Transitioning Cell Culture CURE Labs from Campus to Online: Novel Strategies for a Novel Time.

Course-based undergraduate research experiences (CUREs) provide a way for students to gain research experience in a classroom setting. Few examples of cell culture CUREs or online CUREs exist in the literature. The Cell Biology Education Consortium (CBEC) provides a network and resources for instructors working to incorporate cell-culture based research into the classroom. In this article, we provide examples from six instructors from the CBEC network on how they structure their cell-culture CUREs and how they transitioned the labs to online in the spring semester of 2020. We intend for these examples to provide instructors with ideas for strategies to set up cell culture CUREs, how to change that design mid-term, and for creating online CUREs in the future.

Photogeologic Map of the Perseverance Rover Field Site in Jezero Crater Constructed by the Mars 2020 Science Team.

The Mars 2020 Perseverance rover landing site is located within Jezero crater, a ∼ 50 km diameter impact crater interpreted to be a Noachian-aged lake basin inside the western edge of the Isidis impact structure. Jezero hosts remnants of a fluvial delta, inlet and outlet valleys, and infill deposits containing diverse carbonate, mafic, and hydrated minerals. Prior to the launch of the Mars 2020 mission, members of the Science Team collaborated to produce a photogeologic map of the Perseverance landing site in Jezero crater. Mapping was performed at a 1:5000 digital map scale using a 25 cm/pixel High Resolution Imaging Science Experiment (HiRISE) orthoimage mosaic base map and a 1 m/pixel HiRISE stereo digital terrain model. Mapped bedrock and surficial units were distinguished by differences in relative brightness, tone, topography, surface texture, and apparent roughness. Mapped bedrock units are generally consistent with those identified in previously published mapping efforts, but this study's map includes the distribution of surficial deposits and sub-units of the Jezero delta at a higher level of detail than previous studies. This study considers four possible unit correlations to explain the relative age relationships of major units within the map area. Unit correlations include previously published interpretations as well as those that consider more complex interfingering relationships and alternative relative age relationships. The photogeologic map presented here is the foundation for scientific hypothesis development and strategic planning for Perseverance's exploration of Jezero crater.

Riboflavin Riboswitch Regulation: Hands-On Learning about the Role of RNA Structures in the Control of Gene Expression in Bacteria.

American Society for Microbiology (ASM) Curriculum Guidelines highlight the importance of instruction about informational flow in organisms, including regulation of gene expression. However, foundational central dogma concepts and more advanced gene regulatory mechanisms are challenging for undergraduate biology students. To increase student comprehension of these principles, we designed an activity for upper-level biology students centered on construction and analysis of physical models of bacterial riboswitches. Students manipulate an inexpensive bag of supplies (beads, pipe cleaners) to model two conformations of a riboswitch in a bacterial transcript. After initial pilot testing, we implemented the activity in three upper-level classes at one research-intensive and two primarily undergraduate institutions. To assess student perceptions of learning gains, we utilized a pre/post-activity 5-point Likert-type survey instrument to characterize student perceptions of confidence in both their understanding of riboswitches and their ability to apply the central dogma to riboswitches. Median post-test ranks were significantly higher than median pre-test ranks (p < 0.0001) when compared by the Wilcoxon signed-rank test (n = 31). Next, we assessed post-activity knowledge via use of a rubric to score student responses on exam questions. More than 80% of students could correctly describe and diagram examples of riboswitches; data from initial iterations were used to enhance curriculum materials for subsequent implementations. We conclude that this riboswitch activity leads to both student-reported increases in confidence in the ASM curriculum dimension of gene regulation, including central dogma concepts, and demonstrated student ability to diagram riboswitches, predict outcomes of riboswitches, and connect riboswitches to evolutionary roles.

Students "Tackle" Quantitative Literacy in their Science Communication with Real-World Football Activity.

Undergraduate introductory biology students struggle when communicating quantitative data. This activity provides students with a real-world research experience to improve their quantitative literacy in science communication. Students were provided with a national sports media report that described a professional football athlete requiring 9,000 calories daily. Students were then asked to determine whether, based on their own research and calculations, the reporter had correctly calculated the total calories coming from the reported foods. Students discovered that their different sources of caloric information provided very different (albeit accurate) calculated totals, ranging from 6,000 to 11,000 calories. Importantly, the students generated professional letters outlining their calculated differences and sent them to the sports reporter. The professional letters to the reporter were assessed via rubric for accuracy of calculations, appropriate research evidence, professionalism, and readability for a nonexpert. A majority of the students provided accurate calculations; however, students scored lower on their professional writing skills, ability to cite appropriate research evidence, and readability for a nonexpert. Additionally, summative quantitative problems were individually completed and assessed, and activity cohorts achieved significantly higher on these problems compared with the non-activity cohort. Finally, surveyed students indicated that the activity helped prepare them for quantitative problems on the summative exam and helped them identify major course learning objectives. In conclusion, given an authentic research activity, students can take ownership of their learning and practice their communication to the general public about quantitative scientific information.

The education of Ehrenfried Walther von Tschirnhaus (1651-1708).

Ehrenfried Walther von Tschirnhaus, mathematician, inventor, and correspondent of Spinoza, is often thought to have studied medicine at Leiden, though documentation of this fact has been lacking. Tschirnhaus' medical education is here documented, along with the nature of his medical practice.

Serum deprivation inhibits the transcriptional co-activator YAP and cell growth via phosphorylation of the 130-kDa isoform of Angiomotin by the LATS1/2 protein kinases.

Large tumor suppressor (LATS)1/2 protein kinases transmit Hippo signaling in response to intercellular contacts and serum levels to limit cell growth via the inhibition of Yes-associated protein (YAP). Here low serum and high LATS1 activity are found to enhance the levels of the 130-kDa isoform of angiomotin (Amot130) through phosphorylation by LATS1/2 at serine 175, which then forms a binding site for 14-3-3. Such phosphorylation, in turn, enables the ubiquitin ligase atrophin-1 interacting protein (AIP)4 to bind, ubiquitinate, and stabilize Amot130. Consistently, the Amot130 (S175A) mutant, which lacks LATS phosphorylation, bound AIP4 poorly under all conditions and showed reduced stability. Amot130 and AIP4 also promoted the ubiquitination and degradation of YAP in response to serum starvation, unlike Amot130 (S175A). Moreover, silencing Amot130 expression blocked LATS1 from inhibiting the expression of connective tissue growth factor, a YAP-regulated gene. Concordant with phosphorylated Amot130 specifically mediating these effects, wild-type Amot130 selectively induced YAP phosphorylation and reduced transcription of connective tissue growth factor in an AIP4-dependent manner versus Amot130 (S175A). Further, Amot130 but not Amot130 (S175A) strongly inhibited the growth of MDA-MB-468 breast cancer cells. The dominant-negative effects of Amot130 (S175A) on YAP signaling also support that phosphorylated Amot130 transduces Hippo signaling. Likewise, Amot130 expression provoked premature growth arrest during mammary cell acini formation, whereas Amot130 (S175A)-expressing cells formed enlarged and poorly differentiated acini. Taken together, the phosphorylation of Amot130 by LATS is found to be a key feature that enables it to inhibit YAP-dependent signaling and cell growth.

Amot130 adapts atrophin-1 interacting protein 4 to inhibit yes-associated protein signaling and cell growth.

The adaptor protein Amot130 scaffolds components of the Hippo pathway to promote the inhibition of cell growth. This study describes how Amot130 through binding and activating the ubiquitin ligase AIP4/Itch achieves these effects. AIP4 is found to bind and ubiquitinate Amot130 at residue Lys-481. This both stabilizes Amot130 and promotes its residence at the plasma membrane. Furthermore, Amot130 is shown to scaffold a complex containing overexpressed AIP4 and the transcriptional co-activator Yes-associated protein (YAP). Consequently, Amot130 promotes the ubiquitination of YAP by AIP4 and prevents AIP4 from binding to large tumor suppressor 1. Amot130 is found to reduce YAP stability. Importantly, Amot130 inhibition of YAP dependent transcription is reversed by AIP4 silencing, whereas Amot130 and AIP4 expression interdependently suppress cell growth. Thus, Amot130 repurposes AIP4 from its previously described role in degrading large tumor suppressor 1 to the inhibition of YAP and cell growth.

Proteomic studies of Syk-interacting proteins using a novel amine-specific isotope tag and GFP nanotrap.

Green fluorescent protein (GFP) and variants have become powerful tools to study protein localization, interactions, and dynamics. We present here a mass spectrometry-based proteomics strategy to examine protein-protein interactions using anti-GFP single-chain antibody V(H)H in a combination with a novel stable isotopic labeling reagent, isotope tag on amino groups (iTAG). We demonstrate that the single-chain V(H)H (GFP nanotrap) allows us to identify interacting partners of the Syk protein-tyrosine kinase bearing a GFP epitope tag with high efficiency and high specificity. Interacting proteins identified include CrkL, BLNK, α- and ß-tubulin, Csk, RanBP5 and DJ-1. The iTAG reagents were prepared with simple procedures and characterized with high accuracy in the determination of peptides in model peptide mixtures and as well as in complex mixture. Applications of the iTAG method and GFP nanotrap to an analysis of the nucleocytoplasmic trafficking of Syk led to the identification of location-specific associations between Syk and multiple proteins. While the results reveal that the new quantitative proteomic strategy is generally applicable to integrate protein interaction data with subcellular localization, extra caution should be taken in evaluating the results obtained by such affinity purification strategies as many interactions appear to occur following cell lysis.