Discussion of Annotated Research Articles Results in Increases in Scientific Literacy within a Cell Biology Course.

As the amount and complexity of scientific knowledge continues to grow, it is essential to educate scientifically literate citizens who can comprehend the process of science and the implications of technological advances. This is especially important when educating science, technology, engineering, and mathematics (STEM) college students, since they may play a central role in the future of scientific research and its communication. A central part of decoding and interpreting scientific information is the ability to analyze scientific research articles. For this reason, many different approaches for reading scientific research articles have been developed and published. Despite the availability of numerous ways of analyzing scientific research articles, biology students can face challenges that may prevent them from fully comprehending the text. We sought to address student challenges with science vocabulary and content knowledge by adding structural supports to in-classroom article discussions through the use of annotated articles from the Science in the Classroom initiative. We describe the pedagogical approach used for discussing scientific research articles within a required biology course. In this context, we found that students' scientific literacy skills increased at the end of the semester. We also found that, for each article discussed, the majority of students could interpret graphical representations of article results and that they could identify and comprehend components of the experimental design of the study.

From Novice To Expert: An Assessment To Measure Strategies Students Implement While Learning To Read Primary Scientific Literature.

Primary Scientific Literature (PSL) has been used in undergraduate classrooms as a way to engage students with the research process and to increase science literacy. Most curricula lack any formal training for undergraduates to critically read PSL even though most undergraduate science courses require students to engage with PSL at some level. In addition, there are limited studies exploring the process by which expertise in reading PSL develops in undergraduates. In this study, we adapted behaviors that expert and novice PSL readers exhibit into a quantitative assessment tool, the PSL Reading Strategies Assessment, to evaluate undergraduates' development of reading strategies when learning to read PSL. Factor analysis and reliability measures were implemented to determine the structure of our assessment tool. Our results show the PSL Reading Strategies Assessment is sensitive enough to measure differences among student populations, suggesting that it can be used as a diagnostic tool to guide instructors and researchers as they change curricula, implement new teaching strategies, and strive to develop students' science literacy. Moreover, our data show that developing expert-like reading strategies in students learning to read PSL is not easy. Simply reading a PDF does little to promote the development of reading strategies in students learning to read PSL.

Insights on biology student motivations and challenges when reading and analyzing primary literature.

Reading primary literature is a popular classroom practice that exposes students to the process of science. However, the analysis of primary literature can be taxing and time-consuming for students. For this reason, it is important to determine the source of student challenges and what motivates them to read primary literature. To better understand students' challenges, preferences, and motivations towards analyzing primary literature, we held focus groups with biology undergraduates where we asked them about their thoughts and perceptions on this practice. Students felt they struggle with understanding the big picture of an article, certain aspects of scientific literacy like data interpretation and experimental setup, and lack of knowledge of terms and techniques. Further analysis of the data using the achievement goal and expectancy-value theories of motivation revealed that students: 1) demonstrate mastery and performance approach goal orientations, which are typically associated with positive learning outcomes, 2) value the usefulness of reading primary literature, and 3) feel most engaged in the process of reading an article when the topic interests them. We provide pedagogical recommendations based on our findings.

Characterization and Genome Sequence of the Mycobacteriophage Donny.

We report the discovery of the novel bacteriophage Donny, a Siphoviridae virus that infects Mycobacterium smegmatis mc2155. Donny has a genome length of 69,691 bp and a G+C content of 68.5%. Donny shares 99% and 93% nucleotide identity with bacteriophages Acadian and Baee, respectively.

Development of a structure-analysis pipeline using multiple-solvent crystal structures of barrier-to-autointegration factor.

The multiple-solvent crystal structure (MSCS) approach uses high concentrations of organic solvents to characterize the interactions and effects of solvents on proteins. Here, the method has been further developed and an MSCS data-handling pipeline is presented that uses the Detection of Related Solvent Positions (DRoP) program to improve data quality. DRoP is used to selectively model conserved water molecules, so that an advanced stage of structural refinement is reached quickly. This allows the placement of organic molecules more accurately and convergence on high-quality maps and structures. This pipeline was applied to the chromatin-associated protein barrier-to-autointegration factor (BAF), resulting in structural models with better than average statistics. DRoP and Phenix Structure Comparison were used to characterize the data sets and to identify a binding site that overlaps with the interaction site of BAF with emerin. The conserved water-mediated networks identified by DRoP suggested a mechanism by which water molecules are used to drive the binding of DNA. Normalized and differential B-factor analysis is shown to be a valuable tool to characterize the effects of specific solvents on defined regions of BAF. Specific solvents are identified that cause stabilization of functionally important regions of the protein. This work presents tools and a standardized approach for the analysis and comprehension of MSCS data sets.

Characterization and Genome Sequence of Mycobacterium Phage XianYue.

Novel mycobacteriophage XianYue was isolated in Northeast Georgia and infects Mycobacteria smegmatis mc2155. Actinobacteriophages which share at least 50% nucleotide identity are grouped into clusters, with XianYue in cluster A2. Its genome is 52,907 bp with 91 open reading frames (ORFs) and 62.9% GC content, and it shares 86.51% nucleotide identity with mycobacteriophage Trixie.

BAF: roles in chromatin, nuclear structure and retrovirus integration.

Barrier-to-autointegration factor (BAF) is an essential protein that is highly conserved in metazoan evolution. BAF binds directly to double-stranded DNA, nuclear LEM-domain proteins, lamin A and transcription activators. BAF is also a host cell component of retroviral pre-integration complexes. BAF binds matrix, a retroviral protein, and facilitates efficient retroviral DNA integration in vitro through unknown mechanisms. New findings suggest that BAF has structural roles in nuclear assembly and chromatin organization, represses gene expression and might interlink chromatin structure, nuclear architecture and gene regulation in metazoans.

Barrier-to-autointegration factor: major roles in chromatin decondensation and nuclear assembly.

Barrier-to-autointegration factor (BAF) is a DNA-bridging protein, highly conserved in metazoans. BAF binds directly to LEM (LAP2, emerin, MAN1) domain nuclear membrane proteins, including LAP2 and emerin. We used site-directed mutagenesis and biochemical analysis to map functionally important residues in human BAF, including those required for direct binding to DNA or emerin. We also tested wild-type BAF and 25 point mutants for their effects on nuclear assembly in Xenopus egg extracts, which contain approximately 12 microM endogenous BAF dimers. Exogenous BAF caused two distinct effects: at low added concentrations, wild-type BAF enhanced chromatin decondensation and nuclear growth; at higher added concentrations, wild-type BAF completely blocked chromatin decondensation and nuclear growth. Mutants fell into four classes, including one that defines a novel functional surface on the BAF dimer. Our results suggest that BAF, unregulated, potently compresses chromatin structure, and that BAF interactions with both DNA and LEM proteins are critical for membrane recruitment and chromatin decondensation during nuclear assembly.

Expert-Novice Comparison Reveals Pedagogical Implications for Students' Analysis of Primary Literature.

Student engagement in the analysis of primary scientific literature increases critical thinking, scientific literacy, data evaluation, and science process skills. However, little is known about the process by which expertise in reading scientific articles develops. For this reason, we decided to compare how faculty experts and student novices engage with a research article. We performed think-aloud interviews of biology faculty and undergraduates as they read through a scientific article. We analyzed these interviews using qualitative methods. We grounded data interpretation in cognitive load theory and the ICAP (interactive, constructive, active, and passive) framework. Our results revealed that faculty have more complex schemas than students and that they reduce cognitive load through two main mechanisms: summarizing and note-taking. Faculty also engage with articles at a higher cognitive level, described as constructive by the ICAP framework, when compared with students. More complex schemas, effectively lowering cognitive load, and deeper engagement with the text may help explain why faculty encounter fewer comprehension difficulties than students in our study. Finally, faculty analyze and evaluate data more often than students when reading the text. Findings include a discussion of successful pedagogical approaches for instructors wishing to enhance undergraduates' comprehension and analysis of research articles.

Genome Sequence of Mycobacterium Phage LilHazelnut.

Here, we describe LilHazelnut, a novel mycobacteriophage that infects Mycobacterium smegmatis mc2155. LilHazelnut is a cluster Q phage that shares 99% nucleotide identity with phage Giles, is 53,746 bp in length, and has a G+C content of 67.5%. LilHazelnut is a temperate Siphoviridae virus, as is typical of cluster Q family members.

QR Code Lecture Activity as a Tool for Increasing Nonmajors Biology Students' Enjoyment of Interaction with Their Local Environment.

The impact of the Internet on education has been recognized for decades, and as technology advances, the ways in which students can access Internet content is ever increasing. Most students have some kind of portable smart device with which they access Internet content without the locational constraints of a desktop computer. This mobility has prompted abundant literature suggesting ways that Quick Response Codes (QR codes), a kind of two dimensional barcode, could be used to advance student learning. However, very few studies have tested the usefulness of QR codes in undergraduate science classes. We report on our development of a campus "scavenger hunt" activity using QR codes. We found that this activity develops application skills of the concepts of native and invasive species and enjoyment of coverage of content relative to traditional lecture in a nonmajors Environmental Science class at a four-year teaching institution.

Genome Sequence of Mycobacterium Phage CrystalP.

Mycobacteriophage CrystalP is a newly isolated phage infecting Mycobacterium smegmatis strain mc2155. CrystalP has a 76,483-bp genome and is predicted to contain 143 protein-coding and 2 tRNA genes, including repressor and integrase genes consistent with a temperate lifestyle. CrystalP is related to the mycobacteriophages Toto and Kostya and to other Cluster E phages.

Nuclear membrane protein emerin: roles in gene regulation, actin dynamics and human disease.

Loss of emerin, a nuclear membrane protein, causes Emery-Dreifuss muscular dystrophy (EDMD), characterized by muscle weakening, contractures of major tendons and potentially lethal cardiac conduction system defects. Emerin has a LEM-domain and therefore binds barrier-to-autointegration factor (BAF), a conserved chromatin protein essential for cell division. BAF recruits emerin to chromatin and regulates higher-order chromatin structure during nuclear assembly. Emerin also binds filaments formed by A-type lamins, mutations in which also cause EDMD. Other partners for emerin include nesprin-1alpha and transcriptional regulators such as germ cell-less (GCL). The binding affinities of these partners range from 4nM (nesprin-1alpha) to 200 nM (BAF), and are physiologically significant. Biochemical studies therefore provide a valid means to predict the properties of emerin-lamin complexes in vivo. Emerin and lamin A together form stable complexes with either BAF or GCL in vitro. BAF, however, competes with GCL for binding to emerin in vitro. These and additional partners, notably actin and nuclear myosin II, suggest disease-relevant roles for emerin in gene regulation and the mechanical interity of the nucleus.

Nesprin-1alpha self-associates and binds directly to emerin and lamin A in vitro.

Nesprin-1alpha is a spectrin repeat (SR)-containing, transmembrane protein of the inner nuclear membrane, and is highly expressed in muscle cells. A yeast two-hybrid screen for nesprin-1alpha-interacting proteins showed that nesprin-1alpha interacted with itself. Blot overlay experiments revealed that nesprin-1alpha's third SR binds the fifth SR. The carboxy-terminal half of nesprin-1alpha directly bound lamin A, a nuclear intermediate filament protein. Biochemical analysis demonstrated that nesprin-1alpha dimers bind directly to the nucleoplasmic domain of emerin, an inner nuclear membrane protein, with an affinity of 4 nM. Binding was optimal for full nucleoplasmic dimers of nesprin-1alpha, since nesprin fragments SR1-5 and SR5-7 bound emerin as monomers with affinities of 53 nM and 250 mM, respectively. We propose that membrane-anchored nesprin-1alpha antiparallel dimers interact with both emerin and lamin A to provide scaffolding at the inner nuclear membrane.

The CREATE Method Does Not Result in Greater Gains in Critical Thinking than a More Traditional Method of Analyzing the Primary Literature.

Analysis of the primary literature in the undergraduate curriculum is associated with gains in student learning. In particular, the CREATE (Consider, Read, Elucidate hypotheses, Analyze and interpret the data, and Think of the next Experiment) method is associated with an increase in student critical thinking skills. We adapted the CREATE method within a required cell biology class and compared the learning gains of students using CREATE to those of students involved in less structured literature discussions. We found that while both sets of students had gains in critical thinking, students who used the CREATE method did not show significant improvement over students engaged in a more traditional method for dissecting the literature. Students also reported similar learning gains for both literature discussion methods. Our study suggests that, at least in our educational context, the CREATE method does not lead to higher learning gains than a less structured way of reading primary literature.

Barrier-to-autointegration factor-like (BAF-L): a proposed regulator of BAF.

Barrier-to-autointegration factor (BAF) is an essential chromatin protein conserved in metazoans. BAF has roles in nuclear assembly, chromatin organization, gene expression, and gonad development and is exploited by retroviruses. BAF forms stable dimers that bind nonspecifically to dsDNA and specifically to LEM-domain proteins (e.g., LAP2beta, emerin, MAN1), homeodomain transcription factors, histones, and lamin A. We characterized a protein named BAF-Like (BAF-L) that in humans is 40% identical to BAF. Overexpression studies in HeLa cells show that BAF-L, like BAF, is a predominantly nuclear protein. Recombinant BAF-L forms stable homodimers and heterodimerizes with BAF in vitro and also interacts with BAF in vivo. BAF-L does not bind significantly to DNA, LAP2beta, or emerin but can form ternary complexes in vitro with BAF plus DNA, or BAF plus LAP2beta. Levels of BAF-L mRNA were high in pancreas and testis, suggesting functions in the germline. BAF-L mRNA was detectable at low levels in eleven other tissues and undetectable in heart and skeletal muscle which are specifically affected by Emery-Dreifuss muscular dystrophy, a disease caused by mutations in either emerin or lamin A. We propose that BAF-L regulates BAF function via heterodimerization and might thereby influence tissue-specific roles of BAF.

Barrier-to-autointegration factor is required to segregate and enclose chromosomes within the nuclear envelope and assemble the nuclear lamina.

Barrier-to-autointegration factor (BAF) binds dsDNA, LEM-domain proteins, and lamins. Caenorhabditis elegans BAF requires Ce-lamin and two LEM-domain proteins (Ce-emerin and Ce-MAN1) to localize during nuclear assembly. It was unknown whether Ce-lamin and LEM proteins, in turn, depend on Ce-BAF (mutually dependent structural roles). RNA interference-mediated down-regulation of Ce-BAF caused gross defects in chromosome segregation, chromatin decondensation, and mitotic progression as early as the two-cell stage, and embryos died at the approximately 100-cell stage. Nuclear pores reassembled, whereas Ce-lamin, Ce-emerin, and Ce-MAN1 bound chromatin but remained patchy and disorganized. The nuclear membranes formed but failed to enclose anaphase-bridged chromatin. Time-lapse imaging showed two phenotypes: anaphase-bridged chromatin that eventually resolved, and segregated chromatin that returned to the midzone. Thus, the assembly of BAF, lamins, and LEM-domain proteins is mutually dependent, and is required to capture segregated chromosomes within the nascent nuclear envelope. Embryos that escaped lethality by down-regulation of Ce-BAF grew into sterile adults with misplaced distal tip cells and gonads, further suggesting that mild postembryonic reductions in BAF disrupt tissue-specific functions.

Dynamic interaction between BAF and emerin revealed by FRAP, FLIP, and FRET analyses in living HeLa cells.

Barrier-to-autointegration factor (BAF) is a conserved 10 kDa DNA-binding protein. BAF interacts with LEM-domain proteins including emerin, LAP2 beta, and MAN1 in the inner nuclear membrane. Using fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP), we compared the mobility of BAF to its partners emerin, LAP2 beta, and MAN1 in living HeLa cells. Like endogenous BAF, GFP-BAF was enriched at the nuclear envelope, and found inside the nucleus and in the cytoplasm during interphase. At every location, FRAP and FLIP analysis showed that GFP-BAF diffused rapidly; the halftimes for recovery in a 0.8 microm square area were 260 ms at the nuclear envelope, and even faster inside the nucleus and in the cytoplasm. GFP-fused emerin, LAP2 beta, and MAN1 were all relatively immobile, with recovery halftimes of about 1 min, for a 2 microm square area. Thus, BAF is dynamic and mobile during interphase, in stark contrast to its nuclear envelope partners. FLIP results further showed that rapidly diffusing cytoplasmic and nuclear pools of GFP-BAF were distinctly regulated, with nuclear GFP-BAF unable to replenish cytoplasmic BAF. Fluorescence resonance energy transfer (FRET) results showed that CFP-BAF binds directly to YFP-emerin at the inner nuclear membrane of living cells. We propose a "touch-and-go" model in which BAF binds emerin frequently but transiently during interphase. These findings contrast with the slow mobility of both GFP-BAF and GFP-emerin during telophase, when they colocalized at the 'core' region of telophase chromosomes at early stages of nuclear assembly.

MAN1 and emerin have overlapping function(s) essential for chromosome segregation and cell division in Caenorhabditis elegans.

Emerin and MAN1 are LEM domain-containing integral membrane proteins of the vertebrate nuclear envelope. The function of MAN1 is unknown, whereas emerin is known to interact with nuclear lamins, barrier-to-autointegration factor (BAF), nesprin-1 alpha, and a transcription repressor. Mutations in emerin cause X-linked recessive Emery-Dreifuss muscular dystrophy. Emerin and MAN1 homologs are both conserved in Caenorhabditis elegans, but loss of Ce-emerin has no detectable phenotype. We therefore used C. elegans to test the hypothesis that Ce-MAN1 overlaps functionally with Ce-emerin. Supporting this model, Ce-MAN1 interacted directly with Ce-lamin and Ce-BAF in vitro and required Ce-lamin for its nuclear envelope localization. Interestingly, RNA interference-mediated removal of approximately 90% of Ce-MAN1 was lethal to approximately 15% of embryos. However, in the absence of Ce-emerin, approximately 90% reduction of Ce-MAN1 was lethal to all embryos by the 100-cell stage, with a phenotype involving repeated cycles of anaphase chromosome bridging and cytokinesis ["cell untimely torn" (cut) phenotype]. Immunostaining showed that the anaphase-bridged chromatin specifically retained a mitosis-specific phosphohistone H3 epitope and failed to recruit detectable Ce-lamin or Ce-BAF. These findings show that LEM domain proteins are essential for cell division and that Ce-emerin and Ce-MAN1 share at least one and possibly multiple overlapping functions, which may be relevant to Emery-Dreifuss muscular dystrophy.