Supplemental Material

This article contains the following supporting material:

• Figure 1 - 3D reconstruction of the ER.
  Wild type (NY2647) cells expressing the ER marker Sec61-GFP were grown at 25°C in SC medium. Z-stacks of cells were taken using a confocal microscope. The reconstruction was performed using Imaris program (BitPlan).

• Figure 2 - 3D reconstruction of the ER.
  rtn1Δ mutant (NY2648) cells expressing the ER marker Sec61-GFP were grown at 25°C in SC medium. Z-stacks of cells were taken using a confocal microscope. The reconstruction was performed using Imaris program (BitPlan).

• Figure 3 - Effect of rtn1Δ on other organelles.
  Wild type and rtn1Δ mutant cells expressing the late Golgi marker Sec7-GFP (NY2652, NY2653), the vacuolar marker Vph1-GFP (NY2655, NY2654) or the mitochondrial marker Atp9-DsRed (NY2657, NY2656) were grown at 25°C in SC medium with 2% glucose. Cells were examined using an epifluorescence microscope.

• Figure 4 - 3D reconstruction of the ER.
  Wild type (NY2658) cells expressing Rtn1-GFP were grown at 25°C in SC medium. Z-stacks of cells were taken using a confocal microscope. The reconstruction was performed using Imaris program.

• Figure 5 - The RTN2 gene is induced by 1M NaCl.
  A) A wild type strain expressing Rtn2-GFP (NY2680) was grown at 25°C in SC medium with 2% glucose. The culture was divided in two with 1M NaCl added to one half and the cells were left to grow for two hrs. Cells were examined using an epifluorescence microscope. B) A wild type strain expressing Rtn2-3HA (NY2681) or an untagged control strain (NY1210) were grown at 25°C in SC medium with 2% glucose. The culture was divided in two with 1M NaCl added to one half and the cells were left to grow for two hrs. Total extracts were prepared, loaded on a SDS-PAGE and probed with HA antibody.

• Figure 6 - RTN2 induction does not suppress rtn1Δ.
  A) Wild type (NY2647), rtn1Δ (NY2648) and rtn1Δrtn2Δ (NY2682) cells expressing the ER marker Sec61-GFP were grown at 25°C in SC medium. Two hrs prior to examining the cells using an epifluorescence microscope, 1M NaCl was added to the cultures. Arrows point to cortical regions devoid of fluorescence. B) Quantification of the cortical ER phenotype was done by scoring at least 300 cells with wild type or altered ER morphology, defined as the presence of large gaps in fluorescence at the cell cortex and a more cisternal appearance of the cortical ER. The graph shows the percentage of cells having wild type ER in each strain.