Panel A shows fluorescence microscopy images and fluorescence intensity profiles comparing endogenous Col1A1, Hsp47, and GalNT localization within intracellular regions. The horizontal axes represent distance in micrometers, while the vertical axes represent fluorescence intensity in arbitrary units. Panel B shows a scatter plot quantifying Col1A1 fluorescence intensities within endoplasmic reticulum and Golgi regions under transforming growth factor beta treatment conditions. The vertical axis represents Col1A1 intensity in arbitrary units, while the horizontal axis represents organelle and treatment categories. Panel C shows fluorescence microscopy images and zoomed insets illustrating colocalization between endogenous Col1A1 and Hsp47 chaperone proteins. Panel D shows fluorescence microscopy images and zoomed insets displaying colocalization between endogenous Col1A1 and calreticulin proteins. Panel E shows fluorescence microscopy images and zoomed insets illustrating colocalization between endogenous Col1A1 and PDIA1 proteins. Panel F shows fluorescence microscopy images and zoomed insets displaying Col1A1-mNeonGreen and PDIA6 protein colocalization patterns. Panel G shows fluorescence microscopy images and zoomed insets illustrating Col1A1-mNeonGreen and calnexin colocalization patterns within cells. Panel H shows a scatter plot comparing Manders colocalization coefficients between Col1A1 and multiple chaperone proteins. The vertical axis represents colocalization coefficient values, while the horizontal axis represents chaperone protein categories. Panel I shows time-lapse microscopy images tracking dissipation of Col1A1 condensates after thapsigargin treatment over time intervals. Panel J shows a scatter plot quantifying condensate numbers per cell before and after thapsigargin treatment. The vertical axis represents number of condensates per cell, while the horizontal axis represents treatment conditions.
PC1 condensates are enriched in chaperones and sensitive to ER stress. (A) Fluorescence images of fixed LX-2 cells activated with TGF-β and stained for antibodies against endogenous Col1A1 (green), Hsp47 (magenta), and GalNT (orange) (top), and fluorescence intensity profiles (bottom) along the dashed white line. White arrows show distinct Col1A1 pools in the ER and Golgi, demonstrating PC1 condensates are localized in the ER (scale bar: 10 µm). (B) Plot quantifying Col1A1 intensities in the ER and Golgi in LX-2 cells grown with or without TGF-β, showing increased collagen levels in both organelles upon TGF-β treatment. A total of 38 different cells with TGF-β and 34 cells without TGF-β from two independent experiments were analyzed (****P < 0.0001). (C) Fluorescence images of fixed LX-2 cells activated with TGF-β and stained for antibodies against endogenous Col1A1 (green) and Hsp47 (magenta) (top), and zoomed insets (bottom) (scale bar: 10 µm; inset: 2 µm). (D) Fluorescence images of fixed LX-2 cells activated with TGF-β and stained for antibodies against endogenous Col1A1 (green) and calreticulin (CALR) (magenta) (top), and zoomed insets (bottom) (scale bar: 10 µm; inset: 2 µm). (E) Fluorescence images of fixed LX-2 cells activated with TGF-β and stained for antibodies against endogenous Col1A1 (green) and protein disulfide isomerase A1 (PDIA1) (magenta) (top), and zoomed insets (bottom) (scale bar: 10 µm; inset: 2 µm). (F) Fluorescence images of fixed LX-2 cells stably expressing Col1A1-mNG (green) and protein disulfide isomerase A6 (PDIA6) (magenta) (top), and zoomed insets (bottom) (scale bar: 10 µm; inset: 2 µm). (G) Fluorescence images of fixed LX-2 cells stably expressing Col1A1-mNG (green) and calnexin (CANX) (magenta) (top), and zoomed insets (bottom) (scale bar: 10 µm; inset: 2 µm). (H) Manders’ colocalization coefficient M2 for Col1A1 and the respective chaperones. A total of 33, 15, 16, 24, and 17 cells for Hsp47, CALR, PDIA1, PDIA6, and CANX from two independent experiments were analyzed (ns—not significant; ****P < 0.0001). (I) Time-lapse images from Video 4 showing dissipation of PC1 condensates upon thapsigargin treatment (scale bar: 2 µm). (J) Plot quantifying PC1 condensate number per cell before and after thapsigargin treatment. A total 17 cells from two independent experiments were analyzed (****P < 0.0001).