Sexual and gender minority content in undergraduate medical education in the United States and Canada: current state and changes since 2011.

PURPOSE: To characterize current lesbian, gay, bisexual, transgender, queer, and intersex (LGBTQI +) health-related undergraduate medical education (UME) curricular content and associated changes since a 2011 study and to determine the frequency and extent of institutional instruction in 17 LGBTQI + health-related topics, strategies for increasing LGBTQI + health-related content, and faculty development opportunities. METHOD: Deans of medical education (or equivalent) at 214 allopathic or osteopathic medical schools in Canada and the United States were invited to complete a 36-question, Web-based questionnaire between June 2021 and September 2022. The main outcome measured was reported hours of LGBTQI + health-related curricular content. RESULTS: Of 214 schools, 100 (46.7%) responded, of which 85 (85.0%) fully completed the questionnaire. Compared to 5 median hours dedicated to LGBTQI + health-related in a 2011 study, the 2022 median reported time was 11 h (interquartile range [IQR], 6-16 h, p < 0.0001). Two UME institutions (2.4%; 95% CI, 0.0%-5.8%) reported 0 h during the pre-clerkship phase; 21 institutions (24.7%; CI, 15.5%-33.9%) reported 0 h during the clerkship phase; and 1 institution (1.2%; CI, 0%-3.5%) reported 0 h across the curriculum. Median US allopathic clerkship hours were significantly different from US osteopathic clerkship hours (4 h [IQR, 1-6 h] versus 0 h [IQR, 0-0 h]; p = 0.01). Suggested strategies to increase content included more curricular material focusing on LGBTQI + health and health disparities at 55 schools (64.7%; CI, 54.6%-74.9%), more faculty willing and able to teach LGBTQI + -related content at 49 schools (57.7%; CI, 47.1%-68.2%), and more evidence-based research on LGBTQI + health and health disparities at 24 schools (28.2%; CI, 18.7%-37.8%). CONCLUSION: Compared to a 2011 study, the median reported time dedicated to LGBTQI + health-related topics in 2022 increased across US and Canadian UME institutions, but the breadth, efficacy, or quality of instruction continued to vary substantially. Despite the increased hours, this still falls short of the number of hours based on recommended LGBTQI + health competencies from the Association of American Medical Colleges. While most deans of medical education reported their institutions' coverage of LGBTQI + health as 'fair,' 'good,' or 'very good,' there continues to be a call from UME leadership to increase curricular content. This requires dedicated training for faculty and students.

Water and chloride as allosteric inhibitors in WNK kinase osmosensing.

Previous observations showed that chloride and osmotic stress regulate the autophosphorylation and activity of the kinase domains of WNK1 and WNK3. Further, prior crystallography on the asymmetric dimeric of the unphosphorylated WNK1 kinase domain (WNK1/S382A, WNK1/SA) revealed conserved waters in the active site. Here we show by crystallography that PEG400 applied to crystals of dimeric WNK1/SA grown in space group P1 induces de-dimerization with a change in space group to P2 1 . Both the conserved waters, referred to here as conserved water network 1 (CWN1) and the chloride binding site are disrupted by PEG400. CWN1 is surrounded and stabilized by a pan-WNK-conserved cluster of charged residues. Here we mutagenized these charges in WNK3 to probe the importance of the CWN1 to WNK regulation. Two mutations at E314 in the Activation Loop (WNK3/E314Q and WNK3/E314A) enhanced activity, consistent with the idea that the CWN1 is inhibitory. Mutations of other residues in the cluster had similar or less activity than wild-type. PEG400 activation of WNK3 was not significantly reduced in the point mutants tested. The crystallographic and assay data support a role for CWN1 and the charged cluster in stabilizing an inactive configuration of WNKs and suggest that water functions as an allosteric inhibitor of WNKs.

Hydrostatic Pressure Sensing by WNK kinases.

Previous study has demonstrated that the WNK kinases 1 and 3 are direct osmosensors consistent with their established role in cell-volume control. WNK kinases may also be regulated by hydrostatic pressure. Hydrostatic pressure applied to cells in culture with N2 gas or to Drosophila Malpighian tubules by centrifugation induces phosphorylation of downstream effectors of endogenous WNKs. In vitro, the autophosphorylation and activity of the unphosphorylated kinase domain of WNK3 (uWNK3) is enhanced to a lesser extent than in cells by 190 kPa applied with N2 gas. Hydrostatic pressure measurably alters the structure of uWNK3. Data from size exclusion chromatography in line with multi-angle light scattering (SEC-MALS), SEC alone at different back pressures, analytical ultracentrifugation (AUC), NMR, and chemical crosslinking indicate a change in oligomeric structure in the presence of hydrostatic pressure from a WNK3 dimer to a monomer. The effects on the structure are related to those seen with osmolytes. Potential mechanisms of hydrostatic pressure activation of uWNK3 and the relationships of pressure activation to WNK osmosensing are discussed.

Case report: Persistent Müllerian duct syndrome and enlarged prostatic utricle in a male dog.

A 1-year-old male intact Miniature Schnauzer mix was presented for chronic intermittent hematuria. Abdominal ultrasonography revealed a large, fluid-filled cystic structure extending cranially and dorsally to the prostate. Computed tomography scan images revealed that the fluid-filled cavity resembled a uterus, with both horns entering the scrotum through the inguinal canal adjacent to the testes. On cytogenetic analysis, the dog was found to have a homozygote mutation on AMHRII consistent with persistent Müllerian duct syndrome (PMDS). A gonadohysterectomy was performed, and surgical and histologic findings confirmed the presence of a uterus, oviducts, vagina, and testes in this dog. Additionally, an intraoperative fluoroscopy exam revealed a communication between the uterus and the bladder via an enlarged utricle, explaining the hematuria and urine in the reproductive tract (urometra). To our knowledge, this is the first clinical report of a phenotypically intact male dog with PMDS and urometra due to an enlarged prostatic utricle. This case illustrates a combination of a disorder of sex and urogenital sinus development.

Implementation of Evidence-Based Psychotherapies for Posttraumatic Stress Disorder: A Systematic Review.

Guidelines strongly recommend trauma-focused therapies to treat posttraumatic stress disorder. Implementation of cognitive processing therapy (CPT) and prolonged exposure (PE) in Veterans Health Administration (VHA) and non-VHA settings began in 2006. We conducted a systematic review of implementation facilitators and challenges and strategies to address barriers. We searched MEDLINE, Embase, PsycINFO, and CINAHL from inception until March 2021 for English-language articles. Two individuals reviewed eligibility and rated quality. Quantitative results were abstracted by one reviewer and verified by a second. Qualitative results were independently coded by two reviewers and finalized through consensus. We used RE-AIM and CFIR frameworks to synthesize findings. 29 eligible studies addressed CPT/PE, mostly conducted in VHA. Training/education with audit/feedback was the primary implementation strategy and was linked to improved provider CPT/PE perceptions and self-efficacy. Use was not widespread. Only six studies tested other implementation strategies with mixed impact. Following VHA implementation, strong support for training, perceived effectiveness for patients and benefits for clinics, and positive patient experiences and relationships with providers were reported. However, barriers persisted including perceived protocol inflexibility, complex referral processes and patient complexity and competing needs. In non-VHA settings, providers perceived fewer barriers, but few were CPT/PE trained. Across both settings, fewer studies targeted patient factors. Training/education with audit/feedback improved perceptions and the availability of CPT/PE, but not consistent use. Studies testing implementation strategies to address post-training challenges, including patient-level factors, are needed. A few studies are underway in VHA to test patient-focused and other implementation strategies. Research assessing actual vs perceived barriers in non-VHA settings is needed to elucidate unique challenges experienced.

Evaluation of Implementation Strategies for Cognitive Behavioral Therapy (CBT), Acceptance and Commitment Therapy (ACT), and Mindfulness-Based Stress Reduction (MBSR): a Systematic Review.

BACKGROUND: Improving access to evidence-based psychotherapies (EBPs) is a Veterans Health Administration (VHA) priority. Cognitive behavioral therapy (CBT), acceptance and commitment therapy (ACT), and mindfulness-based stress reduction (MBSR) are effective for chronic pain and several mental health conditions. We synthesized evidence on implementation strategies to increase EBP access and use. METHODS: We searched MEDLINE, Embase, PsycINFO, and CINAHL from inception until March 2021 for articles on EBP implementation within integrated health systems to treat chronic pain or chronic mental health conditions. Reviewers independently screened articles, extracted results, coded qualitative findings, and rated quality using modified criteria from Newcastle-Ottawa (quantitative results) or Critical Appraisal Skills Programme (qualitative results). We categorized implementation strategies using the Expert Recommendations for Implementing Change (ERIC) framework and classified outcomes using RE-AIM domains (Reach, Effectiveness, Adoption, Implementation, Maintenance). RESULTS: Twelve articles (reporting results from 10 studies) evaluated CBT (k = 11) and ACT (k = 1) implementation strategies in large integrated healthcare systems. No studies evaluated MBSR implementation. Eight articles evaluated strategies within VHA. Six articles reported on national VHA EBP implementation programs; all involved training/education, facilitation, and audit/feedback. CBT and ACT implementation demonstrated moderate to large improvements in patient symptoms and quality of life. Trainings increased mental health provider self-efficacy in delivering EBPs, improved provider EBP perceptions, and increased provider EBP use during programs, but had unclear impacts on Reach. It was unclear whether external facilitation added benefit. Provider EBP maintenance was modest; barriers included competing professional time demands and patient barriers. DISCUSSION: Multi-faceted CBT and ACT implementation programs increased provider EBP Adoption but had unclear impacts on Reach. Future implementation efforts should further evaluate Reach, Adoption, and Maintenance; assess the added value of external facilitation; and consider strategies targeting patient barriers. Future work should use implementation frameworks to guide evaluations of barriers and facilitators, processes of change, and outcomes. REGISTRATION: PROSPERO registration number CRD42021252038.

Barriers and Facilitators of Evidence-Based Psychotherapies for Chronic Pain in Adults: A Systematic Review.

Cognitive behavioral therapy (CBT), acceptance and commitment therapy (ACT), and mindfulness-based stress reduction (MBSR) have demonstrated effectiveness for improving outcomes in chronic pain. These evidence-based psychotherapies (EBPs) remain underutilized in clinical practice, however. To identify research gaps and next steps for improving uptake of EBPs, we conducted a systematic review of patient-, provider-, and system-level barriers and facilitators of their use for chronic pain. We searched MEDLINE, Embase, PsycINFO, and CINAHL databases from inception through September 2022. Prespecified eligibility criteria included outpatient treatment of adults with chronic pain; examination of barriers and facilitators and/or evaluation of implementation strategies; conducted in the United States (US), United Kingdom (UK), Ireland, Canada or Australia; and publication in English. Two reviewers independently assessed eligibility and rated quality. We conducted a qualitative synthesis of results using a best-fit framework approach building upon domains of the Consolidated Framework for Implementation Research (CFIR). We identified 34 eligible studies (33 moderate or high quality), most (n = 28) of which addressed patient-level factors. Shared barriers across EBPs included variable patient buy-in to therapy rationale and competing responsibilities for patients; shared facilitators included positive group or patient-therapist dynamics. Most studies examining ACT and all examining MBSR assessed only group formats. No studies compared barriers, facilitators, or implementation strategies of group CBT to individual CBT, or of telehealth to in-person EBPs. Conceptual mismatches of patient knowledge and beliefs with therapy principles were largely analyzed qualitatively, and studies did not explore how these mismatches were addressed to support engagement. Future research on EBPs for chronic pain in real-world practice settings is needed to explore provider and system-level barriers and facilitators, heterogeneity of effects and uptake, and both effects and uptake of EBPs delivered in various formats, including group vs individual therapy and telehealth or asynchronous digital approaches. PERSPECTIVE: This systematic review synthesizes evidence on barriers and facilitators to uptake of cognitive behavioral therapy, acceptance and commitment therapy, and mindfulness-based stress reduction for chronic pain. Findings can guide future implementation work to increase availability and use of evidence-based psychotherapies for treatment of chronic pain. REGISTRATION: PROSPERO number CRD42021252038.

Opioid-related Mortality in United States Death Certificate Data: A Quantitative Bias Analysis With Expert Elicitation of Bias Parameters.

BACKGROUND: Opioid-related mortality is an important public health problem in the United States. Incidence estimates rely on death certificate data generated by health care providers and medical examiners. Opioid overdoses may be underreported when other causes of death appear plausible. We applied physician-elicited death certificate bias parameters to quantitative bias analyses assessing potential age-related differential misclassification in US opioid-related mortality estimates. METHODS: We obtained cause-of-death data (US, 2017) from the National Center for Health Statistics and calculated crude opioid-related outpatient death counts by age category (25-54, 55-64, 65+). We elicited beliefs from 10 primary care physicians on sensitivity of opioid-related death classification from death certificates. We summarized elicited sensitivity estimates, calculated plausible specificity values, and applied resulting parameters in a probabilistic bias analysis. RESULTS: Physicians estimated wide sensitivity ranges for classification of opioid-related mortality by death certificates, with lower estimated sensitivities among older age groups. Probabilistic bias analyses adjusting for physician-estimated misclassification indicated 3.1 times more (95% uncertainty interval: 1.2-23.5) opioid-related deaths than the observed death count in the 65+ age group. All age groups had substantial increases in bias-adjusted death counts. CONCLUSIONS: We developed and implemented a feasible method of eliciting physician expert opinion on bias parameters for sensitivity of a medical record-based death indicator and applied findings in quantitative bias analyses adjusting for differential misclassification. Our findings are consistent with the hypothesis that opioid-related mortality rates may be substantially underestimated, particularly among older adults, due to misclassification in cause-of-death data from death certificates.

Identification of a Class of WNK Isoform-Specific Inhibitors Through High-Throughput Screening.

Introduction: WNK [with no lysine (K)] kinases are serine/threonine kinases associated with familial hyperkalemic hypertension (FHHt). WNKs are therapeutic targets for blood pressure regulation, stroke and several cancers including triple negative breast cancer and glioblastoma. Here, we searched for and characterized novel WNK kinase inhibitors. Methods: We used a ~210,000-compound library in a high-throughput screen, re-acquisition and assay, commercial specificity screens and crystallography to identify WNK-isoform-selective inhibitors. Results: We identified five classes of compounds that inhibit the kinase activity of WNK1: quinoline compounds, halo-sulfones, cyclopropane-containing thiazoles, piperazine-containing compounds, and nitrophenol-derived compounds. The compounds are strongly pan-WNK selective, inhibiting all four WNK isoforms. A class of quinoline compounds was identified that further shows selectivity among the WNK isoforms, being more potent toward WNK3 than WNK1. The crystal structure of the quinoline-derived SW120619 bound to the kinase domain of WNK3 reveals active site binding, and comparison to the WNK1 structure reveals the potential origin of isoform specificity. Discussion: The newly discovered classes of compounds may be starting points for generating pharmacological tools and potential drugs treating hypertension and cancer.

ABORTION AND NEONATAL MORTALITY DUE TO TOXOPLASMA GONDII IN BIGHORN SHEEP (OVIS CANADENSIS).

Low lamb recruitment can be an obstacle to bighorn sheep (Ovis canadensis) conservation and restoration. Causes of abortion and neonate loss in bighorn sheep, which may affect recruitment, are poorly understood. Toxoplasma gondii is a major cause of abortion and stillbirth in domestic small ruminants worldwide, but no reports exist documenting abortion or neonatal death in bighorn sheep attributable to toxoplasmosis. Between March 2019 and May 2021, eight fetal and neonatal bighorn lamb cadavers from four western US states (Idaho, Montana, Nebraska, and Washington) were submitted to the Washington Animal Disease Diagnostic Laboratory for postmortem examination, histologic examination, and ancillary testing to determine the cause of abortion or neonatal death. Necrotizing encephalitis characteristic of toxoplasmosis was identified histologically in six of eight cases, and T. gondii infection was confirmed by PCR in five cases with characteristic lesions. Other lesions attributable to toxoplasmosis were pneumonia (3/5 cases) and myocarditis (2/5 cases). Protozoal cysts were identified histologically within brain, lung, heart, skeletal muscle, adipose tissue, or a combination of samples in all five sheep with PCR-confirmed T. gondii infections. Seroprevalence of T. gondii ranged from 40-81% of adult females sampled in the Washington population in October and November 2018-2021, confirming high rates of exposure before detection of Toxoplasma abortions in this study. Of 1,149 bighorn sheep postmortem samples submitted to Washington Animal Disease Diagnostic Laboratory between January 2000 and May 2021, 21 of which were from fetuses or neonates, a single case of chronic toxoplasmosis was diagnosed in one adult ewe. Recent identification of Toxoplasma abortions in bighorn sheep suggests that toxoplasmosis is an underappreciated cause of reproductive loss. Abortions and neonatal mortalities should be investigated through postmortem and histologic examination, particularly in herds that are chronically small, demographically stagnant, or exhibit reproductive rates lower than expected.

Intracellular Ion Control of WNK Signaling.

The with no lysine (K) (WNK) kinases are an evolutionarily ancient group of kinases with atypical placement of the catalytic lysine and diverse physiological roles. Recent studies have shown that WNKs are directly regulated by chloride, potassium, and osmotic pressure. Here, we review the discovery of WNKs as chloride-sensitive kinases and discuss physiological contexts in which chloride regulation of WNKs has been demonstrated. These include the kidney, pancreatic duct, neurons, and inflammatory cells. We discuss the interdependent relationship of osmotic pressure and intracellular chloride in cell volume regulation. We review the recent demonstration of potassium regulation of WNKs and speculate on possible physiological roles. Finally, structural and mechanistic aspects of intracellular ion and osmotic pressure regulation of WNKs are discussed.

An Analysis of the Role of Mental Health in a Randomized Trial of a Walking Intervention for Black Veterans With Chronic Pain.

Black patients and those with co-occurring mental health disorders are disproportionately affected by chronic pain, but few interventions target these populations. This is a secondary analysis of a randomized trial of a walking-focused proactive counseling intervention for Black Veterans with chronic musculoskeletal pain (ACTION). The primary aim was to examine intervention effectiveness among Veterans with an electronic health record-documented mental health diagnosis [depressive disorder, anxiety disorder, substance use disorder, post-traumatic stress disorder or serious mental illness (n = 205)] and those without a diagnosis (n = 175). About 380 Black Veterans receiving care at the Atlanta VA Health Care System were enrolled from 2016 to 2019 and randomized to the intervention or usual care (UC) (1:1). The intervention featured 6 telephone coaching sessions over 8-14 weeks to encourage walking. Participants with a mental health disorder were more likely to complete all counseling sessions (56% vs 38%) and reported improvements in global perceptions of pain and pain intensity/interference (secondary outcomes) at 3-months vs UC. Among participants without a mental health disorder, the intervention was associated with an improvement in pain-related disability at 6-months (primary outcome). Black chronic pain patients with co-occurring mental health disorders may require more intensive treatment to affect improvement in pain-related disability. PERSPECTIVE: This study examines the effectiveness of a walking intervention for chronic pain among Black Veterans with a mental health disorder. These patients were more engaged with the intervention than those without a mental health disorder. However, they did not experience reductions in pain-related disability, suggesting more intensive treatment is needed.

Synthesis and Structural Characterization of Novel Trihalo-sulfone Inhibitors of WNK1.

With No lysine (K) [WNK] kinases are structurally unique serine/threonine protein kinases that have therapeutic potential for blood pressure regulation and cancer. A novel class of trihalo-sulfone compounds was identified by high-throughput screening. Trihalo-sulfone 1 emerged as an effective inhibitor of WNK1 with an IC50 value of 1.6 µM. Herein, we define chemical features necessary for inhibition of WNK1 using chemical synthesis and X-ray crystallography. Analogues that probed the role of specific functional groups to the inhibitory activity were synthesized. X-ray structures of trihalo-sulfone 1 and a second trihalo-sulfone 23 bound to WNK1 revealed active site binding to two of the three previously defined canonical inhibitor binding pockets as well as a novel binding site for the trihalo-sulfone moiety. The elucidation of these novel interaction sites may allow for the strategic design of even more selective and potent WNK inhibitors.

Comparative Effectiveness of Opioid Tapering or Abrupt Discontinuation vs No Dosage Change for Opioid Overdose or Suicide for Patients Receiving Stable Long-term Opioid Therapy.

Importance: Opioid dosage tapering has emerged as a strategy to reduce harms associated with long-term opioid therapy; however, evidence supporting this approach is limited. Objective: To identify the association of opioid tapering or abrupt discontinuation with opioid overdose and suicide events among patients receiving stable long-term opioid therapy without evidence of opioid misuse. Design, Setting, and Participants: This comparative effectiveness study with a trial emulation approach used a large US claims data set of individuals with commercial insurance or Medicare Advantage who were aged 18 years or older and receiving stable long-term opioid therapy without evidence of opioid misuse between January 1, 2010, and December 31, 2018. Statistical analysis was performed from January 17, 2020, through November 12, 2021. Interventions: Three opioid dosage strategies: stable dosage, tapering (dosage reduction ≥15%), or abrupt discontinuation. Main Outcomes and Measures: Time to opioid overdose or suicide event identified from International Classification of Diseases, Ninth Revision and International Statistical Classification of Diseases and Related Health Problems, Tenth Revision diagnosis codes in medical claims over 11 months of follow-up. Inverse probability weighting was used to adjust for baseline confounders. The primary analysis used an intention-to-treat approach; follow-up after assignment regardless of changes in opioid dose was included. A per-protocol analysis was also conducted, in which episodes were censored for lack of adherence to assigned treatment. Results: A cohort of 199 836 individuals (45.1% men; mean [SD] age, 56.9 [12.4] years; and 57.6% aged 45-64 years) had 415 123 qualifying, long-term opioid therapy episodes; 87.1% of episodes were considered stable, 11.1% were considered a taper, and 1.8% were considered abrupt discontinuation. The adjusted cumulative incidence of opioid overdose or suicide events 11 months after baseline was 0.96% (95% CI, 0.92%-0.99%) with a stable dosage strategy, 1.10% (95% CI, 0.99%-1.22%) with a tapered dosage strategy, and 1.28% (95% CI, 0.93%-1.38%) with an abrupt discontinuation strategy. The risk difference between a taper and a stable dosage was 0.15% (95% CI, 0.03%-0.26%), and the risk difference between abrupt discontinuation and a stable dosage was 0.33% (95% CI, -0.03% to 0.74%). Results were similar using the per-protocol approach. Conclusions and Relevance: This study identified a small absolute increase in risk of harms associated with opioid tapering compared with a stable opioid dosage. These results do not suggest that policies of mandatory dosage tapering for individuals receiving a stable long-term opioid dosage without evidence of opioid misuse will reduce short-term harm via suicide and overdose.

WNK1 collaborates with TGF-ß in endothelial cell junction turnover and angiogenesis.

Angiogenesis is essential for growth of new blood vessels, remodeling existing vessels, and repair of damaged vessels, and these require reorganization of endothelial cell-cell junctions through a partial endothelial-mesenchymal transition. Homozygous disruption of the gene encoding the protein kinase WNK1 results in lethality in mice near embryonic day (E) 12 due to impaired angiogenesis. This angiogenesis defect can be rescued by endothelial-specific expression of an activated form of the WNK1 substrate kinase OSR1. We show that inhibition of WNK1 kinase activity not only prevents sprouting of endothelial cells from aortic slices but also vessel extension in inhibitor-treated embryos ex vivo. Mutations affecting TGF-ß signaling also result in abnormal vascular development beginning by E10 and, ultimately, embryonic lethality. Previously, we demonstrated cross-talk of WNK1 with TGF-ß-regulated SMAD signaling, and OSR1 was identified as a component of the TGF-ß interactome. However, molecular events jointly regulated by TGF-ß and WNK1/OSR1 have not been delineated. Here, we show that inhibition of WNK1 promotes TGF-ß-dependent degradation of the tyrosine kinase receptor AXL, which is involved in TGF-ß-mediated cell migration and angiogenesis. We also show that interaction between OSR1 and occludin, a protein associated with endothelial tight junctions, is an essential step to enable tight junction turnover. Furthermore, we show that these phenomena are WNK1 dependent, and sensitive to TGF-ß. These findings demonstrate intimate connections between WNK1/OSR1 and multiple TGF-ß-sensitive molecules controlling angiogenesis and suggest that WNK1 may modulate many TGF-ß-regulated functions.

Nuclear speckle integrity and function require TAO2 kinase.

Nuclear speckles are non-membrane-bound organelles known as storage sites for messenger RNA (mRNA) processing and splicing factors. More recently, nuclear speckles have also been implicated in splicing and export of a subset of mRNAs, including the influenza virus M mRNA that encodes proteins required for viral entry, trafficking, and budding. However, little is known about how nuclear speckles are assembled or regulated. Here, we uncovered a role for the cellular protein kinase TAO2 as a constituent of nuclear speckles and as a factor required for the integrity of these nuclear bodies and for their functions in pre-mRNA splicing and trafficking. We found that a nuclear pool of TAO2 is localized at nuclear speckles and interacts with nuclear speckle factors involved in RNA splicing and nuclear export, including SRSF1 and Aly/Ref. Depletion of TAO2 or inhibition of its kinase activity disrupts nuclear speckle structure, decreasing the levels of several proteins involved in nuclear speckle assembly and splicing, including SC35 and SON. Consequently, splicing and nuclear export of influenza virus M mRNA were severely compromised and caused a disruption in the virus life cycle. In fact, low levels of TAO2 led to a decrease in viral protein levels and inhibited viral replication. Additionally, depletion or inhibition of TAO2 resulted in abnormal expression of a subset of mRNAs with key roles in viral replication and immunity. Together, these findings uncovered a function of TAO2 in nuclear speckle formation and function and revealed host requirements and vulnerabilities for influenza infection.

WNK1 Enhances Migration and Invasion in Breast Cancer Models.

Metastasis is the major cause of mortality in patients with breast cancer. Many signaling pathways have been linked to cancer invasiveness, but blockade of few protein components has succeeded in reducing metastasis. Thus, identification of proteins contributing to invasion that are manipulable by small molecules may be valuable in inhibiting spread of the disease. The protein kinase with no lysine (K) 1 (WNK1) has been suggested to induce migration of cells representing a range of cancer types. Analyses of mouse models and patient data have implicated WNK1 as one of a handful of genes uniquely linked to invasive breast cancer. Here, we present evidence that inhibition of WNK1 slows breast cancer metastasis. We show that depletion or inhibition of WNK1 reduces migration of several breast cancer cell lines in wound healing assays and decreases invasion in collagen matrices. Furthermore, WNK1 depletion suppresses expression of AXL, a tyrosine kinase implicated in metastasis. Finally, we demonstrate that WNK inhibition in mice attenuates tumor progression and metastatic burden. These data showing reduced migration, invasion, and metastasis upon WNK1 depletion in multiple breast cancer models suggest that WNK1 contributes to the metastatic phenotype, and that WNK1 inhibition may offer a therapeutic avenue for attenuating progression of invasive breast cancers.