Biology subjects

Chase, S. E.

Publications and source records attributed to Chase, S. E..

3 recordsLinked to original sources

bioRxiv2022

Loss of Myosin-1e biases MMTV-PyMT induced breast cancer towards a 4 differentiated and secretory state

Expression of the unconventional myosin, Myosin-1e (Myo1e), has been shown to contribute to tumor progression in the MMTV-PyMT mouse model of mammary tumorigenesis and is associated with poor outcome in breast cancer patients. However, the specific effects of Myo1e expression on the mammary tumor cells remain unidentified. Here, we used Myo1e-KO and wild-type (WT) MMTV-PyMT mice on a pure genetic background to further investigate the molecular and cellular effects of Myo1e expression. Myo1e-WT tumors were characterized by an increased abundance of intra-epithelial macrophages and lower amounts of the extracellular matrix. Transcriptomic profiling of WT and Myo1e-KO tumors identified a pattern of differential expression of tumor suppressor and tumor-promoting genes that was consistent with the observed differences in tumor progression and morphology between the genotypes, and also revealed differential expression of genes associated with secretion and cell-cell adhesion. In agreement with the RNA-seq findings, Myo1e-expressing tumor cells exhibited increased proliferation and elevated nuclear enrichment of YAP1 transcriptional activator compared to Myo1e-KO tumor cells. To investigate tumor cell-autonomous effects of Myo1e expression, we used the epithelial cell line PY-230 derived from the MMTV-PyMT-induced mouse tumor to create Myo1e-depleted cells by Crispr-mediated genome editing. Cells deficient in Myo1e had increased expression of genes encoding milk components compared to the wild-type cells. Electric cell-substrate impedance sensing (ECIS) measurements showed that depletion of Myo1e in PY-230 cells resulted in increased resistance to electrical current indicating enhanced epithelial barrier function. Overall, we find that Myo1e expression biases tumors towards a less-differentiated, pro-tumorigenic state, and that depletion of Myo1e is associated with a pro-secretory, more differentiated state.

cancer biology↗

bioRxiv2022

Myo1e knockout in the adult podocytes leads to proteinuria but has less severe consequences for kidney function than Myo1e loss during renal development

Myosin 1e (Myo1e) is expressed in specialized epithelial cells in the kidney (podocytes) and plays an important role in renal filtration. Knockout of Myo1e in mice and mutations in the MYO1E gene in humans cause proteinuria and disrupt the ultrastructure of glomeruli, the initial segments of renal nephrons that are responsible for selective excretion of waste products without the loss of proteins from the bloodstream. Previous studies have demonstrated that the loss of Myo1e early in development results in severe defects in renal function in mice and may lead to end-stage renal disease in patients homozygous for MYO1E mutations. However, little is known about the effects of Myo1e loss later in life. In this study, we used inducible knockout of Myo1e in mouse podocytes to examine the effects of Myo1e loss from the adult kidneys. We have found that Myo1e loss after the completion of renal development causes proteinuria and podocyte defects but the effects are milder and more variable compared to the effects of Myo1e knockout in developing podocytes. These findings indicate that Myo1e plays an important role in podocyte development and differentiation but may also contribute to the maintenance of the glomerular barrier in the adult kidneys.

cell biology↗

bioRxiv2021

Steroid-resistant nephrotic syndrome associated MYO1E mutations have differential effects on myosin 1e localization, dynamics, and activity

Myo1e is a non-muscle motor protein enriched in the podocyte foot processes. Mutations in MYO1E are associated with steroid-resistant nephrotic syndrome (SRNS). Here, we set out to differentiate between the pathogenic and neutral MYO1E variants identified in SRNS patients by exome sequencing. Based on protein sequence conservation and structural predictions, two mutations in the motor domain, T119I and D388H, were selected for this study. EGFP-tagged Myo1e constructs were delivered into the Myo1e-KO podocytes via adenoviral infection to analyze Myo1e protein stability, Myo1e localization, and clathrin-dependent endocytosis, which is known to involve Myo1e activity. Furthermore, truncated Myo1e constructs were expressed using the baculoviral expression system and used to measure Myo1e ATPase and motor activity in vitro. Both mutants were expressed as full-length proteins in the Myo1e-KO podocytes. However, unlike wild-type (WT) Myo1e, the T119I variant was not enriched at the cell junctions or clathrin-coated vesicles (CCVs) in podocytes. In contrast, the D388H variant localization was similar to the WT. Surprisingly, the dissociation of the D388H variant from cell-cell junctions and CCVs was decreased, suggesting that this mutation also affects Myo1e activity. The ATPase activity and the ability to translocate actin filaments were drastically reduced for the D388H mutant, supporting the findings from cell-based experiments. The experimental pipeline developed in this study allowed us to determine that the T119I and D388H mutations appear to be pathogenic and gain additional knowledge in the Myo1e role in podocytes. This workflow can be applied to the future characterization of novel MYO1E variants associated with SRNS.

cell biology↗