Figure 2.

cER Morphology in ER-PM MCS Tether Mutants

(A) Domain structure of the main ER-PM tethers. Ist2 is an ER multipass transmembrane protein with a long and presumably unstructured cytosolic tail. The C-terminal sorting signal (SS) binds the PM. Scs2 and Scs22 are ER transmembrane proteins containing an N-terminal MSP domain. Tcb proteins are anchored to the ER membrane by a hairpin sequence. In their cytoplasmic C-terminus, Tcbs contain an SMP domain and a variable number of C2 domains. Panels B through F show 1.4-nm-thick tomographic slices of cER in the indicated strains (left) and 3D renderings in two perpendicular orientations upon a 90° rotation along an axis parallel to the PM (right). cER: cortical ER (pink); Nuc: nucleus; PM: plasma membrane (gold). (B) WT cell, (C) Ist2-only cell, (D) Scs2/22-only cell, (E) Tcb1/2/3-only cell, (F) Δtether cell. Insets in (B) and (E) show cER peaks (blue arrowheads). Scale bars: 300 nm (main panels); 25 nm (insets). Panels G, H, and I show quantifications of cER-PM distance (G), cER thickness (H) and cER peak density per μm² of cER membrane area (I). In G and H the violin plots show the complete distribution of values for all MCS analyzed. A white dot represents the median, a black slab the interquartile range, and a black line 1.5 times the interquartile range. Panel I shows average values (gray bars) and SE (error bars). HS: heat shock (42°C for 10 min). ^∗, ^∗∗, and ^∗∗∗ indicate, respectively, p < 0.05, p < 0.01 and p < 0.01 by unpaired t test (G, H) or Mann-Whitney U test (I). N = 6 (WT), 7 (WT HS), 5 (Ist2-only), 5(Scs2/22-only), 9 (Tcb1/2/3-only), 5 (tcb1Δ), 5 (tcb2Δ), 5 (tcb3Δ), 5 (tcb1/2Δ), 5 (tcb1/2/3Δ), and 5 (tcb1/2/3Δ HS) cER-PM MCS. See also Figures S1 and S2; Table S1.