Amphibian metamorphosis is driven by thyroid hormone (TH). We used prometamorphic tadpoles and a cell line of the African clawed frog (Xenopus laevis) to examine immediate effects of dioxin exposure on TH. Gene expression patterns suggest cross-talk between the thyroid hormone receptor (TR) and aryl hydrocarbon receptor (AHR) signaling pathways. In XLK-WG cells, expression of Cytochrome P450 1A6 (cyp1A6), an AHR target, was induced 1000-fold by 100 nM TCDD (2, 3, 7, 8 tetrachlorodibenzo-p-dioxin). Kru ¨ppel-Like Factor 9 (klf9), the first gene induced in a cascade of TH responses tied to metamorphosis, was upregulated over 5-fold by 50 nM triiodothyronine (T3) and 2-fold by dioxin. Co-exposure to T3 and TCDD boosted both responses, further inducing cyp1A6 by 75% and klf9 about 60%. Additional canonical targets of each receptor, including trba and trbb (TR) and udpgt1a (AHR) responded similarly. Induction of TH targets by TCDD in XLK-WG cells predicts that exposure could speed metamorphosis. We tested this hypothesis in two remodeling events: tail resorption and hind limb growth. Resorption of ex vivo cultured tails was accelerated by 10 nM T3, while a modest increase in resorption by 100 nM TCDD lacked statistical significance. Hind limbs doubled in length over four days following 1 nM T3 treatment, but limb length was unaffected by 100 nM TCDD. TCDD co-exposure reduced the T3 effect by nearly 40%, despite TCDD induction of klf9 in whole tadpoles, alone or with T3. These results suggest that tissue-specific TCDD effects limit or reverse the increased metamorphosis rate predicted by klf9 induction.
Glioblastomas (GBMs) are heterogeneous, treatment-resistant tumors driven by populations of cancer stem cells (CSCs). However, few molecular mechanisms critical for CSC population maintenance have been exploited for therapeutic development. We developed a spatially resolved loss-of-function screen in GBM patient-derived organoids to identify essential epigenetic regulators in the SOX2-enriched, therapy-resistant niche and identified WDR5 as indispensable for this population. WDR5 is a component of the WRAD complex, which promotes SET1 family-mediated Lys4 methylation of histone H3 (H3K4me), associated with positive regulation of transcription. In GBM CSCs, WDR5 inhibitors blocked WRAD complex assembly and reduced H3K4 trimethylation and expression of genes involved in CSC-relevant oncogenic pathways. H3K4me3 peaks lost with WDR5 inhibitor treatment occurred disproportionally on POU transcription factor motifs, including the POU5F1(OCT4)::SOX2 motif. Use of a SOX2/OCT4 reporter demonstrated that WDR5 inhibitor treatment diminished cells with high reporter activity. Furthermore, WDR5 inhibitor treatment and WDR5 knockdown altered the stem cell state, disrupting CSC in vitro growth and self-renewal, as well as in vivo tumor growth. These findings highlight the role of WDR5 and the WRAD complex in maintaining the CSC state and provide a rationale for therapeutic development of WDR5 inhibitors for GBM and other advanced cancers.
The development and implementation of a scientific outreach activity comes with a number of challenges. A successful outreach event must match the sophistication of content to the audience, be engaging, expand the knowledge base for participants, and be inclusive for a diverse audience. Ideally, a successful event will also convey the importance of scientific outreach for future scientists and citizens. In this paper, we present a simple, hands-on guide to a scientific outreach event targeted to kindergarten learners. This activity also pursued a second goal: the inclusion of undergraduate students in the development and delivery of the event. We provided a detailed set of four activities, focusing on the blind Mexican cavefish, which were enthusiastically received by kindergarten audiences. The engagement of undergraduate students in the development of this activity encouraged public outreach involvement and fostered new scientific and communication skills. The format of the outreach event we describe is flexible. We provide a set of guidelines and suggestions for adapting this approach to other biological topics. The activity and approach we describe enables the implementation of effective scientific outreach, using active learning approaches, which benefits both elementary school learners and undergraduate students.
Background Breast cancer (BC) is the most common cancer in women and the leading cause of cancer-associated mortality in women. In particular, triple-negative BC (TNBC) has the highest rate of mortality due in large part to the lack of targeted treatment options for this subtype. Thus, there is an urgent need to identify new molecular targets for TNBC treatment. RALA and RALB are small GTPases implicated in growth and metastasis of a variety of cancers, although little is known of their roles in BC. Methods The necessity of RALA and RALB for TNBC tumor growth and metastasis were evaluated in vivo using orthotopic and tail-vein models. In vitro, 2D and 3D cell culture methods were used to evaluate the contributions of RALA and RALB during TNBC cell migration, invasion, and viability. The association between TNBC patient outcome and RALA and RALB expression was examined using publicly available gene expression data and patient tissue microarrays. Finally, small molecule inhibition of RALA and RALB was evaluated as a potential treatment strategy for TNBC in cell line and patient-derived xenograft (PDX) models. Results Knockout or depletion of RALA inhibited orthotopic primary tumor growth, spontaneous metastasis, and experimental metastasis of TNBC cells in vivo. Conversely, knockout of RALB increased TNBC growth and metastasis. In vitro, RALA and RALB had antagonistic effects on TNBC migration, invasion, and viability with RALA generally supporting and RALB opposing these processes. In BC patient populations, elevated RALA but not RALB expression is significantly associated with poor outcome across all BC subtypes and specifically within TNBC patient cohorts. Immunohistochemical staining for RALA in patient cohorts confirmed the prognostic significance of RALA within the general BC population and the TNBC population specifically. BQU57, a small molecule inhibitor of RALA and RALB, decreased TNBC cell line viability, sensitized cells to paclitaxel in vitro and decreased tumor growth and metastasis in TNBC cell line and PDX models in vivo. Conclusions Together, these data demonstrate important but paradoxical roles for RALA and RALB in the pathogenesis of TNBC and advocate further investigation of RALA as a target for the precise treatment of metastatic TNBC.
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