Figure 1.

Live-Cell Imaging of M. oryzae IH and Rice Cell Membranes.

(A) and (B) Differential staining patterns with RB and FM4-64 in rice sheath epidermal cells invaded by EYFP-labeled fungal strain KV1 (green). Arrows indicate sites where appressoria had penetrated into host cells.

(A) RB dye (purple) stained the endoplasmic reticulum inside rice cells and fungal IH at 36 hpi. Bar = 10 μm.

(B) PM and endocytotic membranes in rice cells were stained to saturation with FM4-64 (red). Narrow primary hyphae (P) extending from the penetration site differentiated into bulbous IH inside two invaded rice cells at 27 hpi. This image is a three-dimensional projection of 20 optical sections acquired with a z-interval of 0.44 μm. Bar = 10 μm.

(C) to (E) FM4-64 outlines IH but is not internalized by them. These images show separate and merged fluorescence channels for the upper rice cell in Figure 1B. Shown are EYFP fluorescence (C), FM4-64 fluorescence (white in this image) (D), and merged channels (E). Bars = 5 μm.

(F) to (H) Invasive-like hyphae formed in vitro on dialysis membrane internalize FM4-64. Shown are EYFP fluorescence (F), FM4-64 fluorescence (red) (G), and merged channels (H). Bars = 10 μm.

(I) The membrane encasing the IH had an FM4-64–stained connection (arrow) to rice membrane at the cell periphery. This is an enlarged view of a single optical section from the infection site in (C) to (E). The bright-field channel is included in this view (gray scale). Bar = 5 μm.

(J) and (K) TEM images show EIHM surrounding an IH inside an epidermal cell.

(J) Transverse section of an IH at 26 hpi. The arrow indicates a fibrillar inclusion inside the generally close-fitting EIHM. RC, rice cell. Bar = 500 nm.

(K) High-magnification view of the IH–host interface from the cell in (J). FCW, fungal cell wall; FPM, fungal plasma membrane. Bar = 150 nm.