A low-cost cure for CUREs: An undergraduate microbiology course engaging students in authentic research using publicly available datasets.

Undergraduate research experiences are key to preparing STEM students for a range of careers and graduate programs, and to impacting retention in STEM. Providing undergraduate research experiences can be challenging for institutions due to the high cost associated with equipment and reagents, lab space, and research mentors. In this study, we present an upper-level microbiology seminar course that does not require these resources, as each student chooses and performs their own research project using data obtained from publicly available datasets. The faculty member provides hands-on instruction and regular feedback to mentor the cohort of students through all stages of their research projects, from honing a research question, to choosing a dataset, to data analysis and visualization. Students build science communication skills through each writing a scientific paper, and creating and presenting a scientific poster. These papers and presentations, along with results from student pre- and post-surveys, demonstrate that students built research and communication skills, while also building their confidence and interest in science careers. To access this research experience, students only need to register for this course; no application or selection is required, and no prior research experience is expected. The use of publicly available data makes this course a low-cost way to integrate authentic research projects into the college curriculum, and can be adapted to courses in any discipline. Such "low-cost CUREs" (course-based undergraduate research experiences) can be used to build capacity for undergraduate research experiences that are so crucial to preparing students for opportunities in and beyond college.

A Novel Undergraduate Seminar Course Celebrating Scientific Contributions by Scientists from Historically Marginalized Communities.

Scientific contributions by members from historically marginalized communities (HMCs) have been largely ignored, uncredited, and in some cases erased from history. This has contributed to science, technology, engineering, and math (STEM) curricula lacking diversity. In this study, we present an Honors seminar course aimed to highlight the discoveries of scientists from HMCs, centered around reading primary literature in a way that builds our students' research skills. The course provides students with opportunities for active learning, skill building, and mentorship that are key for persistence of students in the STEM "leaky pipeline." Students also read biographies of scientists from HMCs, interact with guest speakers, and choose scientists to highlight (in final papers and presentations) and publicize (through the creation of Wikipedia pages). Additionally, students use community-building methodologies to build a safe classroom and gain tools to have conversations about diversity, inequities, and intersectionality in STEM. In self-reporting surveys, 93.7% of students strongly agreed that their appreciation for marginalized scientists increased and 92.6% reported that the course met very well the goal of refining their research skills. These findings support the effectiveness of this novel course. We provide two lists (one of 137 scientists and one of 57 scientist biographies) that will allow faculty teaching a wide range of science classes to select examples of scientists and discoveries to highlight in their courses. This course represents a novel platform to diversify STEM curricula while engaging and empowering students from historically marginalized communities.

The Health Equity Scholars Program: Innovation in the Leaky Pipeline.

Despite attempts to increase enrollment of under-represented minorities (URMs: primarily Black/African American, Hispanic/Latino, and Native American students) in health professional programs, limited progress has been made. Compelling reasons to rectify this situation include equity for URMs, better prepared health professionals when programs are diverse, better quality and access to health care for UMR populations, and the need for diverse talent to tackle difficult questions in health science and health care delivery. However, many students who initiate traditional "pipeline" programs designed to link URMs to professional schools in health professions and the sciences, do not complete them. In addition, program requirements often restrict entry to highly qualified students while not expanding opportunities for promising, but potentially less well-prepared candidates. The current study describes innovations in an undergraduate pipeline program, the Health Equity Scholars Program (HESP) designed to address barriers URMs experience in more traditional programs, and provides evaluative outcomes and qualitative feedback from participants. A primary outcome was timely college graduation. Eighty percent (80%) of participants, both transfer students and first time students, so far achieved this outcome, with 91% on track, compared to the campus average of 42% for all first time students and 58-67% for transfers. Grade point averages also improved (p = 0.056) after program participation. Graduates (94%) were working in health care/human services positions and three were in health-related graduate programs. Creating a more flexible program that admits a broader range of URMs has potential to expand the numbers of URM students interested and prepared to make a contribution to health equity research and clinical care.

Intragenic and extragenic suppressors of temperature sensitive mutations in the replication initiation genes dnaD and dnaB of Bacillus subtilis.

BACKGROUND: The Bacillus subtilis genes dnaD and dnaB are essential for the initiation of DNA replication and are required for loading of the replicative helicase at the chromosomal origin of replication oriC. Wild type DnaD and DnaB interact weakly in vitro and this interaction has not been detected in vivo or in yeast two-hybrid assays. METHODOLOGY/PRINCIPAL FINDINGS: We isolated second site suppressors of the temperature sensitive phenotypes caused by one dnaD mutation and two different dnaB mutations. Five different intragenic suppressors of the dnaD23ts mutation were identified. One intragenic suppressor was a deletion of two amino acids in DnaD. This deletion caused increased and detectable interaction between the mutant DnaD and wild type DnaB in a yeast two-hybrid assay, similar to the increased interaction caused by a missense mutation in dnaB that is an extragenic suppressor of dnaD23ts. We isolated both intragenic and extragenic suppressors of the two dnaBts alleles. Some of the extragenic suppressors were informational suppressors (missense suppressors) in tRNA genes. These suppressor mutations caused a change in the anticodon of an alanine tRNA so that it would recognize the mutant codon (threonine) in dnaB and likely insert the wild type amino acid (alanine). CONCLUSIONS/SIGNIFICANCE: The intragenic suppressors should provide insights into structure-function relationships in DnaD and DnaB, and interactions between DnaD and DnaB. The extragenic suppressors in the tRNA genes have important implications regarding the amount of wild type DnaB needed in the cell. Since missense suppressors are typically inefficient, these findings indicate that production of a small amount of wild type DnaB, in combination with the mutant protein, is sufficient to restore some DnaB function.

Multicopy plasmids affect replisome positioning in Bacillus subtilis.

The DNA replication machinery, various regions of the chromosome, and some plasmids occupy characteristic subcellular positions in bacterial cells. We visualized the location of a multicopy plasmid, pHP13, in living cells of Bacillus subtilis using an array of lac operators and LacI-green fluorescent protein (GFP). In the majority of cells, plasmids appeared to be highly mobile and randomly distributed. In a small fraction of cells, there appeared to be clusters of plasmids located predominantly at or near a cell pole. We also monitored the effects of the presence of multicopy plasmids on the position of DNA polymerase using a fusion of a subunit of DNA polymerase to GFP. Many of the plasmid-containing cells had extra foci of the replisome, and these were often found at uncharacteristic locations in the cell. Some of the replisome foci were dynamic and highly mobile, similar to what was observed for the plasmid. In contrast, replisome foci in plasmid-free cells were relatively stationary. Our results indicate that in B. subtilis, plasmid-associated replisomes are recruited to the subcellular position of the plasmid. Extending this notion to the chromosome, we postulated that the subcellular position of the chromosomally associated replisome is established by the subcellular location of oriC at the time of initiation of replication.

Control of DNA replication initiation by recruitment of an essential initiation protein to the membrane of Bacillus subtilis.

The Bacillus subtilis proteins DnaD and DnaB are essential for replication initiation and are conserved in low G+C content Gram-positive bacteria. Previous work indicated that DnaD and DnaB are involved in helicase loading during the process of restarting stalled replication forks. We have investigated the roles of DnaD and DnaB in replication initiation at oriC in vivo. Using chromatin immunoprecipitation (ChIP), we found that DnaD and DnaB functions are needed to load the replicative helicase at oriC. To investigate further the functions of DnaD and DnaB in replication initiation, we isolated and characterized suppressors of the temperature sensitivity of dnaD and dnaB mutant cells. In both cases, we isolated the identical missense mutation in dnaB, dnaBS371P. Using yeast two-hybrid analysis, we found that dnaBS371P uncovers a previously undetected physical interaction between DnaD and DnaB. We also found that DnaBS371P constitutively recruits DnaD to the membrane fraction of cells, where DnaB and oriC are enriched. Phenotypes of cells expressing DnaBS371P are consistent with aberrant replication control. We hypothesize that B. subtilis regulates replication initiation by regulating a physical interaction between two proteins essential for helicase loading at chromosomal origins.

Conserved and clustered RNA recognition sequences are a critical feature of signals directing RNA localization in Xenopus oocytes.

Although it is widely regarded that the targeting of RNA molecules to subcellular destinations depends upon the recognition of cis-elements found within their 3' untranslated regions (UTR), relatively little is known about the specific features of these cis-sequences that underlie their function. Interaction between specific repeated motifs within the 3' UTR and RNA-binding proteins has been proposed as a critical step in the localization of Vg1 RNA to the vegetal pole of Xenopus oocytes. To understand the relative contributions of repeated localization element (LE) sequences, we used comparative functional analysis of Vg1 LEs from two frog species, Xenopus laevis and Xenopus borealis. We show that clusters of repeated VM1 and E2 motifs are required for efficient localization. However, groups of either site alone are not sufficient for localization. In addition, we present evidence that the X. borealis Vg1 LE is recognized by the same set of RNA-binding proteins as the X. laevis Vg1 LE and is capable of productive interactions with the X. laevis transport machinery as it is sufficient to direct vegetal localization in X. laevis oocytes. These results suggest that clustered sets of cis-acting sites within the LE direct vegetal transport through specific interactions with the localization machinery.