Biology subjects

Waschow, M.

Publications and source records attributed to Waschow, M..

3 recordsLinked to original sources

bioRxiv2019

Induced aneuploidy disrupts MCF10A acini formation and CCND1 expression

Abnormal karyotypes, namely aneuploidy, can be detected in nearly all cancer entities at different grades. The impact of these altering mutations on epigenetic regulation, especially on promoter-enhancer interactions are not well understood. Here, we applied a 3D model of MCF10A cells in a high-content screen to measure induced aneuploidy by RNA interference of 82 mitotic genes associated with aneuploidy and breast cancer. Perturbation of ESPL1 and TOP2A expression led to increased mitotic instability and subsequent aneuploidy and polylobed nuclei. During acinus formation these polylobed cells disrupted proper acinus rotation inhibiting the development of a hollow lumen and a polarized outer cell layer. Further, gene expression profiling identified upregulated CCND1 among other breast cancer related genes. We show that acquisition of aneuploidy affects the morphogenesis of MCF10A acini and expression of cancer relevant genes. By conducting 4C chromosome capturing experiments we linked the alteration of interactions of the promoter region to CCND1 upregulation.

cancer biology↗

bioRxiv2018

pheno-seq - linking 3D phenotypes of clonal tumor spheroids to gene expression

3D-culture systems have advanced cancer modeling by reflecting physiological characteristics of in-vivo tissues, but our understanding of functional intratumor heterogeneity including visual phenotypes and underlying gene expression is still limited. Single-cell RNA-sequencing is the method of choice to dissect transcriptional tumor cell heterogeneity in an unbiased way, but this approach is limited in correlating gene expression with contextual cellular phenotypes.\n\nTo link morphological features and gene expression in 3D-culture systems, we present pheno-seq for integrated high-throughput imaging and transcriptomic profiling of clonal tumor spheroids. Specifically, we identify characteristic EMT expression signatures that are associated with invasive growth behavior in a 3D breast cancer model. Additionally, pheno-seq determined transcriptional programs containing lineage-specific markers that can be linked to heterogeneous proliferative capacity in a patient-derived 3D model of colorectal cancer. Finally, we provide evidence that pheno-seq identifies morphology-specific genes that are missed by scRNA-seq and inferred single-cell regulatory states without acquiring additional single cell expression profiles. We anticipate that directly linking molecular features with patho-phenotypes of cancer cells will improve the understanding of intratumor heterogeneity and consequently be useful for translational research.

genomics↗

bioRxiv2017

Unraveling mitotic protein networks by 3D multiplexed epitope drug screening

Three-dimensional protein localization intricately determines the functional coordination of cellular processes. The complex spatial context of protein landscape has been assessed by multiplexed immunofluorescent staining1-3 or mass spectrometry4, applied to 2D cell culture with limited physiological relevance5 or tissue sections. Here, we present 3D SPECS, an automated technology for 3D Spatial characterization of Protein Expression Changes by microscopic Screening. This workflow encompasses iterative antibody staining of proteins, high-content imaging, and machine learning based classification of mitotic states. This is followed by mapping of spatial protein localization into a spherical, cellular coordinate system, the basis used for model-based prediction of spatially resolved affinities of various mitotic proteins. As a proof-of-concept, we mapped twelve epitopes in 3D cultured epithelial breast spheroids and investigated the network effects of mitotic cancer drugs with known limited success in clinical trials6-8. Our approach reveals novel insights into spindle fragility and global chromatin stress, and predicts unknown interactions between proteins in specific mitotic pathways. 3D SPECSs ability to map potential drug targets by multiplexed immunofluorescence in 3D cell cultured models combined with our automized high content assay will inspire future functional protein expression and drug assays.

systems biology↗