Immigrants' Experience of Health Care Access in Canada: A Recent Scoping Review.

Existing literature has reported inequities in access to Canadian health care services among immigrants. The aim of this scoping review was (a) to explore research regarding Canadian immigrants' unique experiences in accessing healthcare, and (b) to provide suggestions for future research and programming considering the identified immigrant-specific service gaps in health care. We searched MEDLINE, CINAHL, EMBASE, and Google Scholar, following the Arksey and O'Malley (2005) framework. The review's findings suggest unmet health care access needs specific to immigrants in Canada, with the most common access barriers including communication, socioeconomic, and cultural barriers. The scoping review expands on the immigrant health care experiences and accessibility factors through a thematic analysis. Findings suggest that developing community-based programming, improving training for health care providers in culturally competent care, and policies that address the social determinants of health can improve health care accessibility among immigrants.

Dynamic quality control machinery that operates across compartmental borders mediates the degradation of mammalian nuclear membrane proteins.

Many human diseases are caused by mutations in nuclear envelope (NE) proteins. How protein homeostasis and disease etiology are interconnected at the NE is poorly understood. Specifically, the identity of local ubiquitin ligases that facilitate ubiquitin-proteasome-dependent NE protein turnover is presently unknown. Here, we employ a short-lived, Lamin B receptor disease variant as a model substrate in a genetic screen to uncover key elements of NE protein turnover. We identify the ubiquitin-conjugating enzymes (E2s) Ube2G2 and Ube2D3, the membrane-resident ubiquitin ligases (E3s) RNF5 and HRD1, and the poorly understood protein TMEM33. RNF5, but not HRD1, requires TMEM33 both for efficient biosynthesis and function. Once synthesized, RNF5 responds dynamically to increased substrate levels at the NE by departing from the endoplasmic reticulum, where HRD1 remains confined. Thus, mammalian protein quality control machinery partitions between distinct cellular compartments to address locally changing substrate loads, establishing a robust cellular quality control system.

Cranberry Ingestion Modulated Drug Transporters and Metabolizing Enzymes: Gefitinib Used as a Probe Substrate in Rats.

Cranberry, a polyphenol-rich functional food, is commonly used for the prophylaxis of urinary tract infections. Gefitinib, an anticancer agent clinically prescribed to treat non-small-cell lung cancer, is a substrate of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), and metabolized mainly by cytochrome P450 (CYP) 3A4 and CYP2D6. This study used gefitinib as a probe substrate to investigate the modulation of cranberry on P-gp, BCRP, CYP3A4 and CYP2D6. Rats were administered gefitinib with and without 5.0 g/kg of cranberry as juice (CJ). The concentration of gefitinib in serum was determined by LC-MS/MS. The results showed that CJ significantly increased the Cmax and AUC0-t of gefitinib by 28% and 55%, respectively. Mechanism studies indicated that CJ activated P-gp, and cranberry metabolites (CM) inhibited CYP2D6. Moreover, the protein level of P-gp in rat enterocytes was decreased, whereas that in hepatocytes was increased. In addition, the protein levels of BCRP, CYP3A4 and CYP2D6 in enterocytes and hepatocytes were decreased. In conclusion, CJ ingestion affected the activities and protein levels of P-gp, BCRP, CYP3A4 and CYP2D6.

Humoral, Cellular and Cytokine Immune Responses Against SARS-CoV-2 Variants in COVID-19 Convalescent and Confirmed Patients With Different Disease Severities.

Objectives: To assess humoral and cellular immune responses against SARS-CoV-2 variants in COVID-19 convalescent and confirmed patients, to explore the correlation between disease severity, humoral immunity, and cytokines/chemokines in confirmed patients, and to evaluate the ADE risk of SARS-CoV-2. Methods: Anti-RBD IgG were quantified using an ELISA. Neutralization potency was measured using pseudovirus and real virus. Cellular immunity was measured using ELISpot. Cytokine/chemokine levels were detected using multiplex immunoassays. In vitro ADE assays were performed using Raji cells. Results: One-month alpha convalescents exhibited spike-specific antibodies and T cells for alpha and delta variants. Notably, the RBD-specific IgG towards the delta variant decreased by 2.5-fold compared to the alpha variant. Besides, serum from individuals recently experienced COVID-19 showed suboptimal neutralizing activity against the delta and omicron variants. Humoral immune response, IL-6, IP-10 and MCP-1 levels were greater in patients with severe disease. Moreover, neither SARS-CoV-1 nor SARS-CoV-2 convalescent sera significantly enhanced SARS-CoV-2 pseudovirus infection. Conclusions: Significant resistance of the delta and omicron variants to the humoral immune response generated by individuals who recently experienced COVID-19. Furthermore, there was a significant correlation among disease severity, humoral immune response, and specific cytokines/chemokine levels. No evident ADE was observed for SARS-CoV-2.

Self-Esteem as a Predictor of Mental Adjustment in Patients with Breast Cancer.

This study aimed to explore the relationship between self-esteem and mental adjustment and examine the directional effects in patients with breast cancer using path modeling. This was a cross-sectional, descriptive, and correlational study. A total of 128 patients with breast cancer were selected through convenience sampling at a medical center in northern Taiwan. They completed a basic characteristics questionnaire, the Memorial Symptom Assessment Scale short form, the Rosenberg Self-Esteem Scale, and the mini-Mental Adjustment to Cancer Scale. Descriptive statistics, regression analysis, and path analysis were used to analyze the data. The results showed that higher self-esteem was associated with better mental adjustment (ß = 0.9, 95% confidence interval 0.6~1.3, p < 0.001). Age, religious beliefs, employment, cancer stage, and symptom distress were correlated with mental adjustment. Path modeling demonstrated that self-esteem, cancer stage, performance status, and symptom distress directly affected mental adjustment in patients with breast cancer. These findings suggest that health professionals should evaluate self-esteem, performance status, and symptom distress in patients with breast cancer immediately upon admission. This can facilitate early implementation of relevant nursing interventions and, consequently, improve self-esteem and symptom distress and increase mental adjustment in these patients.

Dissociative Effects of Neuropeptide S Receptor Deficiency and Nasal Neuropeptide S Administration on T-Maze Discrimination and Reversal Learning.

Cognitive flexibility refers to the ability to modify learned behavior in response to changes in the environment. In laboratory rodents, cognitive flexibility can be assessed in reversal learning, i.e., the change of contingencies, for example in T-maze discrimination learning. The present study investigated the role of the neuropeptide S (NPS) system in cognitive flexibility. In the first experiment, mice deficient of NPS receptors (NPSR) were tested in T-maze discrimination and reversal learning. In the second experiment, C57BL/6J mice were tested in the T-maze after nasal administration of NPS. Finally, the effect of nasal NPS on locomotor activity was evaluated. NPSR deficiency positively affected the acquisition of T-maze discrimination but had no effects on reversal learning. Nasal NPS administration facilitated reversal learning and supported an allocentric learning strategy without affecting acquisition of the task or locomotor activity. Taken together, the present data show that the NPS system is able to modulate both acquisition of T-maze discrimination and its reversal learning. However, NPSR deficiency only improved discrimination learning, while nasal NPS administration only improved reversal learning, i.e., cognitive flexibility. These effects, which at first glance appear to be contradictory, could be due to the different roles of the NPS system in the brain regions that are important for learning and cognitive flexibility.

Antiviral Activity of Compound L3 against Dengue and Zika Viruses In Vitro and In Vivo.

Dengue virus (DENV) and Zika virus (ZIKV) are mosquito-borne flaviviruses that cause severe illness after infection. Currently, there are no specific or effective treatments against DENV and ZIKV. Previous studies have shown that tyrosine kinase activities and signal transduction are involved in flavivirus replication, suggesting a potential therapeutic strategy for DENV and ZIKV. In this study, we found that compound L3 can significantly reduce viral protein expression and viral titers in HEK-293, MCF-7, HepG2, and Huh-7 cells and exhibits superior therapeutic efficacy against flaviviral infection compared to other tyrosine kinase inhibitors. In addition, compound L3 can decrease endogenous HER2 activation and inhibit the phosphorylation of the HER2 downstream signaling molecules Src and ERK1/2, the levels of which have been associated with viral protein expression in MCF-7 cells. Moreover, silencing HER2 diminished DENV-2 and ZIKV expression in MCF-7 cells, which suggests that HER2 activity is involved in flavivirus replication. Furthermore, in DENV-2-infected AG129 mice, treatment with compound L3 increased the survival rates and reduced the viremia levels. Overall, compound L3 demonstrates therapeutic efficacy both in vitro and in vivo and could be developed as a promising antiviral drug against emerging flaviviruses or for concurrent DENV and ZIKV outbreaks.

Hepatitis C Virus Entry: An Intriguingly Complex and Highly Regulated Process.

Hepatitis C virus (HCV) is a major cause of chronic hepatitis and liver disease worldwide. Its tissue and species tropism are largely defined by the viral entry process that is required for subsequent productive viral infection and establishment of chronic infection. This review provides an overview of the viral and host factors involved in HCV entry into hepatocytes, summarizes our understanding of the molecular mechanisms governing this process and highlights the therapeutic potential of host-targeting entry inhibitors.

Anti-inflammatory Compound Shows Therapeutic Safety and Efficacy against Flavivirus Infection.

Flaviviruses comprise several medically important viruses, including Japanese encephalitis virus, West Nile virus, dengue virus (DENV), yellow fever virus, and Zika virus (ZIKV). A large outbreak of DENV and ZIKV occurred recently, leading to many cases of illness and death. However, despite decades of effort, we have no clinically specific therapeutic drugs against DENV and ZIKV. Previous studies showed that inflammatory responses play a critical role in dengue and Zika virus pathogenesis. Thus, in this study, we examined a series of novel anti-inflammatory compounds and found that treatment with compound 2d could dose dependently reduce viral protein expression and viral progeny production in HEK-293 and Raw264.7 cells infected with four serotypes of DENV and ZIKV. In addition, considering medication safety, compound 2d could not suppress cyclooxygenase-1 (COX-1) enzymatic activities and thus could prevent the side effect of bleeding. Moreover, compound 2d significantly inhibited COX-2 enzymatic activities and prostaglandin E2 levels, associated with viral replication, compared to results with a selective COX-2 inhibitor, celecoxib. Furthermore, administering 5 mg/kg compound 2d to DENV-2-infected AG129 mice prolonged survival and reduced viremia and serum cytokine levels. Overall, compound 2d showed therapeutic safety and efficacy in vitro and in vivo and could be further developed as a potential therapeutic agent for flavivirus infection.

Methodologies to monitor protein turnover at the inner nuclear membrane.

Lamin B receptor (LBR) is an inner nuclear membrane protein that associates with the nuclear lamina and harbors sterol reductase activity essential for cholesterol biosynthesis. Several LBR mutations implicated in human congenital disorders give rise to C-terminal truncations which render LBR metabolically unstable, resulting in their rapid turnover in the nucleus. These LBR variants serve as model substrates for investigating the poorly understood protein quality control pathways in the mammalian nuclear envelope (NE). Here we describe a split-GFP-based method for tagging these model substrates to enable live cell imaging and flow cytometry for the identification and characterization of NE-resident protein turnover machinery. Furthermore, we describe a facile subcellular fractionation method to isolate a soluble LBR degradation intermediate, allowing the deconvolution of the membrane extraction and proteasomal turnover steps. The combination of imaging-based and biochemical approaches described here facilitates detailed mechanistic studies to dissect protein turnover in the nuclear compartment.

Protein deficiency after gastric bypass: The role of common limb length in revision surgery.

BACKGROUND: Bariatric surgery, especially the gastric bypass procedure, is an effective therapy for morbid obesity, but may reduce protein absorption and induce protein deficiency (PD). A recent study reported an issue about common limb length for PD. OBJECTIVE: This study aimed to examine the prevalence of PD after gastric bypass surgery and investigate the role of common limb length in PD-related revision surgery. SETTING: Hospital-based bariatric center. METHODS: From 2001 to 2016, 2397 patients with morbid obesity who underwent bariatric/metabolic surgery with 1-year follow-up were recruited. Serum albumin and total protein were measured before and 1 year after surgery. Medical records of patients who underwent revision surgery due to PD were reviewed. RESULTS: The overall prevalence of PD was .5% preoperatively. The prevalence of PD increased to 2.0% at 1 year after surgery. The incidence was highest in one-anastomosis gastric bypass (2.8%) followed by Roux-en-Y gastric bypass (1.8%). Until the end of follow-up, all 19 patients who underwent revision surgery for intractable PD had a relatively short common limb length of <400 cm. After elongation of the common limb length to >400 cm in revision surgery, PD improved in all patients. CONCLUSIONS: A subset of patients can develop PD after gastric bypass surgery when the common limb length is <400 cm. In patients with intractable PD after gastric bypass surgery, revision surgery for elongation of common limb length to >400 cm is mandatory to avoid PD-related complications.

Laparoscopic single-anastomosis duodenal-jejunal bypass with sleeve gastrectomy (SADJB-SG): Surgical risk and long-term results.

BACKGROUND: Single-anastomosis duodeno-jejunal bypass with sleeve gastrectomy (SADJB-SG) was developed as a simplified technique of DJB-SG, but long-term data are lacking. OBJECTIVE: To report the long-term data of SADJB-SG. SETTING: Tertiary Teaching Hospital. METHODS: A total of 148 SADJB-SG was performed from 2011 to 2016 with mean age of 42.0 ± 10.9-years old (14-71), female 64.9%, and mean body mass index 34.2 ± 5.9 kg/m2. All patients were evaluated and managed under a strict multidisciplinary team approach. A retrospective analysis of a prospective bariatric database and telephone interview of patients who defaulted clinic follow-up at 5-year was conducted. RESULTS: The mean operating time, intraoperative blood loss, and hospital stay of SADJB-SG were 189.6 ± 32.1 minutes, 43.5 ± 17.9 mL, and 5.0 ± 5.1 days, respectively. The 30-days postoperative major complication occurred in 7(4.7%) patients, all in patients with type 2 diabetes (T2D). At postoperative 1, 2, and 5 years, the mean percentage of total weight loss and excess weight loss of SADJB-SG patients were 25.5%, 22.8%, 22.5%, and 83.9%, 76.1%, 58.6%, respectively. Among 118 patients with T2D, 62 (52.5%) achieved complete remission (hemoglobin A1C <60%) at 1 year and 36.5% at 5 years after surgery. A total of 15 patients needed reoperation at follow-up, due to reflux disease (n = 11), weight regain, and recurrent of T2D (n = 2), ileus (n = 1), and peritonitis (n = 1). Among them, 8 were converted to RYGB and the others remained in same anatomy. At 5 years, the overall revision rate was 12.9% (8/62) and 24.5% (14/57) of the remaining required proton pump inhibitor for reflux symptoms. CONCLUSION: Our results show that primary SADJB-SG is a durable primary bariatric procedure with sustained weight loss and a high resolution of T2D at 5 years, but de novo GERD is the major side effect.

Thirteen-Year Experience of Laparoscopic Sleeve Gastrectomy: Surgical Risk, Weight Loss, and Revision Procedures.

BACKGROUND: Laparoscopic sleeve gastrectomy (LSG) has been validated as a safe and effective treatment for morbid obesity. However, data of the long-term outcome remains lacking. METHODS: A total of 1759 LSG was performed as primary bariatric procedure from 2005 to 2017 with mean age of 35.2 ± 10.3 years old (14-71), female 69.7%, mean body mass index (BMI) 37.9 ± 7.7 kg/m2, and mean waist width 113.7 ± 17.9 cm. All patients were evaluated and managed under a strict multidisciplinary team approach. A retrospective analysis of a prospective bariatric database and telephone interview of patients who defaulted clinic follow-up at 10 years was conducted. RESULTS: The mean operating time, intraoperative blood, and hospital stay of LSG were 121.5 ± 36.5 min, 40.8 ± 69.7 ml, and 2.8 ± 2.7 days, respectively. The 30-day postoperative major complication occurred in 25 (1.4%) patients. The major complication rate was 15% at first year and 0% at the last year. The follow-up rate at 1, 5 and 10 years were 89.3%, 52.1 and 64.4%. At postoperative 1, 5, and 10 years, the mean percentage of total weight loss (%TWL) and excess weight loss (EWL%) of LSG patients were 33.4, 28.3, and 26.6% and 92.2, 80.1, and 70.5%, respectively. The mean BMI became 27, 26.2, and 27.1 kg/m2 at postoperative 1, 5, and 10 years. At follow-up, a total 69 patients needed surgical revision due to reflux disease (n = 45), weight regain (n = 19), persistent diabetes (n = 2), and chronic fistula (n = 3). The type of revision procedures were hiatal repair and gastropexy (n = 29), Roux-en Y gastric bypass (RYGB) (n = 23), and single anastomosis bypass (n = 17) with median time to revision 33 months (range 3-62). At 10 years, the overall revision rate was 21.5% (14/65) and 11(16.9%) of 65 patients were converted to RYGB. The other 54 patients remained at LSG anatomy, but 45% of them required proton pump inhibitor for reflux symptoms. CONCLUSIONS: Our results showed that primary LSG is a durable primary bariatric procedure with sustained weight loss and a high resolution of comorbidities at 10 years, but about half the patients had de novo GERD. The need for conversion to RYGB was 16.9% at 10 years.

Dietary Intake and Weight Changes 5 Years After Laparoscopic Sleeve Gastrectomy.

BACKGROUND: Laparoscopic sleeve gastrectomy (LSG) is becoming a leading primary bariatric surgery but long-term outcome remains unclear. The amount of food eaten is drastically reduced after LSG and may lead to nutritional deficiencies potentially. The aim of this study is to investigate long-term dietary intake and weight status after LSG. METHODS: Forty patients underwent LSG had more than 5-year follow-up with complete clinical data and food frequency questionnaires were analyzed. RESULTS: The mean age of subjects is 33.5 years old with mean body mass index (BMI) 37.9 kg/m2. Mean BMI loss at 5 years after LSG is 10.6 kg/m2. Weight regain appeared in 20% of patients. Dietary composition analysis at 5 years showed mean calorie intake of 1230 kcal/day, protein 70 g/day (22.5% of calorie), fat 50 g/day (36.1%), carbohydrate 126 g (41.4%), iron 7.5 mg/day, calcium 536.2 mg/day, and fiber 11.7 g/day. Calorie intake at 5 years after LSG is correlated with weight loss but weight regain is not related to a higher calorie intake. All comorbidities were significantly improved after LSG but hemoglobin and parathyroid hormone significantly changed. Incidence of iron deficiency anemia increased from 7.5% at pre-operation to 41.2% after LSG. Incidence of secondary hyperparathyroidism increased from 17.5 to 60.7%. CONCLUSION: LSG is an effective and durable bariatric procedure but with significant changes in nutritional status. Dietary instruction for LSG should include foods rich in protein, iron, calcium, and fiber.

Dissecting Torsin/cofactor function at the nuclear envelope: a genetic study.

The human genome encodes four Torsin ATPases, the functions of which are poorly understood. In this study, we use CRISPR/Cas9 engineering to delete all four Torsin ATPases individually and in combination. Using nuclear envelope (NE) blebbing as a phenotypic measure, we establish a direct correlation between the number of inactivated Torsin alleles and the occurrence of omega-shaped herniations within the lumen of the NE. A similar, although not identical, redundancy is observed for LAP1 and LULL1, which serve as regulatory cofactors for a subset of Torsin ATPases. Unexpectedly, deletion of Tor2A in a TorA/B/3A-deficient background results in a stark increase of bleb formation, even though Tor2A does not respond to LAP1/LULL1 stimulation. The robustness of the observed phenotype in Torsin-deficient cells enables a structural analysis via electron microscopy tomography and a compositional analysis via immunogold labeling. Ubiquitin and nucleoporins were identified as distinctively localizing components of the omega-shaped bleb structure. These findings suggest a functional link between the Torsin/cofactor system and NE/nuclear pore complex biogenesis or homeostasis and establish a Torsin-deficient cell line as a valuable experimental platform with which to decipher Torsin function.

The Lamin B receptor is essential for cholesterol synthesis and perturbed by disease-causing mutations.

Lamin B receptor (LBR) is a polytopic membrane protein residing in the inner nuclear membrane in association with the nuclear lamina. We demonstrate that human LBR is essential for cholesterol synthesis. LBR mutant derivatives implicated in Greenberg skeletal dysplasia or Pelger-Huët anomaly fail to rescue the cholesterol auxotrophy of a LBR-deficient human cell line, consistent with a loss-of-function mechanism for these congenital disorders. These disease-causing variants fall into two classes: point mutations in the sterol reductase domain perturb enzymatic activity by reducing the affinity for the essential cofactor NADPH, while LBR truncations render the mutant protein metabolically unstable, leading to its rapid degradation at the inner nuclear membrane. Thus, metabolically unstable LBR variants may serve as long-sought-after model substrates enabling previously impossible investigations of poorly understood protein turnover mechanisms at the inner nuclear membrane of higher eukaryotes.

Effect of probiotics on postoperative quality of gastric bypass surgeries: a prospective randomized trial.

BACKGROUND: Gastric bypass surgery is the recommended treatment for severely obese patients. However, postoperative symptomatic gastrointestinal (GI) episodes are common complaints. OBJECTIVES: To determine if administration of probiotics improves symptomatic GI episodes after gastric bypass surgery. SETTING: Hospital-based bariatric center. METHODS: This double-blind, randomized trial was conducted between March 2010 and September 2010 with 60 patients who underwent gastric bypass for severe obesity and experienced postoperative symptomatic GI episodes. Patients were randomly assigned to the probiotics group A (n = 20; 1 g Clostridium butyricum MIYAIRI twice daily); probiotics group B (n = 20; Bifidobacterium longum BB536 twice daily); or digestive enzymes group (n = 20; Aczym, containing 100 mg takadiastase N, 20 mg cellulase AP, 50 mg lipase MY, and 100 mg pancreatin, twice daily). Quality of life was measured using the modified Gastrointestinal Quality of Life Index (mGIQLI) before and after the 2-week intervention. RESULTS: Preintervention patient characteristics and mGIQLI scores were similar among the 3 groups. After the 2-week intervention, the mean mGIQLI score improved from 57.4 to 63.9 points in the entire sample and also within each group for 7 items specifically for 7: excessive passage of gas, foul smell of flatulence, belching, heartburn, abdominal noises, abdominal bloating, and abdominal pain. CONCLUSIONS: Administration of probiotics or digestive enzymes may improve symptomatic GI episodes after gastric bypass surgeries and improve quality of life, at least initially.

The Torsin Activator LULL1 Is Required for Efficient Growth of Herpes Simplex Virus 1.

UNLABELLED: TorsinA is a membrane-tethered AAA+ ATPase implicated in nuclear envelope dynamics as well as the nuclear egress of herpes simplex virus 1 (HSV-1). The activity of TorsinA and the related ATPase TorsinB strictly depends on LAP1 and LULL1, type II transmembrane proteins that are integral parts of the Torsin/cofactor AAA ring, forming a composite, membrane-spanning assembly. Here, we use CRISPR/Cas9-mediated genome engineering to create single- and double knockout (KO) cell lines of TorA and TorB as well as their activators, LAP1 and LULL1, to investigate the effect on HSV-1 production. Consistent with LULL1 being the more potent Torsin activator, a LULL1 KO reduces HSV-1 growth by one order of magnitude, while the deletion of other components of the Torsin system in combination causes subtle defects. Notably, LULL1 deficiency leads to a 10-fold decrease in the number of viral genomes per host cell without affecting viral protein production, allowing us to tentatively assign LULL1 to an unexpected role that precedes HSV-1 nuclear egress. IMPORTANCE: In this study, we conduct the first comprehensive genetic and phenotypic analysis of the Torsin/cofactor system in the context of HSV-1 infection, establishing LULL1 as the most important component of the Torsin system with respect to viral production.

Cellular RNA binding proteins NS1-BP and hnRNP K regulate influenza A virus RNA splicing.

Influenza A virus is a major human pathogen with a genome comprised of eight single-strand, negative-sense, RNA segments. Two viral RNA segments, NS1 and M, undergo alternative splicing and yield several proteins including NS1, NS2, M1 and M2 proteins. However, the mechanisms or players involved in splicing of these viral RNA segments have not been fully studied. Here, by investigating the interacting partners and function of the cellular protein NS1-binding protein (NS1-BP), we revealed novel players in the splicing of the M1 segment. Using a proteomics approach, we identified a complex of RNA binding proteins containing NS1-BP and heterogeneous nuclear ribonucleoproteins (hnRNPs), among which are hnRNPs involved in host pre-mRNA splicing. We found that low levels of NS1-BP specifically impaired proper alternative splicing of the viral M1 mRNA segment to yield the M2 mRNA without affecting splicing of mRNA3, M4, or the NS mRNA segments. Further biochemical analysis by formaldehyde and UV cross-linking demonstrated that NS1-BP did not interact directly with viral M1 mRNA but its interacting partners, hnRNPs A1, K, L, and M, directly bound M1 mRNA. Among these hnRNPs, we identified hnRNP K as a major mediator of M1 mRNA splicing. The M1 mRNA segment generates the matrix protein M1 and the M2 ion channel, which are essential proteins involved in viral trafficking, release into the cytoplasm, and budding. Thus, reduction of NS1-BP and/or hnRNP K levels altered M2/M1 mRNA and protein ratios, decreasing M2 levels and inhibiting virus replication. Thus, NS1-BP-hnRNPK complex is a key mediator of influenza A virus gene expression.

Host modulators of H1N1 cytopathogenicity.

Influenza A virus infects 5-20% of the population annually, resulting in ~35,000 deaths and significant morbidity. Current treatments include vaccines and drugs that target viral proteins. However, both of these approaches have limitations, as vaccines require yearly development and the rapid evolution of viral proteins gives rise to drug resistance. In consequence additional intervention strategies, that target host factors required for the viral life cycle, are under investigation. Here we employed arrayed whole-genome siRNA screening strategies to identify cell-autonomous molecular components that are subverted to support H1N1 influenza A virus infection of human bronchial epithelial cells. Integration across relevant public data sets exposed druggable gene products required for epithelial cell infection or required for viral proteins to deflect host cell suicide checkpoint activation. Pharmacological inhibition of representative targets, RGGT and CHEK1, resulted in significant protection against infection of human epithelial cells by the A/WS/33 virus. In addition, chemical inhibition of RGGT partially protected against H5N1 and the 2009 H1N1 pandemic strain. The observations reported here thus contribute to an expanding body of studies directed at decoding vulnerabilities in the command and control networks specified by influenza virulence factors.