Navigating a research career with a disability.

In recent years, we have seen an increasing focus in the academic environment on equity, diversity and inclusion. However, one broad group often left out of these discussions are disabled scientists/scientists with disabilities, who often face severe challenges entering the research profession and navigating their careers. Building on the success of the 2022 Young Embryologist Network's meeting, which included a session on 'Working in science with a disability' ( Morgan, 2023) we learn here from the lived experiences of five biologists who share the challenges and successes of undertaking a scientific career with a disability, as well as accommodations that can make science, technology, engineering, mathematics and medicine (STEMM) careers more accessible and inclusive.

Alveolar progenitor differentiation and lactation depends on paracrine inhibition of notch via ROBO1/CTNNB1/JAG1.

In the mammary gland, how alveolar progenitor cells are recruited to fuel tissue growth with each estrus cycle and pregnancy remains poorly understood. Here, we identify a regulatory pathway that controls alveolar progenitor differentiation and lactation by governing Notch activation in mouse. Loss of Robo1 in the mammary gland epithelium activates Notch signaling, which expands the alveolar progenitor cell population at the expense of alveolar differentiation, resulting in compromised lactation. ROBO1 is expressed in both luminal and basal cells, but loss of Robo1 in basal cells results in the luminal differentiation defect. In the basal compartment, ROBO1 inhibits the expression of Notch ligand Jag1 by regulating ß-catenin (CTNNB1), which binds the Jag1 promoter. Together, our studies reveal how ROBO1/CTTNB1/JAG1 signaling in the basal compartment exerts paracrine control of Notch signaling in the luminal compartment to regulate alveolar differentiation during pregnancy.

SLIT Loss or Sequestration Increases Mammary Alveologenesis and Lactogenesis.

SLITs comprise a family of secreted proteins that function as ligands for Roundabout (ROBO) receptors. Previous research showed that ROBO1 promotes the differentiation of milk-producing alveolar cells by inhibiting Notch signaling in mammary luminal cells. Here, we show enhanced alveolar development and increased milk production in Slit2-/-;Slit3-/- knockout mammary gland epithelia. This result can also be achieved by intraperitoneal delivery of recombinant ROBO1 extracellular domain fragment, ROBO1-5Ig-Fc, which sequesters SLITs. Together, our phenotypic studies suggest that SLITs restrict alveologenesis and lactogenesis by inhibiting ROBO1.

A Suite of Mouse Reagents for Studying Amelogenesis.

Amelogenesis, the formation of dental enamel, is driven by specialized epithelial cells called ameloblasts, which undergo successive stages of differentiation. Ameloblasts secrete enamel matrix proteins (EMPs), proteases, calcium, and phosphate ions in a stage-specific manner to form mature tooth enamel. Developmental defects in tooth enamel are common in humans, and they can greatly impact the well-being of affected individuals. Our understanding of amelogenesis and developmental pathologies is rooted in past studies using epithelial Cre driver and knockout alleles. However, the available mouse models are limited, as most do not allow targeting different ameloblast sub-populations, and constitutive loss of EMPs often results in severe phenotype in the mineral, making it difficult to interpret defect mechanisms. Herein, we report on the design and verification of a toolkit of twelve mouse alleles that include ameloblast-stage specific Cre recombinases, fluorescent reporter alleles, and conditional flox alleles for the major EMPs. We show how these models may be used for applications such as sorting of live stage specific ameloblasts, whole mount imaging, and experiments with incisor explants. The full list of new alleles is available at https://dev.facebase.org/enamelatlas/mouse-models/ .

SRSF1 governs progenitor-specific alternative splicing to maintain adult epithelial tissue homeostasis and renewal.

Alternative splicing generates distinct mRNA variants and is essential for development, homeostasis, and renewal. Proteins of the serine/arginine (SR)-rich splicing factor family are major splicing regulators that are broadly required for organ development as well as cell and organism viability. However, how these proteins support adult organ function remains largely unknown. Here, we used the continuously growing mouse incisor as a model to dissect the functions of the prototypical SR family protein SRSF1 during tissue homeostasis and renewal. We identified an SRSF1-governed alternative splicing network that is specifically required for dental proliferation and survival of progenitors but dispensable for the viability of differentiated cells. We also observed a similar progenitor-specific role of SRSF1 in the small intestinal epithelium, indicating a conserved function of SRSF1 across adult epithelial tissues. Thus, our findings define a regulatory mechanism by which SRSF1 specifically controls progenitor-specific alternative splicing events to support adult tissue homeostasis and renewal.

Salivary gland: A budding genius.

During salivary gland development, branches arise through formation of new end buds (budding) and division of existing buds (clefting). In a recent Cell study, Wang et al., (2021) report that mouse salivary gland budding is reliant on a delicate interplay between epithelial cells and the extracellular matrix surrounding them.

Generation of Mosaic Mammary Organoids by Differential Trypsinization.

Organoids offer self-organizing, three-dimensional tissue structures that recapitulate physiological processes in the convenience of a dish. The murine mammary gland is composed of two distinct epithelial cell compartments, serving different functions: the outer, contractile myoepithelial compartment and the inner, secretory luminal compartment. Here, we describe a method by which the cells comprising these compartments are isolated and then combined to investigate their individual lineage contributions to mammary gland morphogenesis and differentiation. The method is simple and efficient and does not require sophisticated separation technologies such as fluorescence activated cell sorting. Instead, we harvest and enzymatically digest the tissue, seed the epithelium on adherent tissue culture dishes, and then use differential trypsinization to separate myoepithelial from luminal cells with ~90% purity. The cells are then plated in an extracellular matrix where they organize into bilayered, three-dimensional (3D) organoids that can be differentiated to produce milk after 10 days in culture. To test the effects of genetic mutations, cells can be harvested from wild type or genetically engineered mouse models, or they can be genetically manipulated prior to 3D culture. This technique can be used to generate mosaic organoids that allow investigation of gene function specifically in the luminal or myoepithelial compartment.

Loss of miR-203 regulates cell shape and matrix adhesion through ROBO1/Rac/FAK in response to stiffness.

Breast tumor progression is accompanied by changes in the surrounding extracellular matrix (ECM) that increase stiffness of the microenvironment. Mammary epithelial cells engage regulatory pathways that permit dynamic responses to mechanical cues from the ECM. Here, we identify a SLIT2/ROBO1 signaling circuit as a key regulatory mechanism by which cells sense and respond to ECM stiffness to preserve tensional homeostasis. We observed that Robo1 ablation in the developing mammary gland compromised actin stress fiber assembly and inhibited cell contractility to perturb tissue morphogenesis, whereas SLIT2 treatment stimulated Rac and increased focal adhesion kinase activity to enhance cell tension by maintaining cell shape and matrix adhesion. Further investigation revealed that a stiff ECM increased Robo1 levels by down-regulating miR-203. Consistently, patients whose tumor expressed a low miR-203/high Robo1 expression pattern exhibited a better overall survival prognosis. These studies show that cells subjected to stiffened environments up-regulate Robo1 as a protective mechanism that maintains cell shape and facilitates ECM adherence.

Variables psicológicas y adhesión en personas con VIH: evaluación en función del tiempo de infección / Psychological variables and adherence among HIV persons: evaluation based on length of infection

Se realizó un estudio transversal con el objetivo de evaluar la influencia de un conjunto de variables psicológicas sobre los comportamientos de adhesión al tratamiento en función del tiempo de infección en meses. Participaron 93 personas con VIH, quienes contestaron dos instrumentos: 1) Variables psicológicas y comportamientos de adhesión, y 2) Situaciones vinculadas con estrés. Se emplearon los siguientes procedimientos estadísticos: la ji al cuadrado de Pearson, la U de Mann-Whitney y un análisis de regresión múltiple. Los análisis de regresión revelaron una influencia diferenciada de las variables psicológicas en función del tiempo de infección: en el grupo de personas con < 42 meses, los predictores de los comportamientos de adhesión fueron una buena motivación, un óptimo desempeño competencial presente y bajos niveles de estrés vinculados con tolerancia a la frustración. En el grupo con 43 o más meses los predictores fueron una buena motivación y bajos niveles de estrés vinculados con toma de decisiones. El tiempo de infección en meses constituye una variable crítica respecto de la forma en que operan las variables psicológicas sobre la práctica de los comportamientos de adhesión en personas con VIH expuestas a tratamiento con medicamentos antirretrovirales.

This cross-sectional study was carried out to assess the influence of psychological variables on antiretroviral treatment adherence behaviors, based on length of infection in months. Participants included ninety-three HIV-positive persons, who answered two self-administered questionnaires: 1) Psychological variables and adherence behaviors, and 2) Stress-related situations. Three consecutive statistical testing procedures were applied for data analysis: Pearson's chi-square, Mann-Whitney U, and multiple regressions. Regression analyses found psychological variables influencing adherence behaviors in different ways based on length of infection: in the group of persons with < 42 months, psychological predictors were a good motivation, an optimal competential performance and low stress-related with tolerance to frustration, whereas in the group of persons with 43 or more months predictors were a good motivation and low stress-related with decision-making. Results strongly support the tenet that length of infection is a critical variable related to psychological variables influencing adherence behaviors among HIV-positive persons under antiretroviral treatment.