Abstract
The Golgi apparatus is a critical effector of tumor cell polarization and directional motility during EMT, but how the organization and functions of the Golgi are regulated during EMT remains unclear. Here, we present a quantitative assay to measure the structure-function dynamics of the Golgi apparatus at below the diffraction limit, using HyVolution - a combination of high sensitivity detector equipped confocal platforms to acquire diffraction limited high-resolution images using defined sets of acquisition parameters and subsequent computational super-resolution by image deconvolution. HyVolution has been reported to achieve a lateral resolution of 140 nm, which is approximately 1.6 times higher than the resolution achievable with conventional confocal microscopy. This assay is amenable to fixed and live cell and tissue imaging formats, using commercially available fluorophores and enabled screening of the Golgi morphology and function in panels of cell lines. Using this approach, we found that the EMT- activating transcription factor ZEB1 leads to the formation of a polarized and compact Golgi structure with improved ribbon linking, enhanced Golgi matrix connectivity, and increased anterograde vesicle trafficking. Progestin and adiponectin q receptor 11 (PAQR11), a ZEB1 derepressed Golgi-resident protein is required for ZEB1 to induce the structure-function changes in the Golgi, anterograde trafficking, cell migration and metastasis. Prevalent optical microscopy based assays of Golgi structure and function allows diffraction-limited quantitative assessment of Golgi compaction but does not allow precise measurement of Golgi element size and number, ribbon linking or compares compactness across several cell types. Our HyVolution based technique allowed comparison of panels of murine and human lung cancer cells based on their Golgi compaction, element size, number, ribbon linking and correlate the same with cell migration, invasion and metastasis.
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