Cancer Modeling by Transgene Electroporation in Adult Zebrafish (TEAZ)

Montal, Emily; Suresh, Shruthy; Ma, Yilun; Tagore, Mohita M.; White, Richard M.

doi:10.1007/978-1-0716-3401-1_5

Cited by 7 publications

(1 citation statement)

References 16 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Tumors were generated by Transgene Electroporation in Adult Zebrafish 71,84 (TEAZ). To generate hmgb2a / hmgb2b knockout melanomas, adult (3-6 month old) fish were injected with the following plasmids: miniCoopR-GFP, mitfa:Cas9, Tol2, U6-sgptena, U6-sgptenb, and either 394-zU6-3XsgRNA[hmgb2a] and 394-zU6-3XsgRNA[hmgb2b], or 394-zU6-3XsgRNA[NT].…”

Section: Zebrafish In Vivo Electroporationmentioning

confidence: 99%

Mechanical confinement governs phenotypic plasticity in melanoma

Hunter,

Montal,

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

Phenotype switching is a form of cellular plasticity in which cancer cells reversibly move between two opposite extremes - proliferative versus invasive states. While it has long been hypothesised that such switching is triggered by external cues, the identity of these cues has remained elusive. Here, we demonstrate that mechanical confinement mediates phenotype switching through chromatin remodelling. Using a zebrafish model of melanoma coupled with human samples, we profiled tumor cells at the interface between the tumor and surrounding microenvironment. Morphological analysis of these rare cells showed flattened, elliptical nuclei suggestive of mechanical confinement by adjacent tissue. Spatial and single-cell transcriptomics demonstrated that the interface cells adopted a gene program of neuronal invasion, including acquisition of an acetylated tubulin cage that protects the nucleus during migration. We identified the DNA-bending protein HMGB2 as a confinement-induced mediator of the neuronal state. HMGB2 is upregulated in confined cells, and quantitative modelling revealed that confinement prolongs contact time between HMGB2 and chromatin, leading to changes in chromatin configuration that favor the neuronal phenotype. Genetic disruption of HMGB2 showed that it regulates the trade-off between proliferative and invasive states, in which confined HMGB2hightumor cells are less proliferative but more drug resistant. Our results implicate the mechanical microenvironment as a mechanism driving phenotype switching in melanoma.

show abstract

Section: Zebrafish In Vivo Electroporationmentioning

confidence: 99%

Mechanical confinement governs phenotypic plasticity in melanoma

Hunter,

Montal,

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

Identifyingin vivogenetic dependencies of melanocyte and melanoma development

Perlee,

Ma,

Hunter

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

The advent of large-scale sequencing in both development and disease has identified large numbers of candidate genes that may be linked to important phenotypes. Validating the function of these candidates in vivo is challenging, due to low efficiency and low throughput of most model systems. This is especially the case in skin cells such as melanocytes, where the background mutation rate is high. We have developed a rapid and scalable system for assessing the role of candidate genes in a melanocyte specific manner using zebrafish. We generated transgenic zebrafish in which Cas9 was knocked-in to the endogenous mitfa locus, a master transcription factor of the melanocyte lineage. By introducing single guide RNA expression cassettes into mitfaCas9 embryos, we were able to achieve highly efficient melanocyte-specific mutation of genes important for melanocyte patterning and survival. These animals can be used to screen for dominant or recessive pigment defects in both the F0 generation (3 days) and F1 generation (3 months). We also utilized the mitfaCas9 line to study the role of melanoma genetic dependencies such as SOX10, demonstrating that loss of SOX10 reduces melanoma initiation yet promotes tumor progression by a switch to a SOX9hi state. This SOX10 to SOX9 switch has previously been observed in human patients, indicating that our system can be used to rapidly uncover biological states with relevance to human disease. Our high efficiency genetic approach can be readily applied to other cell lineages, with relevance to both development and disease.

show abstract

Restraint of melanoma progression by cells in the local skin environment

Ma,

Tagore,

Hunter

et al. 2024

Preprint

View full text Add to dashboard Cite

Keratinocytes, the dominant cell type in the melanoma microenvironment during tumor initiation, exhibit diverse effects on melanoma progression. Using a zebrafish model of melanoma and human cell co-cultures, we observed that keratinocytes undergo an Epithelial-Mesenchymal Transition (EMT)-like transformation in the presence of melanoma, reminiscent of their behavior during wound healing. Surprisingly, overexpression of the EMT transcription factor Twist in keratinocytes led to improved overall survival in zebrafish melanoma models, despite no change in tumor initiation rates. This survival benefit was attributed to reduced melanoma invasion, as confirmed by human cell co-culture assays. Single-cell RNA-sequencing revealed a unique melanoma cell cluster in the Twist-overexpressing condition, exhibiting a more differentiated, less invasive phenotype. Further analysis nominated homotypic jam3b-jam3b and pgrn-sort1a interactions between Twist-overexpressing keratinocytes and melanoma cells as potential mediators of the invasive restraint. Our findings suggest that EMT in the tumor microenvironment (TME) may limit melanoma invasion through altered cell-cell interactions.

show abstract

Cancer Modeling by Transgene Electroporation in Adult Zebrafish (TEAZ)

Cited by 7 publications

References 16 publications

Mechanical confinement governs phenotypic plasticity in melanoma

Mechanical confinement governs phenotypic plasticity in melanoma

Identifyingin vivogenetic dependencies of melanocyte and melanoma development

Restraint of melanoma progression by cells in the local skin environment

Contact Info

Product

Resources

About