Decoding the variance in 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 high-dimensional 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 performed principal component analysis on the fitting coefficients and we found the first five components explain 85% of the total variance. Each of these components was mapped into a mode corresponding to a distinct biological source of variation. The first mode represented nuclear volume, which increases throughout interphase at timescales of hours. The second mode represented how flat (vs. round) a nucleus appeared in the apical-basal axis (Z-direction), which was linked to changes in colony cell packing dynamics consistent with a timescale of several days for cell packing within colonies. The remaining modes represented how elongated a nucleus appeared in the XY plane and how tilted the nucleus appeared along the major and minor axis. We found these modes to be caused by 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.