Uncovering nuclear pore complexity with innovation.

Advances in imaging and reductionist approaches have provided a high-resolution understanding of nuclear pore complex structure and transport, revealing unexpected mechanistic complexities based on nucleoporin functions and specialized import and export pathways.

The In Vivo and In Vitro Architecture of the Hepatitis C Virus RNA Genome Uncovers Functional RNA Secondary and Tertiary Structures.

Hepatitis C virus (HCV) is a positive-strand RNA virus that remains one of the main contributors to chronic liver disease worldwide. Studies over the last 30 years have demonstrated that HCV contains a highly structured RNA genome and many of these structures play essential roles in the HCV life cycle. Despite the importance of riboregulation in this virus, most of the HCV RNA genome remains functionally unstudied. Here, we report a complete secondary structure map of the HCV RNA genome in vivo, which was studied in parallel with the secondary structure of the same RNA obtained in vitro. Our results show that HCV is folded extensively in the cellular context. By performing comprehensive structural analyses on both in vivo data and in vitro data, we identify compact and conserved secondary and tertiary structures throughout the genome. Genetic and evolutionary functional analyses demonstrate that many of these elements play important roles in the virus life cycle. In addition to providing a comprehensive map of RNA structures and riboregulatory elements in HCV, this work provides a resource for future studies aimed at identifying therapeutic targets and conducting further mechanistic studies on this important human pathogen. IMPORTANCE HCV has one of the most highly structured RNA genomes studied to date, and it is a valuable model system for studying the role of RNA structure in protein-coding genes. While previous studies have identified individual cases of regulatory RNA structures within the HCV genome, the full-length structure of the HCV genome has not been determined in vivo. Here, we present the complete secondary structure map of HCV determined both in cells and from corresponding transcripts generated in vitro. In addition to providing a comprehensive atlas of functional secondary structural elements throughout the genomic RNA, we identified a novel set of tertiary interactions and demonstrated their functional importance. In terms of broader implications, the pipeline developed in this study can be applied to other long RNAs, such as long noncoding RNAs. In addition, the RNA structural motifs characterized in this study broaden the repertoire of known riboregulatory elements.

Improving the rigor and utility of botanical toxicity studies: Recommended resources.

Interest in botanicals, particularly as dietary supplement ingredients, is growing steadily. This growth, and the marketing of new ingredients and combination products as botanical dietary supplements, underscores the public health need for a better understanding of potential toxicities associated with use of these products. This article and accompanying template outline the resources to collect literature and relevant information to support the design of botanical toxicity studies. These resources provide critical information related to botanical identification, characterization, pre-clinical and clinical data, including adverse effects and interactions with pharmaceuticals. Toxicologists using these resources should collaborate with pharmacognosists and/or analytical chemists to enhance knowledge of the botanical material being tested. Overall, this guide and resource list is meant to help locate relevant information that can be leveraged to inform on decisions related to toxicity testing of botanicals, including the design of higher quality toxicological studies.

Dbp5 associates with RNA-bound Mex67 and Nab2 and its localization at the nuclear pore complex is sufficient for mRNP export and cell viability.

In Saccharomyces cerevisiae, the mRNA export receptor Mex67 is recruited to mature nuclear transcripts to mediate mRNA export through the nuclear pore complex (NPC) to the cytoplasm. Mex67 binds transcripts through adaptor proteins such as the poly(A) binding protein Nab2. When a transcript reaches the cytoplasmic face of the NPC, the DEAD-box protein Dbp5 acts to induce a local structural change to release Nab2 and Mex67 in an essential process termed mRNP remodeling. It is unknown how certain proteins (Nab2, Mex67) are released during Dbp5-mediated mRNP remodeling, whereas others remain associated. Here, we demonstrate that Dbp5 associates in close proximity with Mex67 and Nab2 in a cellular complex. Further, fusion of Dbp5 to Nup159 anchors Dbp5 at the cytoplasmic face of the NPC and is sufficient for cell viability. Thus, we speculate that the essential role of Dbp5 in remodeling exporting mRNPs requires its localization to the NPC and is separable from other subcellular functions of Dbp5. This work supports a model where the diverse nuclear, cytoplasmic and NPC functions of Dbp5 in the mRNA lifecycle are not interdependent and that Dbp5 is locally recruited through complex protein-protein interactions to select regions of transcripts for specific removal of transport proteins at the NPC.

Macrophages in ovarian cancer and their interactions with monoclonal antibody therapies.

The unmet clinical need for effective treatments in ovarian cancer has yet to be addressed using monoclonal antibodies (mAbs), which have largely failed to overcome tumour-associated immunosuppression, restrict cancer growth, and significantly improve survival. In recent years, experimental mAb design has moved away from solely targeting ovarian tumours and instead sought to modulate the wider tumour microenvironment (TME). Tumour-associated macrophages (TAMs) may represent an attractive therapeutic target for mAbs in ovarian cancer due to their high abundance and close proximity to tumour cells and their active involvement in facilitating several pro-tumoural processes. Moreover, the expression of several antibody crystallisable fragment (Fc) receptors and broad phenotypic plasticity of TAMs provide opportunities to modulate TAM polarisation using mAbs to promote anti-tumoural phenotypes. In this review, we discuss the role of TAMs in ovarian cancer TME and the emerging strategies to target the contributions of these cells in tumour progression through the rationale design of mAbs.

Depressive symptoms improve over 2 years of type 2 diabetes treatment via a digital continuous remote care intervention focused on carbohydrate restriction.

Depressive symptoms are prevalent among people with type 2 diabetes (T2D) and, even at low severity levels, are associated with worse diabetes outcomes. Carbohydrate restriction is an effective treatment for T2D but its long-term impacts on depressive symptoms are unclear. In the current study we explored changes in depressive symptoms over 2 years among 262 primarily non-depressed T2D patients participating in a continuous remote care intervention emphasizing carbohydrate restriction. Subclinical depressive symptoms decreased over the first 10 weeks and reductions were maintained out to 2 years. Increased frequency of blood ketone levels indicative of adherence to low carbohydrate eating predicted decreases in depressive symptoms. Concerns have been raised with recommending restrictive diets due to potential negative impacts on quality-of-life factors such as mood; however, results of the current study support positive rather than negative long-term impacts of closely monitored carbohydrate restriction on depressive symptoms.

Continuous care intervention with carbohydrate restriction improves physical function of the knees among patients with type 2 diabetes: a non-randomized study.

BACKGROUND: In a previous study, we assessed a novel, remotely monitored carbohydrate restricted diet regimen including nutritional ketosis in patients with type 2 diabetes and reported significant improvements in weight, glycemic control, abdominal fat and inflammation from baseline to 2 years. Knee outcome measures were collected as a secondary outcome in the trial. This study aims to assess the effect of this intervention on knee functional scores and to identify if changes in weight, central abdominal fat (CAF), glycemic status and high sensitivity C-reactive protein (hsCRP) were associated with its improvement. METHODS: This prospective analysis included continuous care intervention (CCI, n = 173) and usual care (UC, n = 69) trial participants with type 2 diabetes that reported knee pain at baseline. Knee outcome measures included the Knee injury and Osteoarthritis Outcome Score (KOOS) pain, symptoms, activities of daily living (ADL), sports and recreation function, and knee-related quality of life subscales, and total KOOS score were assessed from baseline to 2 years. Missing data at each time point were replaced with multiple imputation under the assumption of missing at random. To assess if the primary analysis of the knee scores changed under plausible missing not at random assumptions, sensitivity analysis was also performed using pattern mixture models. In CCI, we also assessed factors associated with the improvement of knee scores. RESULTS: In the primary analysis, CCI participants demonstrated a statistically significant improvement in total KOOS and all KOOS individual subscale scores at 1 year and maintained through 2 years as opposed to UC patients who showed no significant changes from baseline to 2 years. The significant improvement in total KOOS and its individual subscale scores from baseline to 2 years remained relatively stable in CCI in the sensitivity analysis under different missing not at random scenarios confirming the robustness of the findings from the primary analysis. Approximately 46% of the CCI participants met the 10 points minimal clinically important change at 2 years. A reduction in CAF was associated with improvement in total KOOS and KOOS ADL, while a decrease in hsCRP was associated with improvement in KOOS symptoms scores. CONCLUSION: A very low carbohydrate intervention including nutritional ketosis resulted in significant improvements in knee pain and function among patients with T2D. The improvements in knee function were likely secondary to a reduction in central adiposity and inflammation. Future research on the applicability of this intervention in radiographically confirmed OA patients is important. TRIAL REGISTRATION: Clinical trial registration: NCT02519309 (10/08/2015).

Small molecules that target group II introns are potent antifungal agents.

Specific RNA structures control numerous metabolic processes that impact human health, and yet efforts to target RNA structures de novo have been limited. In eukaryotes, the self-splicing group II intron is a mitochondrial RNA tertiary structure that is absent in vertebrates but essential for respiration in plants, fungi and yeast. Here we show that this RNA can be targeted through a process of high-throughput in vitro screening, SAR and lead optimization, resulting in high-affinity compounds that specifically inhibit group IIB intron splicing in vitro and in vivo and lack toxicity in human cells. The compounds are potent growth inhibitors of the pathogen Candida parapsilosis, displaying antifungal activity comparable to that of amphotericin B. These studies demonstrate that RNA tertiary structures can be successfully targeted de novo, resulting in pharmacologically valuable compounds.

Depression in context: Important considerations for youth with type 1 vs type 2 diabetes.

BACKGROUND: Youth with diabetes are at increased risk for depression. However, severity and correlates of depressive symptoms may differ by diabetes type. OBJECTIVE: Associations of depressive symptoms with global health, diabetes duration, and gender were compared between youth with type 1 and type 2 diabetes. METHODS: A sample of 149 youth ages 12 to 21 diagnosed with either type 1 (n = 122) or type 2 (n = 27) diabetes were screened during routine clinic appointments. Regression models were constructed to examine differences by diabetes type. RESULTS: Adolescents with type 2 diabetes had significantly higher depressive symptom scores (4.89 vs 2.99, P = .025) than those with type 1 diabetes. A significant interaction between global health and diabetes type on depressive symptoms revealed inverse associations between global health and depressive symptoms that was stronger among youth with type 2 diabetes (ß = -.98, P < .001) than type 1 (ß = -.48, P < .001). Further probing revealed that among youth with better global health, adolescents with type 1 had more depressive symptoms than those with type 2 diabetes (ß = .33, P = .035). Diabetes duration and depressive symptoms were positively associated among individuals with type 2 (ß = .86, P = .043), but not type 1 diabetes. No gender differences were detected. CONCLUSION: These findings suggest that correlates of depressive symptoms in youth with diabetes differ by diabetes type. Global health appears to be an important correlate among youth with both types, whereas diabetes duration was only a significant factor among those with type 2 diabetes. The current findings can inform future psychosocial intervention efforts within both these populations.

Diabetes-Specific Self-Compassion: A New Measure for Parents of Youth With Type 1 Diabetes.

OBJECTIVE: Given the high daily demands of managing type 1 diabetes (T1D), parents of youth with T1D can experience high levels of emotional distress, burden, and self-criticism, with implications for parent and child well-being and parent self-efficacy for managing diabetes. Diabetes-specific self-compassion (SC), or being kind to oneself when facing challenges related to managing diabetes, may serve as protective for parents. This study aimed to create and assess the psychometric properties of a new tool, the diabetes-specific Self-Compassion Scale (SCS-Dp), to assess diabetes-specific SC in parents of youth with T1D. METHODS: We adapted a parent diabetes-specific SC measure; surveyed parents (N = 198; parent: 88% female; 95% non-Hispanic White; M age = 44 ± 8.9; child: 46% female; M age = 13 ± 3.4, range 2-18 years; 83% insulin pump users; 40% continuous glucose monitor (CGM) users; HbA1c from clinic data available for 76 participants: M HbA1c = 8.1 ± 1.3%) and conducted confirmatory factor analysis, and reliability and construct validity analyses. Validity measures included diabetes distress, diabetes empowerment, diabetes numeracy, and HbA1c. RESULTS: A bifactor structure provided the best fit, with one general factor and two wording-related group factors (positively and negatively worded items). The final 19-item SCS-Dp demonstrated excellent internal consistency (α =.94; range of item-total correlations: .52-.81) and good construct validity. As predicted, greater SC was associated with lower distress (r = -.68, p < .001) and greater empowerment (r = .43, p < .001) and was not associated with diabetes numeracy (p = .61). Diabetes-specific Self-Compassion Scale was not associated with HbA1c (p = .28). CONCLUSIONS: Results provide initial evidence of good reliability and validity of the SCS-Dp to assess diabetes-specific SC in parents.

Nup42 and IP6 coordinate Gle1 stimulation of Dbp5/DDX19B for mRNA export in yeast and human cells.

The mRNA lifecycle is driven through spatiotemporal changes in the protein composition of mRNA particles (mRNPs) that are triggered by RNA-dependent DEAD-box protein (Dbp) ATPases. As mRNPs exit the nuclear pore complex (NPC) in Saccharomyces cerevisiae, this remodeling occurs through activation of Dbp5 by inositol hexakisphosphate (IP6 )-bound Gle1. At the NPC, Gle1 also binds Nup42, but Nup42's molecular function is unclear. Here we employ the power of structure-function analysis in S. cerevisiae and human (h) cells, and find that the high-affinity Nup42-Gle1 interaction is integral to Dbp5 (hDDX19B) activation and efficient mRNA export. The Nup42 carboxy-terminal domain (CTD) binds Gle1/hGle1B at an interface distinct from the Gle1-Dbp5/hDDX19B interaction site. A nup42-CTD/gle1-CTD/Dbp5 trimeric complex forms in the presence of IP6 . Deletion of NUP42 abrogates Gle1-Dbp5 interaction, and disruption of the Nup42 or IP6 binding interfaces on Gle1/hGle1B leads to defective mRNA export in S. cerevisiae and human cells. In vitro, Nup42-CTD and IP6 stimulate Gle1/hGle1B activation of Dbp5 and DDX19B recombinant proteins in similar, nonadditive manners, demonstrating complete functional conservation between humans and S. cerevisiae. Together, a highly conserved mechanism governs spatial coordination of mRNP remodeling during export. This has implications for understanding human disease mutations that perturb the Nup42-hGle1B interaction.

The epidemiology of amyloidosis in Queensland, Australia.

Published studies on the epidemiology of amyloidosis have relied on death certificate data for case ascertainment. We estimated the incidence and mortality burden of amyloidosis among residents of the Australian state, Queensland, aged ≥20 years for the years 1999-2013 based on case ascertainment from histopathology reports. Information systems for participating laboratories were scrutinised to identify histopathology reports that documented a diagnosis of amyloidosis. Case mortality status was determined via linkage to the National Death Index. A total of 447 cases of amyloidosis were identified, with a median age at diagnosis of 66 years. A plasma cell dyscrasia was identified in 72% of patients who had paraprotein studies performed. The estimated incidence for Queenslanders aged ≥20 years was 12·1 cases per million person years. The median survival was 2·45 years. Age at diagnosis, presence of a paraprotein, earlier year of diagnosis, and inner regional location of residence (compared with residence in a major city) were independently associated with reduced survival. Our data confirms previously reported incidence data for amyloidosis of approximately 10 cases per million patient years and indicates that survival for Queensland patients with amyloidosis is improving, though it remains poor for the elderly and patients with AL amyloidosis.

NS3 from Hepatitis C Virus Strain JFH-1 Is an Unusually Robust Helicase That Is Primed To Bind and Unwind Viral RNA.

Hepatitis C viruses (HCV) encode a helicase enzyme that is essential for viral replication and assembly (nonstructural protein 3 [NS3]). This helicase has become the focus of extensive basic research on the general helicase mechanism, and it is also of interest as a novel drug target. Despite the importance of this protein, mechanistic work on NS3 has been conducted almost exclusively on variants from HCV genotype 1. Our understanding of NS3 from the highly active HCV strains that are used to study HCV genetics and mechanism in cell culture (such as JFH-1) is lacking. We therefore set out to determine whether NS3 from the replicatively efficient genotype 2a strain JFH-1 displays novel functional or structural properties. Using biochemical assays for RNA binding and duplex unwinding, we show that JFH-1 NS3 binds RNA much more rapidly than the previously studied NS3 variants from genotype 1b. Unlike NS3 variants from other genotypes, JFH-1 NS3 binds RNA with high affinity in a functionally active form that is capable of immediately unwinding RNA duplexes without undergoing rate-limiting conformational changes that precede activation. Unlike other superfamily 2 (SF2) helicases, JFH-1 NS3 does not require long 3' overhangs, and it unwinds duplexes that are flanked by only a few nucleotides, as in the folded HCV genome. To understand the physical basis for this, we solved the crystal structure of JFH-1 NS3, revealing a novel conformation that contains an open, positively charged RNA binding cleft that is primed for productive interaction with RNA targets, potentially explaining robust replication by HCV JFH-1.IMPORTANCE Genotypes of HCV are as divergent as different types of flavivirus, and yet mechanistic features of HCV variants are presumed to be held in common. One of the most well-studied components of the HCV replication complex is a helicase known as nonstructural protein 3 (NS3). We set out to determine whether this important mechanical component possesses biochemical and structural properties that differ between common strains such as those of genotype 1b and a strain of HCV that replicates with exceptional efficiency (JFH-1, classified as genotype 2a). Indeed, unlike the inefficient genotype 1b NS3, which has been well studied, JFH-1 NS3 is a superhelicase with strong RNA affinity and high unwinding efficiency on a broad range of targets. Crystallographic analysis reveals architectural features that promote enhanced biochemical activity of JFH-1 NS3. These findings show that even within a single family of viruses, drift in sequence can result in the acquisition of radically new functional properties that enhance viral fitness.

Cancer-related loneliness mediates the relationships between social constraints and symptoms among cancer patients.

Cancer patients have high rates of persistent and disabling symptoms. Evidence suggests that social constraints (e.g., avoidance and criticism) negatively impact symptoms, but pathways linking these variables have yet to be identified. This study examined whether cancer-related loneliness (i.e., feeling socially disconnected related to having cancer) mediated the relationships between social constraints and symptoms (i.e., pain interference, fatigue, sleep disturbance, and cognitive complaints) in patients with various cancers (N = 182). Patients (51% female, mean age = 59) were recruited from the Indiana Cancer Registry and completed questionnaires assessing social constraints, cancer-related loneliness, and symptoms. Structural equation modeling was used to evaluate the hypothesized relationships among variables. The model demonstrated good fit. Consistent with our hypothesis, cancer-related loneliness mediated the relationships between social constraints and each symptom. Findings suggest that addressing cancer-related loneliness in symptom management interventions may mitigate the negative impact of social constraints on outcomes.

Predictability of Interruptions During Medication Administration With Related Behavioral Management Strategies.

The aim of this qualitative study was to examine the nature of interruptions during medication administration. Focus groups were conducted with medical/surgical nurses (n = 15), critical care nurses (n = 13), and nurse managers/educators/specialists (n = 6). Most interruptions (78%) were predictable. Nurse-adopted strategies included blocking, engaging, mediating, multitasking, and preventing. Educational content was developed that relates behavioral strategies to respond to predictable and unpredictable interruptions.

Cdc42 regulates epithelial cell polarity and cytoskeletal function during kidney tubule development.

The Rho GTPase Cdc42 regulates key signaling pathways required for multiple cell functions, including maintenance of shape, polarity, proliferation, migration, differentiation and morphogenesis. Although previous studies have shown that Cdc42 is required for proper epithelial development and maintenance, its exact molecular function in kidney development is not well understood. In this study, we define the specific role of Cdc42 during murine kidney epithelial tubulogenesis by deleting it selectively at the initiation of ureteric bud or metanephric mesenchyme development. Deletion in either lineage results in abnormal tubulogenesis, with profound defects in polarity, lumen formation and the actin cytoskeleton. Ultimately, these defects lead to renal failure. Additionally, in vitro analysis of Cdc42-null collecting duct cells shows that Cdc42 controls these processes by regulating the polarity Par complex (Par3-Par6-aPKC-Cdc42) and the cytoskeletal proteins N-Wasp and ezrin. Thus, we conclude that the principal role of Cdc42 in ureteric bud and metanephric mesenchyme development is to regulate epithelial cell polarity and the actin cytoskeleton.

Avoidant coping and self-efficacy mediate relationships between perceived social constraints and symptoms among long-term breast cancer survivors.

OBJECTIVE: Many breast cancer survivors feel constrained in discussing their cancer experience with others. Limited evidence suggests that social constraints (e.g., avoidance and criticism) from loved ones may negatively impact breast cancer survivors' global health, but research has yet to examine relationships between social constraints and common physical symptoms. Informed by social cognitive processing theory, this study examined whether perceived social constraints from partners and healthcare providers (HCPs) were associated with fatigue, sleep disturbance, and attentional functioning among long-term breast cancer survivors (N = 1052). In addition, avoidant coping and self-efficacy for symptom management were examined as potential mediators of these relationships. METHODS: Long-term breast cancer survivors (mean years since diagnosis = 6) completed questionnaires assessing social constraints from partners and HCPs, avoidant coping, self-efficacy for symptom management, and symptoms (i.e., fatigue, sleep disturbance, and attentional functioning). Structural equation modeling was used to evaluate the hypothesized relationships among variables in two models: one focused on social constraints from partners and one focused on social constraints from HCPs. RESULTS: Both models demonstrated good fit. Consistent with theory and prior research, greater social constraints from both partners and HCPs were associated with greater symptom burden (i.e., greater fatigue and sleep disturbance, poorer attentional functioning). In addition, all relationships were mediated by avoidant coping and self-efficacy for symptom management. CONCLUSIONS: Findings are consistent with social cognitive processing theory and suggest that symptom management interventions may be enhanced by addressing the impact of social constraints from survivors' partners and HCPs on their coping and self-efficacy. Copyright © 2016 John Wiley & Sons, Ltd.

Long-term fear of recurrence in young breast cancer survivors and partners.

BACKGROUND: Fear of a breast cancer recurrence is the most prevalent and disruptive source of distress for long-term survivors and their partners. However, few studies have focused on predictors of fear of recurrence. The aim of this study is to test the efficacy of the Social Cognitive Processing Theory (SCPT) in predicting fear of recurrence in long-term breast cancer survivors diagnosed at age 45 years or younger and their partners. METHODS: In a large cross-sectional study, breast cancer survivors (n = 222) 3-8 years from diagnosis and their partners completed a survey assessing demographic characteristics, fear of recurrence, social constraints, and cognitive processing (intrusive thoughts and cognitive avoidance). Mediation analyses were conducted for survivors and partners separately to determine if cognitive processing would mediate the relationship between social constraints and fear of recurrence. RESULTS: Cognitive processing mediated the relationship between social constraints and fear of recurrence both for survivors [F(3,213) = 47.541, R2 = 0.401, p < 0.001] and partners [F(3,215) = 27.917, R2 = 0.280, p < 0.001). Demographic variables were not significant predictors of fear of recurrence. CONCLUSIONS: As predicted, cognitive processing mediated the relationship between social constraints and fear of recurrence. Results expand the utility of the SCPT in long-term survivors and their partners by supporting its use in intervention design. Copyright © 2015 John Wiley & Sons, Ltd.

Molecular Engineering of Near Infrared Absorbing Thienopyrazine Double Donor Double Acceptor Organic Dyes for Dye-Sensitized Solar Cells.

The thienopyrazine (TPz) building block allows for NIR photon absorption in dye-sensitized solar cells (DSCs) when used as a π-bridge. We synthesized and characterized 7 organic sensitizers employing thienopyrazine (TPz) as a π-bridge in a double donor, double acceptor organic dye design. Donor groups are varied based on electron donating strength and sterics at the donor-π bridge bond with the acceptor groups varied as either carboxylic acids or benzoic acids on the π-bridge. This dye design was found to be remarkably tunable with solution absorption onsets ranging from 750 to near 1000 nm. Interestingly, the solution absorption measurements do not accurately approximate the dye absorption on TiO2 films with up to a 250 nm blue-shift of the dye absorption onset on TiO2. This shift in absorption and the effect on electron transfer properties is investigated via computational analysis, time-correlated single photon counting studies, and transient absorption spectroscopy. Structure-performance relationships were analyzed for the dyes in DSC devices with the highest performance observed at 17.6 mA/cm2 of photocurrent and 7.5% PCE for a cosensitized device with a panchromatic IPCE onset of 800 nm.