Glyceollin Transcription Factor GmMYB29A2 Regulates Soybean Resistance to Phytophthora sojae.

Glyceollin isomers I, II, and III are the major pathogen-elicited secondary metabolites (i.e. phytoalexins) of soybean (Glycine max) that, collectively with other 5-deoxyisoflavonoids, provide race-specific resistance to Phytophthora sojae. The NAC-family transcription factor (TF) GmNAC42-1 is an essential regulator of some but not all glyceollin biosynthesis genes, indicating other essential TF(s) of the glyceollin gene regulatory network remain to be identified. Here, we conducted comparative transcriptomics on soybean hairy roots of the variety Williams 82 and imbibing seeds of Harosoy 63 upon treatment with wall glucan elicitor from P. sojae and identified two homologous R2R3-type MYB TF genes, GmMYB29A1 and GmMYB29A2, up-regulated during the times of peak glyceollin biosynthesis. Overexpression and RNA interference silencing of GmMYB29A2 increased and decreased expression of GmNAC42-1, GmMYB29A1, and glyceollin biosynthesis genes and metabolites, respectively, in response to wall glucan elicitor. By contrast, overexpressing or silencing GmMYB29A1 decreased glyceollin I accumulation with marginal or no effects on the expressions of glyceollin synthesis genes, suggesting a preferential role in promoting glyceollin turnover and/or competing biosynthetic pathways. GmMYB29A2 interacted with the promoters of two glyceollin I biosynthesis genes in vitro and in vivo. Silencing GmMYB29A2 in Williams 82, a soybean variety that encodes the resistance gene Rps1k, rendered it compatible with race 1 P. sojae, whereas overexpressing GmMYB29A2 rendered the susceptible Williams variety incompatible. Compatibility and incompatibility coincided with reduced and enhanced accumulations of glyceollin I but not other 5-deoxyisoflavonoids. Thus, GmMYB29A2 is essential for accumulation of glyceollin I and expression of Phytophthora resistance.

The NAC family transcription factor GmNAC42-1 regulates biosynthesis of the anticancer and neuroprotective glyceollins in soybean.

BACKGROUND: Glyceollins are isoflavonoid-derived pathogen-inducible defense metabolites (phytoalexins) from soybean (Glycine max L. Merr) that have important roles in providing defense against pathogens. They also have impressive anticancer and neuroprotective activities in mammals. Despite their potential usefulness as therapeutics, glyceollins are not economical to synthesize and are biosynthesized only transiently and in low amounts in response to specific stresses. Engineering the regulation of glyceollin biosynthesis may be a promising approach to enhance their bioproduction, yet the transcription factors (TFs) that regulate their biosynthesis have remained elusive. To address this, we first aimed to identify novel abiotic stresses that enhance or suppress the elicitation of glyceollins and then used a comparative transcriptomics approach to search for TF gene candidates that may positively regulate glyceollin biosynthesis. RESULTS: Acidity stress (pH 3.0 medium) and dehydration exerted prolonged (week-long) inductive or suppressive effects on glyceollin biosynthesis, respectively. RNA-seq found that all known biosynthetic genes were oppositely regulated by acidity stress and dehydration, but known isoflavonoid TFs were not. Systemic acquired resistance (SAR) genes were highly enriched in the geneset. We chose to functionally characterize the NAC (NAM/ATAF1/2/CUC2)-family TF GmNAC42-1 that was annotated as an SAR gene and a homolog of the Arabidopsis thaliana (Arabidopsis) indole alkaloid phytoalexin regulator ANAC042. Overexpressing and silencing GmNAC42-1 in elicited soybean hairy roots dramatically enhanced and suppressed the amounts of glyceollin metabolites and biosynthesis gene mRNAs, respectively. Yet, overexpressing GmNAC42-1 in non-elicited hairy roots failed to stimulate the expressions of all biosynthesis genes. Thus, GmNAC42-1 was necessary but not sufficient to activate all biosynthesis genes on its own, suggesting an important role in the glyceollin gene regulatory network (GRN). The GmNAC42-1 protein directly bound the promoters of biosynthesis genes IFS2 and G4DT in the yeast one-hybrid (Y1H) system. CONCLUSIONS: Acidity stress is a novel elicitor and dehydration is a suppressor of glyceollin biosynthesis. The TF gene GmNAC42-1 is an essential positive regulator of glyceollin biosynthesis. Overexpressing GmNAC42-1 in hairy roots can be used to increase glyceollin yields > 10-fold upon elicitation. Thus, manipulating the expressions of glyceollin TFs is an effective strategy for enhancing the bioproduction of glyceollins in soybean.

The relationship between ventilatory threshold and repeated-sprint ability in competitive male ice hockey players.

BACKGROUND/OBJECTIVE: The relationship between ventilatory threshold (VT1, VT2) and repeated-sprint ability (RSA) in competitive male ice hockey players was investigated. METHODS: Forty-three male ice hockey players aged 18-23 years competing in NCAA Division I, NCAA Division III, and Junior A level participated. Participants performed an incremental graded exercise test on a skate treadmill to determine V ˙ O2peak, VT1, and VT2 using MedGraphics Breezesuit™ software (v-slope). Participants performed an on-ice repeated shift (RSA) test consisting of 8-maximal skating bouts, lasting approximately 25 s and interspersed with 90 s of passive recovery, to determine first gate, second gate, and total sprint decrement (%dec). Pearson product-moment correlations and multiple regressions were used to assess relationships between ventilatory threshold variables (VT1, VT2, Stage at VT1, and Stage at VT2) and RSA (first gate, second gate, and total course decrement). RESULTS: Stage at VT2 was the only variable substantially correlated with first gate (r = -0.35; P < 0.05), second gate (r = -0.58; P < 0.001) and total course decrement (r = -0.42; P < 0.05). CONCLUSION: The results of this study demonstrated that VT is substantially associated with RSA, and VT2 is more strongly correlated with RSA than V ˙ O2peak. This study suggests that longer duration high-intensity interval training at intensities that increase workrate at VT2 may lead to possible improvements in RSA.

National utilization patterns of warfarin use in older patients with atrial fibrillation: a population-based study of Medicare Part D beneficiaries.

BACKGROUND: Although warfarin therapy reduces stroke incidence in patients with atrial fibrillation (AF), the rate of warfarin use in this population remains low. In 2008, the Medicare Part D program was expanded to pay for medications for Medicare enrollees. OBJECTIVE: To examine rates and predictors of warfarin use in Medicare Part D beneficiaries with AF. METHODS: This population-based retrospective cohort study used claims data from 41,447 Medicare beneficiaries aged 66 and older with at least 2 AF diagnoses in 2007 and at least 1 diagnosis in 2008. All subjects had continuous Medicare Part D prescription coverage in 2008. Statistical analysis using χ(2) was used to examine differences in warfarin use by patient characteristics (age, ethnicity, sex, Medicaid eligibility, comorbidities, contraindications to warfarin, and whether they visited a cardiologist or a primary care physician [PCP]), CHADS(2) score (congestive heart failure, hypertension, age, diabetes, and stroke or transient ischemic attack; higher scores indicate higher risks of stroke), and geographic regions. Using hierarchical generalized linear models restricted to subjects without warfarin contraindications (n = 34,947), we examined the effect of patient characteristics and geographic regions on warfarin use. RESULTS: The overall warfarin use rate was 66.8%. The warfarin use rates varied between hospital referral regions, with highest rates in the Midwestern states and lowest rates in the South. The regional variation persisted even after adjustment for patient characteristics. Multivariable analysis showed that the odds of being on warfarin decreased significantly with age and increasing comorbidity, in blacks, and among those with low income. Seeing a cardiologist (OR 1.10; 95% CI 1.05-1.16), having a PCP (OR 1.23; 95% CI 1.17-1.29), and CHADS(2) score of 2 or greater (OR 1.09; 95% CI 1.01-1.17) were associated with increased odds of warfarin use. CONCLUSIONS: Warfarin use rates vary by patient characteristics and region, with higher rates among residents of the Midwest and among patients seen by cardiologists and PCPs. Preventing stroke-related disability in AF requires implementation of evidence-based initiatives to increase warfarin use.

Improving testing for hepatitis B before treatment with rituximab.

AIMS/OBJECTIVES/BACKGROUND: Individuals with current or previous infection with the hepatitis B virus (HBV) can experience viral reactivation when treated with immunosuppression. Rituximab, an anti-CD20 antibody used to treat many diseases, has potent immunosuppressant effects with a high risk of causing HBV reactivation. Reactivation can range from elevated liver enzymes to acute severe hepatitis with liver failure and a significant mortality risk. HBV screening and appropriate use of prophylactic antiviral therapy can prevent reactivation. This work describes the introduction of a local policy for HBV testing in patients before rituximab treatment and assesses its impact. METHODS AND RESULTS: A baseline review (before policy introduction) of 90 patients showed that only 21 (23%) had hepatitis B surface antigen (HBsAg) and 17 (19%) had hepatitis B core antibody (anti-HBcAb) tested before receiving rituximab. Following introduction of the policy (on the basis of international guidelines), improved laboratory reporting protocols and targeted education sessions, two further reviews of HBV testing rates among patients being initiated onto rituximab were performed. There was a marked increase in pre-rituximab testing for HBsAg from 23 to 79% and for anti-HBcAb from 19 to 78%. Throughout the study period, a total of one (0.8%) HBsAg-positive and six (4.7%) anti-HBcAb-positive patients were identified. CONCLUSIONS: This work clearly indicates that simple strategies can markedly improve appropriate HBV screening. In our cohort, 6% (of whom only 43% had recognized HBV risk factors) required antiviral prophylaxis, which emphasizes the importance of universal screening before rituximab. Reinforcement of the guidelines and ongoing education is needed to further increase testing rates.