysis of a previously published, randomized controlled trial of twice daily prazosin for PTSD, we examined the relative effect of prazosin on individual items of the CAPS for DSM-IV, and tested whether prazosin responsiveness predicted the partial correlation of the changes in symptom intensity at the level of individual subjects. Results were not adjusted for multiple comparisons., Results: Prazosin showed the largest effect for distressing dreams, anhedonia, difficulty falling or staying asleep, difficulty concentrating, and hypervigilance. These items were also (a) of higher baseline severity in the underlying population, and (b) more related in how they fluctuated at the level of individual subjects. Covariance analysis did not support a clear cutoff between highly prazosin responsive items and those showing a smaller, not statistically significant response., Conclusions: In this data set, twice daily prazosin substantially reduced not only nightmares and sleep disruption, but the majority of hyperarousal symptoms, with some evidence of efficacy for avoidance symptoms. The relationship of baseline symptom distribution to which symptoms showed significant response to prazosin reinforces the possibility that differences in a clinical trial's participant populations may significantly influence trial outcome. The pattern of symptom endorsement at the level of individual subjects was consistent with prazosin-responsive items sharing a common pathophysiologic mechanism., Competing Interests: Declaration of Conflicting Interests: The author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: Dr. Raskind is a paid advisory board member of Pfizer Laboratories, Merck, and Takeda Pharmaceuticals. Dr. Peskind is a paid advisory board member for Lilly, Takeda, Merck, and Avanir pharmaceuticals. All other authors report no financial relationships with commercial interests. The views expressed are those of the authors and do not reflect the official policy of the Department of Veterans Affairs or the U.S. Government. The investigators have adhered to the policies for protection of human participants as prescribed in 45 CFR 46., (© The Author(s) 2021.)

DEAD-box RNA helicase DDX21 (also named nucleolar RNA helicase 2) is a nuclear autoantigen with undefined roles in cancer. To explore possible roles of autoimmune recognition in cancer immunity, we examined DDX21 protein expression in colorectal cancer tissue and its association with patient clinical outcomes. Unbiased deep proteomic profiling of two independent colorectal cancer cohorts using mass spectrometry showed that DDX21 protein was significantly upregulated in cancer relative to benign mucosa. We then examined DDX21 protein expression in a validation group of 710 patients, 619 of whom with early stage and 91 with late stage colorectal cancers. DDX21 was detected mostly in the tumor cell nuclei, with high expression in some mitotic cells. High levels of DDX21 protein were found in 28% of stage I, 21% of stage II, 30% of stage III, and 32% of stage IV colorectal cancer cases. DDX21 expression levels correlated with non-mucinous histology in early stage cancers but not with other clinicopathological features such as patient gender, age, tumor location, tumor grade, or mismatch repair status in any cancer stage. Kaplan-Meier analyses revealed that high DDX21 protein levels was associated with longer survival in patients with early stage colorectal cancer, especially longer disease-free survival in patients with microsatellite instability (MSI) cancers, but no such correlations were found for the microsatellite stable subtype or late stage colorectal cancer. Univariate and multivariate analyses also identified high DDX21 protein expression as an independent favorable prognostic marker for early stage MSI colorectal cancer.

Mounting evidence points to the significance of neurovascular-related dysfunction in veterans with blast-related mTBI, which is also associated with reduced [ 18 F]-fluorodeoxyglucose (FDG) uptake. The goal of this study was to determine whether plasma VEGF-A is altered in veterans with blast-related mTBI and address whether VEGF-A levels correlate with FDG uptake in the cerebellum, a brain region that is vulnerable to blast-related injury 72 veterans with blast-related mTBI (mTBI) and 24 deployed control (DC) veterans with no lifetime history of TBI were studied. Plasma VEGF-A was significantly elevated in mTBIs compared to DCs. Plasma VEGF-A levels in mTBIs were significantly negatively correlated with FDG uptake in cerebellum. In addition, performance on a Stroop color/word interference task was inversely correlated with plasma VEGF-A levels in blast mTBI veterans. Finally, we observed aberrant perivascular VEGF-A immunoreactivity in postmortem cerebellar tissue and not cortical or hippocampal tissues from blast mTBI veterans. These findings add to the limited number of plasma proteins that are chronically elevated in veterans with a history of blast exposure associated with mTBI. It is likely the elevated VEGF-A levels are from peripheral sources. Nonetheless, increasing plasma VEGF-A concentrations correlated with chronically decreased cerebellar glucose metabolism and poorer performance on tasks involving cognitive inhibition and set shifting. These results strengthen an emerging view that cognitive complaints and functional brain deficits caused by blast exposure are associated with chronic blood-brain barrier injury and prolonged recovery in affected regions., (Copyright © 2020. Published by Elsevier B.V.)

Background and Significance: Parallel-group randomized controlled trials (PG-RCTs) are the gold standard for detecting differences in mean improvement across treatment conditions. However, PG-RCTs provide limited information about individuals, making them poorly optimized for quantifying the relationship of a biomarker measured at baseline with treatment response. In N-of-1 trials, an individual subject moves between treatment conditions to determine their specific response to each treatment. Aggregated N-of-1 trials analyze a cohort of such participants, and can be designed to optimize both statistical power and clinical or logistical constraints, such as allowing all participants to begin with an open-label stabilization phase to facilitate the enrollment of more acutely symptomatic participants. Here, we describe a set of statistical simulation studies comparing the power of four different trial designs to detect a relationship between a predictive biomarker measured at baseline and subjects' specific response to the PTSD pharmacotherapeutic agent prazosin. Methods: Data was simulated from 4 trial designs: (1) open-label; (2) open-label + blinded discontinuation; (3) traditional crossover; and (4) open label + blinded discontinuation + brief crossover (the N-of-1 design). Designs were matched in length and assessments. The primary outcome, analyzed with a linear mixed effects model, was whether a statistically significant association between biomarker value and response to prazosin was detected with 5% Type I error. Simulations were repeated 1,000 times to determine power and bias, with varied parameters. Results: Trial designs 2 & 4 had substantially higher power with fewer subjects than open label design. Trial design 4 also had higher power than trial design 2. Trial design 4 had slightly lower power than the traditional crossover design, although power declined much more rapidly as carryover was introduced. Conclusions: These results suggest that an aggregated N-of-1 trial design beginning with an open label titration phase may provide superior power over open label or open label and blinded discontinuation designs, and similar power to a traditional crossover design, in detecting an association between a predictive biomarker and the clinical response to the PTSD pharmacotherapeutic prazosin. This is achieved while allowing all participants to spend the first 8 weeks of the trial on open-label active treatment., (Copyright © 2020 Hendrickson, Thomas, Schork and Raskind.)

Reduced working memory is frequently reported by Veterans with a history of blast-related mild traumatic brain injury (mTBI), but can be difficult to quantify on neuropsychological measures. This study aimed to improve our understanding of the impact of blast-related mTBI on the working memory system by using resting state functional magnetic resonance imaging (fMRI) to explore differences in functional connectivity between OEF/OIF/OND Veterans with and without a history of mTBI. Participants were twenty-four Veterans with a history of blast-related mTBI and 17 Veterans who were deployed but had no lifetime history of TBI. Working memory ability was evaluated with the Auditory Consonants Trigrams (ACT) task. Resting state fMRI was used to evaluate intrinsic functional connectivity from frontal seed regions that are known components of the working memory network. No significant group differences were found on the ACT, but the imaging analyses revealed widespread hyper-connectivity from the frontal seed regions in the Veterans with a history of mTBI relative to the deployed control group. Further, within the mTBI group, but not the control group, better performance on the ACT was associated with increased functional connectivity to multiple brain regions, including cerebellar components of the working memory network. These results were present after controlling for age, PTSD symptoms, and estimated premorbid IQ, and suggest that long-term alterations in the functional connectivity of the working memory network following blast-related mTBI may reflect a compensatory change that contributes to intact performance on an objective measure of working memory.

Theranostic agents should ideally be renally cleared and biodegradable. Here, we report the synthesis, characterization and theranostic applications of fluorescent ultrasmall gold quantum clusters that are stabilized by the milk metalloprotein alpha-lactalbumin. We synthesized three types of these nanoprobes that together display fluorescence across the visible and near-infrared spectra when excited at a single wavelength through optical colour coding. In live tumour-bearing mice, the near-infrared nanoprobe generates contrast for fluorescence, X-ray computed tomography and magnetic resonance imaging, and exhibits long circulation times, low accumulation in the reticuloendothelial system, sustained tumour retention, insignificant toxicity and renal clearance. An intravenously administrated near-infrared nanoprobe with a large Stokes shift facilitated the detection and image-guided resection of breast tumours in vivo using a smartphone with modified optics. Moreover, the partially unfolded structure of alpha-lactalbumin in the nanoprobe helps with the formation of an anti-cancer lipoprotein complex with oleic acid that triggers the inhibition of the MAPK and PI3K-AKT pathways, immunogenic cell death and the recruitment of infiltrating macrophages. The biodegradability and safety profile of the nanoprobes make them suitable for the systemic detection and localized treatment of cancer.

Colorectal cancers are among the most common cancers and a leading cause of cancer death. In our pursuit to discover molecular markers for better characterization and precision theranostics of these cancers, we first conducted global deep proteome analyses and identified maspin (serpin B5, peptidase inhibitor 5) as an upregulated protein in tumor tissue. We then validated its expression in a large cohort of 743 patients with colorectal cancers of all stages and found that both cytoplasmic and nuclear expression varied widely between different patients. Comparison with clinicopathological features revealed that maspin expression levels correlate significantly only with mismatch repair (MMR) status but not with other features. To elucidate the prognostic significance of maspin, we analyzed two outcome-annotated cohorts, one of 572 early stage cancer patients and another of 93 late stage cancer patients. Kaplan-Meier survival, univariate, and multivariate analyses revealed that maspin overexpression predicts longer overall and disease-free survival for early stage microsatellite instability (MSI) subtype colorectal cancer, but there is no correlation with survival for patients with early stage cancer of the microsatellite stability (MSS) subtype or late stage cancer. Our study identifies maspin expression as an independent prognostic marker for risk stratification of early stage MSI subtype colorectal cancer and may provide guidance for improved therapeutic management., (Copyright © 2020 Tanaka, Wang, Shia, Zhou, Ogawa, Hendrickson, Klimstra and Roehrl.)

Glutaric aciduria type 1 (GA1) is an inborn error of lysine degradation characterized by a specific encephalopathy that is caused by toxic accumulation of lysine degradation intermediates. Substrate reduction through inhibition of DHTKD1, an enzyme upstream of the defective glutaryl-CoA dehydrogenase, has been investigated as a potential therapy, but revealed the existence of an alternative enzymatic source of glutaryl-CoA. Here, we show that loss of DHTKD1 in glutaryl-CoA dehydrogenase-deficient HEK-293 cells leads to a 2-fold decrease in the established GA1 clinical biomarker glutarylcarnitine and demonstrate that oxoglutarate dehydrogenase (OGDH) is responsible for this remaining glutarylcarnitine production. We furthermore show that DHTKD1 interacts with OGDH, dihydrolipoyl succinyltransferase and dihydrolipoamide dehydrogenase to form a hybrid 2-oxoglutaric and 2-oxoadipic acid dehydrogenase complex. In summary, 2-oxoadipic acid is a substrate for DHTKD1, but also for OGDH in a cell model system. The classical 2-oxoglutaric dehydrogenase complex can exist as a previously undiscovered hybrid containing DHTKD1 displaying improved kinetics towards 2-oxoadipic acid., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The production and secretion of matrix proteins upon stimulation of fibroblasts by transforming growth factor-beta (TGFβ) play a critical role in wound healing. How TGFβ supports the bioenergetic cost of matrix protein synthesis is not fully understood. Here, we show that TGFβ promotes protein translation at least in part by increasing the mitochondrial oxidation of glucose and glutamine carbons to support the bioenergetic demand of translation. Surprisingly, we found that in addition to stimulating the entry of glucose and glutamine carbon into the TCA cycle, TGFβ induced the biosynthesis of proline from glutamine in a Smad4-dependent fashion. Metabolic manipulations that increased mitochondrial redox generation promoted proline biosynthesis, while reducing mitochondrial redox potential and/or ATP synthesis impaired proline biosynthesis. Thus, proline biosynthesis acts as a redox vent, preventing the TGFβ-induced increase in mitochondrial glucose and glutamine catabolism from generating damaging reactive oxygen species (ROS) when TCA cycle activity exceeds the ability of oxidative phosphorylation to convert mitochondrial redox potential into ATP. In turn, the enhanced synthesis of proline supports TGFβ-induced production of matrix proteins., (© 2020 The Authors.)

Colorectal cancer (CRC) is one of the most prevalent and lethal malignancies. Especially for early stage CRC, prognostic molecular markers are needed to guide therapy. In this study, we first extracted total proteomes from matched pairs of fresh cancer and benign mucosal tissues from 22 CRC patients. Global proteomic profiling with Fourier transform liquid chromatography-mass spectrometry sequencing and label free quantitation uncovered that P4HA1 (prolyl 4-hydroxylase alpha 1) was overexpressed in CRC relative to benign colonic mucosa. We then investigated expression by immunohistochemical staining with P4HA1-specific antibodies using CRC tissue microarrays. Independent validation cohorts of 599 cases of early stage CRC and 91 cases of late stage CRC were examined. Multivariate and univariate survival analyses revealed that high expression of P4HA1 protein was an independent poor prognostic marker for patients with early stage CRC, especially of the microsatellite stable subtype. Our study provides strong support for P4HA1 as a predictive protein marker for precision diagnostics, therapeutic decision-making, and drug development for early stage colorectal cancer and demonstrates the utility of proteomic profiling to identify novel protein biomarkers., Competing Interests: CONFLICTS OF INTEREST AT, YZ, JS, FG, MO, RCH, and JYW declare no conflicts of interest related to this study. JYW is founder and equity holder of Curandis. DSK is a consultant for and equity holder in Paige.AI and a consultant for Merck. MHR is member of the Scientific Advisory Boards of Proscia and Trans-Hit. None of these companies had any influence in support, design, execution, data analysis, or any other aspect of this study., (Copyright: © 2020 Tanaka et al.)

Iron is essential for nearly all bacterial pathogens, including Mycobacterium tuberculosis (Mtb), but is severely limited in the human host. To meet its iron needs, Mtb secretes siderophores, small molecules with high affinity for iron, and takes up iron-loaded mycobactins (MBT) and carboxymycobactins (cMBT), from the environment. Mtb is also capable of utilizing heme and hemoglobin which contain more than 70% of the iron in the human body. However, many components of these iron acquisition pathways are still unknown. In this study, a high-density transposon mutagenesis coupled with deep sequencing (TnSeq) showed that Mtb exhibits nearly opposite requirements for 165 genes in the presence of heme and hemoglobin versus MBT and cMBT as iron sources. The ESX-3 secretion system was assessed as essential for siderophore-mediated iron uptake and, surprisingly, also for heme utilization by Mtb. Predictions derived from the TnSeq analysis were validated by growth experiments with isogenic Mtb mutants. These results showed that (i) the efflux pump MmpL5 plays a dominant role in siderophore secretion, (ii) the Rv2047c protein is essential for growth of Mtb in the presence of mycobactin, and (iii) the transcriptional repressor Zur is required for heme utilization by Mtb. The novel genetic determinants of iron utilization revealed in this study will stimulate further experiments in this important area of Mtb physiology., Competing Interests: The authors have declared that no competing interests exist.

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

Epithelial-to-mesenchymal transitions (EMTs) are phenotypic plasticity processes that confer migratory and invasive properties to epithelial cells during development, wound-healing, fibrosis and cancer 1-4 . EMTs are driven by SNAIL, ZEB and TWIST transcription factors 5,6 together with microRNAs that balance this regulatory network 7,8 . Transforming growth factor β (TGF-β) is a potent inducer of developmental and fibrogenic EMTs 4,9,10 . Aberrant TGF-β signalling and EMT are implicated in the pathogenesis of renal fibrosis, alcoholic liver disease, non-alcoholic steatohepatitis, pulmonary fibrosis and cancer 4,11 . TGF-β depends on RAS and mitogen-activated protein kinase (MAPK) pathway inputs for the induction of EMTs 12-19 . Here we show how these signals coordinately trigger EMTs and integrate them with broader pathophysiological processes. We identify RAS-responsive element binding protein 1 (RREB1), a RAS transcriptional effector 20,21 , as a key partner of TGF-β-activated SMAD transcription factors in EMT. MAPK-activated RREB1 recruits TGF-β-activated SMAD factors to SNAIL. Context-dependent chromatin accessibility dictates the ability of RREB1 and SMAD to activate additional genes that determine the nature of the resulting EMT. In carcinoma cells, TGF-β-SMAD and RREB1 directly drive expression of SNAIL and fibrogenic factors stimulating myofibroblasts, promoting intratumoral fibrosis and supporting tumour growth. In mouse epiblast progenitors, Nodal-SMAD and RREB1 combine to induce expression of SNAIL and mesendoderm-differentiation genes that drive gastrulation. Thus, RREB1 provides a molecular link between RAS and TGF-β pathways for coordinated induction of developmental and fibrogenic EMTs. These insights increase our understanding of the regulation of epithelial plasticity and its pathophysiological consequences in development, fibrosis and cancer.

Protein arginine methyltransferase 5 (PRMT5) has emerged as a promising cancer drug target, and three PRMT5 inhibitors are currently in clinical trials for multiple malignancies. In this study, we investigated the role of PRMT5 in human acute myeloid leukemia (AML). Using an enzymatic dead version of PRMT5 and a PRMT5-specific inhibitor, we demonstrated the requirement of the catalytic activity of PRMT5 for the survival of AML cells. We then identified PRMT5 substrates using multiplexed quantitative proteomics and investigated their role in the survival of AML cells. We found that the function of the splicing regulator SRSF1 relies on its methylation by PRMT5 and that loss of PRMT5 leads to changes in alternative splicing of multiple essential genes. Our study proposes a mechanism for the requirement of PRMT5 for leukemia cell survival and provides potential biomarkers for the treatment response to PRMT5 inhibitors.