Molokanova E , Savchenko A , Kramer RH .

R33 CA094304 NCI NIH HHS

	Figure 2. PTK inhibitors reduce the genistein inhibition. (A) Inhibition of closed rod photoreceptor CNG channels by 1 min pre-exposure to 100 μM genistein alone or together with PTK inhibitors (500 μM AMP-PNP or 100 μM erbstatin). Closed channels were activated by application of 2 mM cGMP (control). (B) Inhibition of fully activated channels by genistein alone or together with PTK inhibitors.
	Figure 1. Effect of genistein on native rod CNG channels. (A) Activation of CNG channels by saturating cGMP without genistein. (B) Effect of genistein on closed channels. Response to saturating cGMP after pre-exposure to 100 μM genistein for 1 min. The broken line distinguishes the slowly activating current component inhibited by genistein and the rapidly activating residual current. (C) Effect of 100 μM genistein on fully activated channels, indicated by the broken line.
	Figure 3. Comparison of genistein inhibition of native CNG channels. Dose-inhibition curves of the effect of genistein on native closed (A) and fully activated (B) CNG channels from rod and cone outer segments and olfactory receptor neurons. Continuous curves in A and B show fits of the data to the Hill equation (see materials and methods). (C) Bar graph showing Ki values for genistein inhibition of closed and activated channels from rods, cones, and olfactory neurons. Data represents mean ± SEM for all of the individual experiments included in A and B.
	Figure 4. Subunit dependence of genistein inhibition of closed rod CNG channels. (A) Dose-inhibition curves for genistein effect on closed native rod, homomeric RETα, and heteromeric RETα+β channels. (B) Bar graph showing Ki and Hill coefficient values for genistein inhibition curves. Data represents mean ± SEM for all individual experiments included in A.
	Figure 6. Kinetics of inhibition of fully activated homomeric RETα (A) and heteromeric RETα+β (B) channels by various concentrations of genistein.
	Figure 5. Subunit dependence of genistein inhibition of fully activated rod CNG channels. (A) Dose-inhibition curves for genistein effect on native and homomeric RETα, and heteromeric RETα+β channels. All channels were fully activated with saturating (2 mM) cGMP before genistein application. (B) Bar graph showing Ki and Hill coefficient values for genistein inhibition curves. Data represents mean ± SEM for all individual experiments included in A.
	Figure 9. Schematic diagram of genistein inhibition. Homomeric and heteromeric RET and OLF channels and their interactions with PTKs, symbolized with the shaded crescent symbol. Genistein (G) is depicted bound to the PTK. In the closed state of the homomeric RETα channel (A), two PTKs, and thus two genistein molecules, are required to completely inhibit the channel. cGMP triggers channel opening, which functionally dimerizes RETα subunits. With the PTK spanning two functional dimers, only one PTK, and therefore one genistein, is sufficient to inhibit the channel, although two are still able to bind. In addition, the affinity of the closed channel for the PTK is higher than for the open channel, hence a tighter fit between the channel and the PTK symbol is depicted for the closed channel. For the RETα+β channel (B), only the RETα binds the PTK. Hence there is no change in the number of PTK required to blocked the closed and open channels. The homomeric OLFα channel (C) is unaffected by genistein because OLFα subunits do not contain PTK binding sites. The heteromeric OLFα+OLFβ (D) channel binds one PTK, via the binding sites on the OLFβ subunits.
	Figure 7. Subunit dependence of genistein inhibition of olfactory CNG channels. Dose-inhibition curves for genistein effect on closed (A) and fully activated (B) native (n = 3–5), homomeric OLFα (n = 4), and heteromeric OLFα+β (n = 6) channels.
	Figure 8. Summary data for genistein inhibition of various expressed homomeric and heteromeric channels in closed (A) and fully activated (B) states. n = 5–8 for each channel type.

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