Bridging the digital divide: Promoting equal access to online learning for health professions in an unequal world.

Online learning has the potential to enhance open and equitable access to medical education resources globally. Conversely, there are also concerns that it can perpetuate and exacerbate digital inequalities between developed (global North) and developing (global South) countries. In this article, we describe the historical lack of representation of the global South in the design of online medical education, as well as the resulting consequences and potential solutions. We compare the Northern and Southern views of online learning in medical education and identify the different types of barriers to its adoption. We describe how socioeconomic disparities and the historical dominance of the global North over the global South have led to systemic digital inequalities in the design and implementation of online learning in education generally, and in medical education particularly. The lack of representation of global South voices hinders the development of digital learning solutions relevant to local contexts, therefore limiting their effectiveness and sustainability. Thus, we propose approaches to build more equitable partnerships by soliciting local input and local expertise. Further, we discuss the need to maintain local relevance while setting global standards. Overall, we hope to inform and guide the development of more equitable and accessible online education training for a diverse global population.

Twelve tips for co-production of online learning.

In the digital age, experts in digital learning tools, or learning technologists (LTs), play an increasingly important role in the creation and delivery of online learning in health professions education. However, their expertise in the selection, curation and implementation of digital tools is often underutilized due to imbalanced relationships and lack of effective collaboration between faculty and LTs. Here, we describe how the co-production model can be applied to build equal and synergistic partnerships between faculty and LTs, so as to optimize the use of digital affordances and enhance online learning.

Interprofessional collaboration (or lack thereof) between faculty and learning technologists in the creation of digital learning.

BACKGROUND: As digital learning becomes more prevalent and important in health professions education, learning technologists play increasingly central roles in designing and delivering learning materials. However, little is understood about the process by which learning technologists have integrated into the existing teaching and learning ecosystem, and it seems that they remain marginal and undervalued. Our aim in this paper was therefore to examine the process of interprofessional co-development of course materials as experienced by educators and learning technologists. METHODS: Our approach was qualitative, using individual semi-structured interviews (conducted between July 2021 to May 2022) to explore the working relationship between faculty and learning technologists. Transcripts were analysed abductively. RESULTS: We found that the attitudes of both faculty and learning technologists towards collaborating to drive digital adoption in health professions education fell into two main themes: "embrace" and "replace" - and "conflict", which we present as a third theme. Our results revealed that faculty did not take an active and agentic role in developing their digital practices in respect of education delivery. Learning technologists positioned themselves as a resource to support faculty's knowledge and skill gap in digital competence. There was an obvious power differential between the two groups: learning technologists lacked agency and seemed in the position of servants to faculty masters. This created barriers to effective collaboration. CONCLUSIONS: By examining the process of co-development of course materials by faculty and learning technologists, we open up a space to examine the social, relational and organisational complexities associated with interprofessional collaboration in digital health professions education. Our study also has important implications for guiding educational policy to better position learning technologists to effectively collaborate with faculty and realise the potential of digital health professions education.

Epithelial Monolayers Coalesce on a Viscoelastic Substrate through Redistribution of Vinculin.

The mechanical properties of the microenvironment play a large role in influencing cellular behavior. In particular, the tradeoff between substrate viscosity and elasticity on collective cell migration by adherent cells is highly physiologically relevant, but remains poorly understood. To investigate the specific effects of viscous substrates, we plated epithelial monolayers onto polydimethylsiloxane substrata with a range of viscosities and elasticities. We found that on viscoelastic substrates the monolayers underwent rapid and coordinated movement to generate cell-free areas. To understand the molecular mechanism of this coordinated movement, we imaged various structural and signaling proteins at cell-cell and cell-matrix junctions. Through quantitative image analysis of monolayer disruption and subcellular protein redistribution, we show that the mechanosensor protein, vinculin, is necessary and sufficient for this viscous response, during which it is lost from focal adhesions and recruited by the cadherin complex to intercellular junctions. In addition, the viscous response is dependent upon and enhanced by actomyosin contractility. Our results implicate vinculin translocation in a molecular switching mechanism that senses substrate viscoelasticity and associates with actomyosin contractility.

RPTPα controls epithelial adherens junctions, linking E-cadherin engagement to c-Src-mediated phosphorylation of cortactin.

Epithelial junctions are fundamental determinants of tissue organization, subject to regulation by tyrosine phosphorylation. Homophilic binding of E-cadherin activates tyrosine kinases, such as Src, that control junctional integrity. Protein tyrosine phosphatases (PTPs) also contribute to cadherin-based adhesion and signaling, but little is known about their specific identity or functions at epithelial junctions. Here, we report that the receptor PTP RPTPα (human gene name PTPRA) is recruited to epithelial adherens junctions at the time of cell-cell contact, where it is in molecular proximity to E-cadherin. RPTPα is required for appropriate cadherin-dependent adhesion and for cyst architecture in three-dimensional culture. Loss of RPTPα impairs adherens junction integrity, as manifested by defective E-cadherin accumulation and peri-junctional F-actin density. These effects correlate with a role for RPTPα in cellular (c)-Src activation at sites of E-cadherin engagement. Mechanistically, RPTPα is required for appropriate tyrosine phosphorylation of cortactin, a major Src substrate and a cytoskeletal actin organizer. Expression of a phosphomimetic cortactin mutant in RPTPα-depleted cells partially rescues F-actin and E-cadherin accumulation at intercellular contacts. These findings indicate that RPTPα controls cadherin-mediated signaling by linking homophilic E-cadherin engagement to cortactin tyrosine phosphorylation through c-Src.

Cortactin scaffolds Arp2/3 and WAVE2 at the epithelial zonula adherens.

Cadherin junctions arise from the integrated action of cell adhesion, signaling, and the cytoskeleton. At the zonula adherens (ZA), a WAVE2-Arp2/3 actin nucleation apparatus is necessary for junctional tension and integrity. But how this is coordinated with cadherin adhesion is not known. We now identify cortactin as a key scaffold for actin regulation at the ZA, which localizes to the ZA through influences from both E-cadherin and N-WASP. Using cell-free protein expression and fluorescent single molecule coincidence assays, we demonstrate that cortactin binds directly to the cadherin cytoplasmic tail. However, its concentration with cadherin at the apical ZA also requires N-WASP. Cortactin is known to bind Arp2/3 directly (Weed, S. A., Karginov, A. V., Schafer, D. A., Weaver, A. M., Kinley, A. W., Cooper, J. A., and Parsons, J. T. (2000) J. Cell Biol. 151, 29-40). We further show that cortactin can directly bind WAVE2, as well as Arp2/3, and both these interactions are necessary for actin assembly at the ZA. We propose that cortactin serves as a platform that integrates regulators of junctional actin assembly at the ZA.

MOSH: An Online Tool for Real-Time Monitoring of Student Engagement.

Student engagement is important for classroom management but can be challenging to monitor, especially in large virtual classes. After lessons were moved online due to COVID-19 measures, instructors were unable to directly observe student behaviours, impacting their ability to gauge engagement levels and adjust the pace of delivery for optimal learning outcomes. A widget called MOSH, for Move On/Stay Here, was developed for students to indicate whether they wished to "move on" from or "stay here" on a point of discussion in real-time. By increasing acknowledgment of and response to student feedback, we aimed to enhance the student-instructor feedback loop.

The cytoskeleton and classical cadherin adhesions.

This chapter discusses the biochemical and functional links between classical cadherin adhesion systems and the cytoskeleton. Cadherins are best understood to cooperate with the actin cytoskeleton, but there is increasing evidence for the role of junctional microtubules in regulating cadherin biology. Cadherin adhesions and the junctional cytoskeleton are both highly dynamic systems that undergo continual assembly, turnover and remodeling, and yet maintain steady state structures necessary for intercellular adhesion. This requires the functional coordination of cadherins and cadherin-binding proteins, actin regulatory proteins, organizers of microtubule assembly and structure, and signaling pathways. These components act in concert to regulate junctional organization in response to extracellular forces and changing cellular contexts, which is essential for intercellular cohesion and tissue integrity.

Differential subcellular distributions and trafficking functions of hnRNP A2/B1 spliceoforms.

Trafficking of mRNA molecules from the nucleus to distal processes in neural cells is mediated by heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 trans-acting factors. Although hnRNP A2/B1 is alternatively spliced to generate four isoforms, most functional studies have not distinguished between these isoforms. Here, we show, using isoform-specific antibodies and isoform-specific green fluorescent protein (GFP)-fusion expression constructs, that A2b is the predominant cytoplasmic isoform in neural cells, suggesting that it may play a key role in mRNA trafficking. The differential subcellular distribution patterns of the individual isoforms are determined by the presence or absence of alternative exons that also affect their dynamic behavior in different cellular compartments, as measured by fluorescence correlation spectroscopy. Expression of A2b is also differentially regulated with age, species and cellular development. Furthermore, coinjection of isoform-specific antibodies and labeled RNA into live oligodendrocytes shows that the assembly of RNA granules is impaired by blockade of A2b function. These findings suggest that neural cells modulate mRNA trafficking by regulating alternative splicing of hnRNP A2/B1 and controlling expression levels of A2b, which may be the predominant mediator of cytoplasmic-trafficking functions. These findings highlight the importance of considering isoform-specific functions for alternatively spliced proteins.

Complex evolutionary relationships among four classes of modular RNA-binding splicing regulators in eukaryotes: the hnRNP, SR, ELAV-like and CELF proteins.

Alternative RNA splicing in multicellular organisms is regulated by a large group of proteins of mainly unknown origin. To predict the functions of these proteins, classification of their domains at the sequence and structural level is necessary. We have focused on four groups of splicing regulators, the heterogeneous nuclear ribonucleoprotein (hnRNP), serine-arginine (SR), embryonic lethal, abnormal vision (ELAV)-like, and CUG-BP and ETR-like factor (CELF) proteins, that show increasing diversity among metazoa. Sequence and phylogenetic analyses were used to obtain a broader understanding of their evolutionary relationships. Surprisingly, when we characterised sequence similarities across full-length sequences and conserved domains of ten metazoan species, we found some hnRNPs were more closely related to SR, ELAV-like and CELF proteins than to other hnRNPs. Phylogenetic analyses and the distribution of the RRM domains suggest that these proteins diversified before the last common ancestor of the metazoans studied here through domain acquisition and duplication to create genes of mixed evolutionary origin. We propose that these proteins were derived independently rather than through the expansion of a single protein family. Our results highlight inconsistencies in the current classification system for these regulators, which does not adequately reflect their evolutionary relationships, and suggests that a domain-based classification scheme may have more utility.

Functional implications of the emergence of alternative splicing in hnRNP A/B transcripts.

The heterogeneous nuclear ribonucleoproteins (hnRNPs) A/B are a family of RNA-binding proteins that participate in various aspects of nucleic acid metabolism, including mRNA trafficking, telomere maintenance, and splicing. They are both regulators and targets of alternative splicing, and the patterns of alternative splicing of their transcripts have diverged between paralogs and between orthologs in different species. Surprisingly, the extent of this splicing variation and its implications for post-transcriptional regulation have remained largely unexplored. Here, we conducted a detailed analysis of hnRNP A/B sequences and expression patterns across six vertebrates. Alternative exons emerged via the introduction of new splice sites, changes in the strengths of existing splice sites, and the accumulation of auxiliary splicing regulatory motifs. Observed isoform expression patterns could be attributed to the frequency and strength of cis-elements. We found a trend toward increased splicing variation in mammals and identified novel alternatively spliced isoforms in human and chicken. Pulldown and translational assays demonstrated that the inclusion of alternative exons altered the affinity of hnRNP A/B proteins for their cognate nucleic acids and modified protein expression levels. As the hnRNPs A/B regulate several key steps in mRNA processing, the involvement of diverse hnRNP isoforms in multiple cellular contexts and species implies concomitant differences in the transcriptional output of these systems. We conclude that the emergence of alternative splicing in the hnRNPs A/B has contributed to the diversification of their roles in the regulation of alternative splicing and has thus added an unexpected layer of regulatory complexity to transcription in vertebrates.

A visual framework for sequence analysis using n-grams and spectral rearrangement.

MOTIVATION: Protein sequences are often composed of regions that have distinct evolutionary histories as a consequence of domain shuffling, recombination or gene conversion. New approaches are required to discover, visualize and analyze these sequence regions and thus enable a better understanding of protein evolution. RESULTS: Here, we have developed an alignment-free and visual approach to analyze sequence relationships. We use the number of shared n-grams between sequences as a measure of sequence similarity and rearrange the resulting affinity matrix applying a spectral technique. Heat maps of the affinity matrix are employed to identify and visualize clusters of related sequences or outliers, while n-gram-based dot plots and conservation profiles allow detailed analysis of similarities among selected sequences. Using this approach, we have identified signatures of domain shuffling in an otherwise poorly characterized family, and homology clusters in another. We conclude that this approach may be generally useful as a framework to analyze related, but highly divergent protein sequences. It is particularly useful as a fast method to study sequence relationships prior to much more time-consuming multiple sequence alignment and phylogenetic analysis. AVAILABILITY: A software implementation (MOSAIC) of the framework described here can be downloaded from http://bioinformatics.org.au/mosaic/ CONTACT: m.ragan@uq.edu.au SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Functional diversity of the hnRNPs: past, present and perspectives.

The hnRNPs (heterogeneous nuclear ribonucleoproteins) are RNA-binding proteins with important roles in multiple aspects of nucleic acid metabolism, including the packaging of nascent transcripts, alternative splicing and translational regulation. Although they share some general characteristics, they vary greatly in terms of their domain composition and functional properties. Although the traditional grouping of the hnRNPs as a collection of proteins provided a practical framework, which has guided much of the research on them, this approach is becoming increasingly incompatible with current knowledge about their structural and functional divergence. Hence, we review the current literature to examine hnRNP diversity, and discuss how this impacts upon approaches to the classification of RNA-binding proteins in general.

Differential subnuclear localisation of hnRNPs A/B is dependent on transcription and cell cycle stage.

The heterogeneous nuclear ribonucleoproteins A1, A2/B1 and A3 (hnRNPs A/B) are involved in many nuclear functions that are confined to distinct regions within the nucleus. To characterise and compare the distribution of the hnRNPs A/B in these subnuclear compartments, their colocalisation with spliceosomal components, nascent transcripts and other nuclear markers in HeLa cells was investigated by immunostaining and transfection of GFP constructs. The mechanisms of this localisation were further explored by treating cells with detergent, nucleases and transcription inhibitors. We have also examined the dynamics of A2/B1 throughout the cell cycle. Our results show that hnRNPs A/B have different subnuclear localisations, with A1 differentially localised to the nuclear envelope, and A2/B1 and A3 enriched around nucleoli. This pattern of distribution was dependent on RNA integrity and active transcription. The hnRNPs A/B preferentially colocalised with a subset of splicing factors. Significantly, only rarely did transcription factories colocalise with high levels of these hnRNPs. Moreover, localisation of A2/B1 changed with cell cycle stage. Our findings show that the subnuclear localisation of the hnRNPs A/B is differentially, spatially and temporally regulated, and suggest that this localisation may be relevant to their nuclear functions.