GFI1 functions in transcriptional control and cell fate determination require SNAG domain methylation to recruit LSD1.

Proper hematopoietic cell fate decisions require co-ordinated functions of transcription factors, their associated co-regulators, and histone-modifying enzymes. Growth factor independence 1 (GFI1) is a zinc finger transcriptional repressor and master regulator of normal and malignant hematopoiesis. While several GFI1-interacting proteins have been described, how GFI1 leverages these relationships to carry out transcriptional repression remains unclear. Here, we describe a functional axis involving GFI1, SMYD2, and LSD1 that is a critical contributor to GFI1-mediated transcriptional repression. SMYD2 methylates lysine-8 (K8) within a -(8)KSKK(11)- motif embedded in the GFI1 SNAG domain. Methylation-defective GFI1 SNAG domain lacks repressor function due to failure of LSD1 recruitment and persistence of promoter H3K4 di-methyl marks. Methylation-defective GFI1 also fails to complement GFI1 depletion phenotypes in developing zebrafish and lacks pro-growth and survival functions in lymphoid leukemia cells. Our data show a discrete methylation event in the GFI1 SNAG domain that facilitates recruitment of LSD1 to enable transcriptional repression and co-ordinate control of hematopoietic cell fate in both normal and malignant settings.

Cervical Cancer Screening Barriers and Risk Factor Knowledge Among Uninsured Women.

A steady decline in cervical cancer incidence and mortality in the United States has been attributed to increased uptake of cervical cancer screening tests such as Papanicolau (Pap) tests. However, disparities in Pap test compliance exist, and may be due in part to perceived barriers or lack of knowledge about risk factors for cervical cancer. This study aimed to assess correlates of cervical cancer risk factor knowledge and examine socio-demographic predictors of self-reported barriers to screening among a group of low-income uninsured women. Survey and procedure data from 433 women, who received grant-funded cervical cancer screenings over a span of 33 months, were examined for this project. Data included demographics, knowledge of risk factors, and agreement on potential barriers to screening. Descriptive analysis showed significant correlation between educational attainment and knowledge of risk factors (r = 0.1381, P < 0.01). Multivariate analyses revealed that compared to Whites, Hispanics had increased odds of identifying fear of finding cancer (OR 1.56, 95% CI 1.00-2.43), language barriers (OR 4.72, 95% CI 2.62-8.50), and male physicians (OR 2.16, 95% CI 1.32-3.55) as barriers. Hispanics (OR 1.99, 95% CI 1.16-3.44) and Blacks (OR 2.06, 95% CI 1.15-3.68) had a two-fold increase in odds of agreeing that lack of knowledge was a barrier. Identified barriers varied with age, marital status and previous screening. Programs aimed at conducting free or subsidized screenings for medically underserved women should include culturally relevant education and patient care in order to reduce barriers and improve screening compliance for safety-net populations.

The cost-effectiveness of training US primary care physicians to conduct colorectal cancer screening in family medicine residency programs.

BACKGROUND: Demand for a wide array of colorectal cancer screening strategies continues to outpace supply. One strategy to reduce this deficit is to dramatically increase the number of primary care physicians who are trained and supportive of performing office-based colonoscopies or flexible sigmoidoscopies. This study evaluates the clinical and economic implications of training primary care physicians via family medicine residency programs to offer colorectal cancer screening services as an in-office procedure. METHODS: Using previously established clinical and economic assumptions from existing literature and budget data from a local grant (2013), incremental cost-effectiveness ratios are calculated that incorporate the costs of a proposed national training program and subsequent improvements in patient compliance. Sensitivity analyses are also conducted. RESULTS: Baseline assumptions suggest that the intervention would produce 2394 newly trained residents who could perform 71,820 additional colonoscopies or 119,700 additional flexible sigmoidoscopies after ten years. Despite high costs associated with the national training program, incremental cost-effectiveness ratios remain well below standard willingness-to-pay thresholds under base case assumptions. Interestingly, the status quo hierarchy of preferred screening strategies is disrupted by the proposed intervention. CONCLUSIONS: A national overhaul of family medicine residency programs offering training for colorectal cancer screening yields satisfactory incremental cost-effectiveness ratios. However, the model places high expectations on primary care physicians to improve current compliance levels in the US.

Determinants of variations in self-reported barriers to colonoscopy among uninsured patients in a primary care setting.

Colorectal cancer (CRC) is the third most common type of cancer among both males and females in the United States and the second leading cause of cancer-related deaths. Although largely preventable through screening, early detection and removal of polyps, screening rates are considered sub-optimal. Perceived barriers to screening have been reported to influence screening rates. This paper examines variations in the extent to which uninsured patients identified barriers to CRC screening using colonoscopy based on race/ethnicity, educational attainment, age, gender, marital status and prior colonoscopy. Multivariate analyses showed that compared to Caucasians, African Americans had an increased likelihood of identifying lack of transportation as a barrier [odds ratio (OR) 2.68; 95 % confidence interval (CI) 1.35-5.32] while Hispanics were more likely to identify fear of finding cancer as a barrier (OR 2.09; 95 % CI 1.19-3.66). Compared to those with more than a high school education, there was increased likelihood of identifying lack of knowledge as a barrier among individuals with high school education (OR 3.51; 95 % CI 1.94-6.36) or less than a high school education (OR 2.16; 95 % CI 1.04-4.50). Our findings suggest that strategies aimed at increasing colonoscopy screening rates among underserved populations should take into consideration race/ethnicity, educational attainment, age, and prior colonoscopy experience when developing education and outreach plans to reduce barriers to colonoscopy.

Directional Darwinian Selection in proteins.

BACKGROUND: Molecular evolution is a very active field of research, with several complementary approaches, including dN/dS, HON90, MM01, and others. Each has documented strengths and weaknesses, and no one approach provides a clear picture of how natural selection works at the molecular level. The purpose of this work is to present a simple new method that uses quantitative amino acid properties to identify and characterize directional selection in proteins. METHODS: Inferred amino acid replacements are viewed through the prism of a single physicochemical property to determine the amount and direction of change caused by each replacement. This allows the calculation of the probability that the mean change in the single property associated with the amino acid replacements is equal to zero (H0: µ = 0; i.e., no net change) using a simple two-tailed t-test. RESULTS: Example data from calanoid and cyclopoid copepod cytochrome oxidase subunit I sequence pairs are presented to demonstrate how directional selection may be linked to major shifts in adaptive zones, and that convergent evolution at the whole organism level may be the result of convergent protein adaptations. CONCLUSIONS: Rather than replace previous methods, this new method further complements existing methods to provide a holistic glimpse of how natural selection shapes protein structure and function over evolutionary time.

GFI1 Cooperates with IKZF1/IKAROS to Activate Gene Expression in T-cell Acute Lymphoblastic Leukemia.

Growth factor independence-1 (GFI1) is a transcriptional repressor and master regulator of normal and malignant hematopoiesis. Repression by GFI1 is attributable to recruitment of LSD1-containing protein complexes via its SNAG domain. However, the full complement of GFI1 partners in transcriptional control is not known. We show that in T-acute lymphoblastic leukemia (ALL) cells, GFI1 and IKAROS are transcriptional partners that co-occupy regulatory regions of hallmark T-cell development genes. Transcriptional profiling reveals a subset of genes directly transactivated through the GFI1-IKAROS partnership. Among these is NOTCH3, a key factor in T-ALL pathogenesis. Surprisingly, NOTCH3 expression by GFI1 and IKAROS requires the GFI1 SNAG domain but occurs independent of SNAG-LSD1 binding. GFI1 variants deficient in LSD1 binding fail to activate NOTCH3, but conversely, small molecules that disrupt the SNAG-LSD1 interaction while leaving the SNAG primary structure intact stimulate NOTCH3 expression. These results identify a noncanonical transcriptional control mechanism in T-ALL which supports GFI1-mediated transactivation in partnership with IKAROS and suggest competition between LSD1-containing repressive complexes and others favoring transactivation. IMPLICATIONS: Combinatorial diversity and cooperation between DNA binding proteins and complexes assembled by them can direct context-dependent transcriptional outputs to control cell fate and may offer new insights for therapeutic targeting in cancer.

Update on the evaluation and management of functional dyspepsia.

Dyspepsia affects up to 40 percent of adults each year and is often diagnosed as functional (nonulcer) dyspepsia. The defining symptoms are postprandial fullness, early satiation, or epigastric pain or burning in the absence of causative structural disease. These symptoms may coexist with symptoms of functional gastrointestinal disorders, such as gastroesophageal reflux and irritable bowel syndrome, as well as anxiety and depression. The history and physical examination can help identify other possible causes of the symptoms. Warning signs of serious disease, such as cancer, are unintended weight loss, progressive dysphagia, persistent vomiting, evidence of gastrointestinal bleeding, and a family history of cancer. In these cases, more extensive laboratory investigation, imaging, and endoscopy should be considered as clinically indicated. During the initial evaluation, a test-and-treat strategy to identify and eradicate Helicobacter pylori infection is more effective than empiric treatment and more cost-effective than initial endoscopy. Eradication of H. pylori helps one out of 15 patients with functional dyspepsia diagnosed by endoscopy, but may not be cost-effective. Treatment options that may be beneficial for functional dyspepsia include histamine H2 blockers, proton pump inhibitors, and prokinetic agents. Although psychotropic medications and psychological interventions have no proven benefit in patients with functional dyspepsia, they are appropriate for treating common psychiatric comorbidities.

Growth Factor Independence 1B-Mediated Transcriptional Repression and Lineage Allocation Require Lysine-Specific Demethylase 1-Dependent Recruitment of the BHC Complex.

Growth factor independence 1B (GFI1B) coordinates assembly of transcriptional repressor complexes comprised of corepressors and histone-modifying enzymes to control gene expression programs governing lineage allocation in hematopoiesis. Enforced expression of GFI1B in K562 erythroleukemia cells favors erythroid over megakaryocytic differentiation, providing a platform to define molecular determinants of binary fate decisions triggered by GFI1B. We deployed proteome-wide proximity labeling to identify factors whose inclusion in GFI1B complexes depends upon GFI1B's obligate effector, lysine-specific demethylase 1 (LSD1). We show that GFI1B preferentially recruits core and putative elements of the BRAF-histone deacetylase (HDAC) (BHC) chromatin-remodeling complex (LSD1, RCOR1, HMG20A, HMG20B, HDAC1, HDAC2, PHF21A, GSE1, ZMYM2, and ZNF217) in an LSD1-dependent manner to control acquisition of erythroid traits by K562 cells. Among these elements, depletion of both HMG20A and HMG20B or of GSE1 blocks GFI1B-mediated erythroid differentiation, phenocopying impaired differentiation brought on by LSD1 depletion or disruption of GFI1B-LSD1 binding. These findings demonstrate the central role of the GFI1B-LSD1 interaction as a determinant of BHC complex recruitment to enable cell fate decisions driven by GFI1B.

DNA reference alignment benchmarks based on tertiary structure of encoded proteins.

MOTIVATION: Multiple sequence alignments (MSAs) are at the heart of bioinformatics analysis. Recently, a number of multiple protein sequence alignment benchmarks (i.e. BAliBASE, OXBench, PREFAB and SMART) have been released to evaluate new and existing MSA applications. These databases have been well received by researchers and help to quantitatively evaluate MSA programs on protein sequences. Unfortunately, analogous DNA benchmarks are not available, making evaluation of MSA programs difficult for DNA sequences. RESULTS: This work presents the first known multiple DNA sequence alignment benchmarks that are (1) comprised of protein-coding portions of DNA (2) based on biological features such as the tertiary structure of encoded proteins. These reference DNA databases contain a total of 3545 alignments, comprising of 68 581 sequences. Two versions of the database are available: mdsa_100s and mdsa_all. The mdsa_100s version contains the alignments of the data sets that TBLASTN found 100% sequence identity for each sequence. The mdsa_all version includes all hits with an E-value score above the threshold of 0.001. A primary use of these databases is to benchmark the performance of MSA applications on DNA data sets. The first such case study is included in the Supplementary Material.

The evolution of foot-and-mouth disease virus: impacts of recombination and selection.

Foot-and-mouth disease virus is an economically important animal virus that exhibits extensive genetic and antigenic heterogeneity. To examine the evolutionary forces that have influenced the population dynamics of foot-and-mouth disease virus, individual genes and the coding genomes for the Eurasian (Asia1, A, C, and O) serotypes were examined for phylogenetic relationships, recombination, genetic diversity and selection. Our analyses demonstrate that paraphyletic relationships among serotypes are not as prevalent as previously proposed and suggest that convergent evolution might be obscuring phylogenetic relationships. We provide evidence that identification of recombinant sequences and recombination breakpoint patterns among and within serotypes are heavily dependent on the level of genetic diversity and convergent characters present in a particular data set as well as the methods used to detect recombination. Here, we also investigate the impact of adaptive positive selection on the capsid proteins and the non-structural genes 2B, 2C, 3A, and 3Cpro to identify genome regions involved in genetic diversity and antigenic variation. Two different categories of positive selection at the amino acid level were examined; conservative (stabilizing) selection that maintains particular phenotypic properties of an amino acid residue and radical (destabilizing), and selection that dramatically alters the phenotype and potentially the functional and/or structural features of the protein. Approximately, 29% of residues in the capsid proteins were under positive selection. Of those, 64% were under the influence of destabilizing selection, 80% were under the influence of stabilizing selection, and 44% had phenotypic properties influenced by both selection types. The majority of residues under selection (74%) were located outside of known antigenic sites; suggestive of additional uncharacterized epitopes and genomic regions involved in antigenic drift.

The impact of interprofessional education on family nurse practitioner students' and family medicine residents' knowledge and confidence in screening for breast and cervical cancer.

BACKGROUND AND PURPOSE: This study determined the impact of an interprofessional education (IPE) simulation on family nurse practitioner (FNP) students' and family medicine residents' (FMRs) self-reported confidence in counseling women reluctant to engage in cancer screening or evaluation and assessed knowledge of breast and cervical cancer risk factors. METHOD: A multi-item knowledge survey on breast and cervical cancer risk factors was administered to 76 FNP students and FMRs followed by an IPE simulation with a pre-/postsurvey of self-reported confidence in counseling a woman reluctant to have breast and cervical cancer screening and evaluation. DISCUSSION: Data demonstrated knowledge deficits in breast and cervical cancer risk factors in both disciplines with the average risk factor knowledge score of 8.5/12 for breast cancer and 7.8/12 for cervical cancer. Following IPE simulation, confidence in counseling women reluctant to have breast or cervical cancer screening improved across both disciplines (p < .05) and debrief feedback findings suggest improved attitudes toward collegiality, communication, and understanding of other interprofessional roles among both disciplines. CONCLUSION: Knowledge gaps exist among both FNP students and FMRs in breast and cervical cancer risk factors. This study suggests IPE simulation is effective in building individual provider confidence and team collegiality.

Hybrid insulin cocrystals for controlled release delivery.

The ability to tailor the release profile of a drug by manipulating its formulation matrix offers important therapeutic advantages. We show here that human insulin can be cocrystallized at preselected ratios with the fully active lipophilically modified insulin derivative octanoyl-N(epsilon)-LysB29-human insulin (C8-HI). The cocrystal is analogous to the NPH (neutral protamine Hagedorn) crystalline complex formed with human insulin, which is commonly used as the long-acting insulin component of diabetes therapy. The in vitro and in vivo release rates of the cocrystal can be controlled by adjusting the relative proportions of the two insulin components. We identified a cocrystal composition comprising 75% C8-HI and 25% human insulin that exhibits near-ideal basal pharmacodynamics in somatostatin-treated beagle dogs. The dependence of release rate on cocrystal ratio provides a robust mechanism for modulating insulin pharmacodynamics. These findings show that a crystalline protein matrix may accommodate a chemical modification that alters the dissolution rate of the crystal in a therapeutically useful way, yet that is structurally innocuous enough to preserve the pharmaceutical integrity of the original microcrystalline entity and the pharmacological activity of the parent molecule.

Determinants of HPV vaccine awareness and healthcare providers' discussion of HPV vaccine among females.

Two human papillomavirus (HPV) vaccines are available and can prevent 98% of HPV 16 and 18 infections. This study aimed to explore determinants of 1) HPV vaccine awareness among a cohort of low-income women participating in a cancer prevention program in Central Texas and compare them to United States residents; 2) determinants of healthcare providers' discussion of HPV vaccine among female residents of the United States. Bivariate and multivariable analysis of HPV vaccine awareness using survey data (n = 359) collected between 2014 and 2016 in Central Texas, and the Health Information and Nutrition Survey (HINTS) data which is a nationally representative dataset (unweighted n = 1214) collected in 2013 were conducted. Bivariate and multivariable regression analyses of healthcare providers' discussion of the HPV vaccine using the HINTS survey data were also conducted. Compared to non-Hispanic Whites, there was a decreased likelihood of HPV vaccine awareness among non-Hispanic Blacks (OR = 0.50; 95% CI = 0.28-0.90) and Hispanics (OR = 0.55; 95% CI = 0.30-0.99) in the grant funded program, as well as non-Hispanic Blacks (OR = 0.28; 95% CI = 0.14-0.58) and Hispanics (OR = 0.22; 95% CI = 0.12-0.41) in the HINTS data. There was also a decreased likelihood of healthcare providers discussing the HPV vaccine with respondents who were 35-49 years (OR = 0.50; 95% CI = 0.30-0.84), 50-64 years (OR = 0.26; 95% CI = 0.14-0.49) or ≥ 65 years compared to those who were 18-34 years among the HINTS data respondents. Interventions to increase HPV awareness among non-Hispanic Blacks and Hispanics, as well as encourage healthcare providers' discussion of the HPV vaccination during patient encounters regardless of the patient's age are needed.

Phylogenomics and molecular evolution of polyomaviruses.

We provide in this chapter an overview of the basic steps to reconstruct evolutionary relationships through standard phylogeny estimation approaches as well as network approaches for sequences more closely related. We discuss the importance of sequence alignment, selecting models of evolution, and confidence assessment in phylogenetic inference. We also introduce the reader to a variety of software packages used for such studies. Finally, we demonstrate these approaches throughout using a data set of 33 whole genomes of polyomaviruses. A robust phylogeny of these genomes is estimated and phylogenetic relationships among the polyomaviruses determined using Bayesian and maximum likelihood approaches. Furthermore, population samples of SV40 are used to demonstrate the utility of network approaches for closely related sequences. The phylogenetic analysis suggested a close relationship among the BK viruses, JC viruses, and SV40 with a more distant association with mouse polyomavirus, monkey polymavirus (LPV) and then avian polyomavirus (BFDV).

SUMOylation Regulates Growth Factor Independence 1 in Transcriptional Control and Hematopoiesis.

Cell fate specification requires precise coordination of transcription factors and their regulators to achieve fidelity and flexibility in lineage allocation. The transcriptional repressor growth factor independence 1 (GFI1) is comprised of conserved Snail/Slug/Gfi1 (SNAG) and zinc finger motifs separated by a linker region poorly conserved with GFI1B, its closest homolog. Moreover, GFI1 and GFI1B coordinate distinct developmental fates in hematopoiesis, suggesting that their functional differences may derive from structures within their linkers. We show a binding interface between the GFI1 linker and the SP-RING domain of PIAS3, an E3-SUMO (small ubiquitin-related modifier) ligase. The PIAS3 binding region in GFI1 contains a conserved type I SUMOylation consensus element, centered on lysine-239 (K239). In silico prediction algorithms identify K239 as the only high-probability site for SUMO modification. We show that GFI1 is modified by SUMO at K239. SUMOylation-resistant derivatives of GFI1 fail to complement Gfi1 depletion phenotypes in zebrafish primitive erythropoiesis and granulocytic differentiation in cultured human cells. LSD1/CoREST recruitment and MYC repression by GFI1 are profoundly impaired for SUMOylation-resistant GFI1 derivatives, while enforced expression of MYC blocks granulocytic differentiation. These findings suggest that SUMOylation within the GFI1 linker favors LSD1/CoREST recruitment and MYC repression to govern hematopoietic differentiation.

Expanding Access to Colorectal Cancer Screening: Benchmarking Quality Indicators in a Primary Care Colonoscopy Program.

BACKGROUND: An inadequate supply of physicians who perform colonoscopies contributes to suboptimal screening rates, especially among the underserved. This shortage could be reduced if primary care physicians perform colonoscopies. This purpose of this article is to report quality indicators from colonoscopy procedures performed by family medicine physicians as part of a colorectal cancer prevention program targeting uninsured, low-income individuals. METHODS: A grant-funded colorectal cancer screening program was implemented to increase access to affordable colonoscopies for underinsured or uninsured residents of target counties while providing colonoscopy training to family medicine resident physicians. Colonoscopies were performed or supervised by 4 board-certified family physicians. Data were collected between 2011 and 2014. RESULTS: A total of 1155 colonoscopies were performed on 1101 individuals over a 3-year period. Cecal intubation rate was 96.25%. Adenoma detection rates among men and women >50 years old were 38.15% and 25.96%, respectively. There was 1 perforation, which was referred to a hospital, and 1 instance of postprocedural bleeding, which spontaneously resolved. CONCLUSIONS: Primary care physicians performing colonoscopies met the recommended quality indicators set forth by the American Society for Gastrointestinal Endoscopy.

The role of health literacy and communication habits on previous colorectal cancer screening among low-income and uninsured patients.

OBJECTIVE: To determine the association between health literacy, communication habits and colorectal cancer (CRC) screening among low-income patients. METHODS: Survey responses of patients who received financial assistance for colonoscopy between 2011 and 2014 at a family medicine residency clinic were analyzed using multivariate logistic regression (n = 456). There were two dependent variables: (1) previous CRC screening and (2) CRC screening adherence. Our independent variables of interest were health literacy and communication habits. RESULTS: Over two-thirds (67.13%) of respondents had not been previously screened for CRC. Multivariate analysis showed a decreased likelihood of previous CRC screening among those who had marginal (OR = 0.52; 95% CI = 0.29-0.92) or inadequate health literacy (OR = 0.49; 95% CI = 0.27-0.87) compared to those with adequate health literacy. Controlling for health literacy, the significant association between educational attainment and previous CRC screening was eliminated. Thus, health literacy mediated the relationship between educational attainment and previous CRC screening. There was no significant association between communication habits and previous CRC screening. There was no significant association between screening guideline adherence, and health literacy or communication. CONCLUSION: Limited health literacy is a potential barrier to CRC screening. Suboptimal CRC screening rates reported among those with lower educational attainment may be mediated by limited health literacy.

Detecting molecular selection on single amino acid replacements.

Amino acid property methods have repeatedly proven more sensitive than nucleotide-based methods, especially to the subtle influences of selection on Single Nucleotide Polymorphisms (SNPs). The purpose of this study is to evaluate the effects of population sampling and sliding window size on the sensitivity of one amino acid based method by assessing drug resistant SNPs from the HIV-1 gag-pol gene. The analysis of most of the SNPs produced positive results. There was not a trend in terms of properties affected most often, but sliding window size affected results more than population sampling.

Tired of seeing your patients in the ED? Let's give parent education another look!

This study was conducted to determine if a parent-oriented educational intervention reduces the use of emergency department (ED) services for the care of infants. Infants aged 7 days to 1 year and older children aged 2 to 10 years were tracked for 3 years in two separate primary care (PC) practices in Washington County, Texas, with the last year being the interventional study period. Also, infants aged 7 days to 1 year and older children aged 7 days to 5 years were tracked in a third PC practice in Burleson County, Texas. The study group consisted of all parents of patients aged 7 days to 1 year seen by the pediatric group in Washington County during the 1-year interventional period. Only parents of infants in the study group received a specific educational intervention booklet. Five separate control groups were followed in this study. The control groups received usual care with standardized patient information, but they did not receive the educational intervention booklet. Each group was evaluated by calculating its monthly ED utilization rate, which is the quotient derived from dividing the number of children from that particular group seen in the ED per month by the number of children from the same group seen in the PC clinic per month. A difference of proportions test was applied to test for statistical significance regarding ED utilization. Compared with parents in the control group, parents receiving the intervention booklet (the study group) showed significantly (P < .05) lower use of ED services for care of their infants. We found no change in ED utilization for children of parents receiving other standard educational information.