Image-based siRNA screen to identify kinases regulating Weibel-Palade body size control using electroporation

Area of Science:

• Cell Biology
• Molecular Biology
• Genomics

Background:

• Understanding subcellular organelle biogenesis and function is crucial in cell biology.
• Kinase inhibitors play a significant role in elucidating these cellular mechanisms.
• High-content screening provides a powerful approach for large-scale biological investigations.

Purpose of the Study:

• To present a human kinome siRNA high-content screen for identifying mechanisms of organelle biogenesis and function.
• To describe a novel, open-source, automated image analysis workflow for high-content analysis of subcellular organelles.
• To provide a valuable dataset for analyzing morphological parameters in primary human umbilical vein endothelial cells.

Main Methods:

• Performed a human kinome siRNA high-content screen using primary human umbilical vein endothelial cells (HUVECs).
• Utilized electroporation for efficient cell transfection.
• Developed and described an open-source, automated image analysis workflow for confocal fluorescence microscopy data.

Main Results:

• Generated a confocal fluorescence microscopic image dataset from the siRNA screen.
• The automated workflow enables reproducible and efficient high-content analysis of organelle morphology.
• The dataset is suitable for investigating morphological parameters linked to HUVEC biology.

Conclusions:

• The presented siRNA screen and analysis workflow facilitate the study of organelle biogenesis and function.
• This resource supports research into endothelial cell biology and the role of kinases.
• The open-source nature of the workflow promotes broader application in cell imaging studies.