Decoding the Variance in Integrated Intracellular Organization of the Undifferentiated Hips Cell

The Allen Institute for Cell Science is developing a state space of structural signatures of the undifferentiated human induced pluripotent stem cell (hiPSC) to understand how cells organize and transition between states (cellular morphogenesis). To do this we take advantage of the ∼35 endogenous fluorescently tagged hiPSC lines in the Allen Cell Collection (www.allencell.org), each expressing a monoallelic EGFP-tagged protein representing a particular organelle. We develop image-based assays and segmentations for quantitative analyses, taking advantage of thousands of replicate high-resolution 3D images for each structure. We are investigating biological sources of cellular variation in a dimensionally-reduced space that represents integrated intracellular organization. We applied the Allen Cell Structure Segmenter to images of lamin B1-tagged cells to extract nuclear shapes, which we then fit using spherical harmonics. We analyzed the contributions of the fitting coefficients to shape variations. We found that nuclear shape could be well described by five coefficients representing three distinct sources of biological variation. The first mode represented nuclear volume, which increases throughout interphase (∼one day timescale). The second mode represented how flat (vs. round) a nucleus appeared in the apical-basal axis (Z-direction), which was linked to differences in cell packing in distinct regions of the colony. High cell density regions contained taller cells with rounder nuclei. Individual nuclei exhibited little variation in flatness over a multi-hour time period, consistent with a timescale of several days for cell packing within colonies. The third mode represented how ‘squeezed’ a nucleus appeared in the XY plane due to interactions with neighboring cells occurring at timescales of minutes. We are now applying these analyses to develop biophysical models of nuclear shape. Our framework will be extended to cell shape and key intracellular structures in an integrative fashion.