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i-SNAREs: inhibitory SNAREs that fine-tune the specificity of membrane fusion.

Varlamov O, Volchuk A, Rahimian V, Doege CA, Paumet F, Eng WS, Arango N, Parlati F, Ravazzola M, Orci L, Söllner TH, Rothman JE - J. Cell Biol. (2003)

Bottom Line: A new functional class of SNAREs, designated inhibitory SNAREs (i-SNAREs), is described here.A striking pattern emerges in which certain subunits of the cis-Golgi SNAREpin function as i-SNAREs that inhibit fusion mediated by the trans-Golgi SNAREpin, and vice versa.Although the opposing distributions of the cis- and trans-Golgi SNAREs themselves could provide for a countercurrent fusion pattern in the Golgi stack, the gradients involved would be strongly sharpened by the complementary countercurrent distributions of the i-SNAREs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Biochemistry and Biophysics, Memorial Sloan-Kettering Cancer Center, 1275 York Ave., Box 251, New York, NY 10021, USA.

ABSTRACT
A new functional class of SNAREs, designated inhibitory SNAREs (i-SNAREs), is described here. An i-SNARE inhibits fusion by substituting for or binding to a subunit of a fusogenic SNAREpin to form a nonfusogenic complex. Golgi-localized SNAREs were tested for i-SNARE activity by adding them as a fifth SNARE together with four other SNAREs that mediate Golgi fusion reactions. A striking pattern emerges in which certain subunits of the cis-Golgi SNAREpin function as i-SNAREs that inhibit fusion mediated by the trans-Golgi SNAREpin, and vice versa. Although the opposing distributions of the cis- and trans-Golgi SNAREs themselves could provide for a countercurrent fusion pattern in the Golgi stack, the gradients involved would be strongly sharpened by the complementary countercurrent distributions of the i-SNAREs.

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i-SNAREs for the trans-Golgi fusion reaction. (A) Increasing concentrations of candidate i-SNAREs were incorporated into acceptor liposomes containing the ttrans complex. (B) Proteoliposomes were analyzed by SDS-PAGE and Coomassie blue staining. The position of each candidate i-SNARE is indicated by asterisks. (C) The resulting acceptor liposomes were incubated with donor liposomes containing the v-SNARE Sft1, and relative fusion activities are plotted as a function of the molar ratio of the candidate i-SNARE to the t-SNARE.
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fig3: i-SNAREs for the trans-Golgi fusion reaction. (A) Increasing concentrations of candidate i-SNAREs were incorporated into acceptor liposomes containing the ttrans complex. (B) Proteoliposomes were analyzed by SDS-PAGE and Coomassie blue staining. The position of each candidate i-SNARE is indicated by asterisks. (C) The resulting acceptor liposomes were incubated with donor liposomes containing the v-SNARE Sft1, and relative fusion activities are plotted as a function of the molar ratio of the candidate i-SNARE to the t-SNARE.

Mentions: The trans-Golgi fusion reaction (ttrans = Sed5/Ykt6, Gos1; vtrans = Sft1) is strongly inhibited by the vcis Bet1 (K50 = 0.4) and Tlg1 (K50 = 2.5), and is slightly inhibited by the tcis light chains Bos1 (K50 = 7.5) and Sec22 (K50 = 10; Fig. 3). In contrast, Vti1 and Snc1 had no significant effect (Fig. 3). During the course of our work, we noticed that a high concentration of Tlg1 inhibits incorporation of the t-SNAREs into liposomes. However, the lower concentrations of Tlg1 used for calculation of K50 did not have a significant effect on t-SNARE reconstitution, yet resulted in at least 50% inhibition of fusion (Fig. 2 B and Fig. 3 B; Tlg1, lanes 2 and 3).


i-SNAREs: inhibitory SNAREs that fine-tune the specificity of membrane fusion.

Varlamov O, Volchuk A, Rahimian V, Doege CA, Paumet F, Eng WS, Arango N, Parlati F, Ravazzola M, Orci L, Söllner TH, Rothman JE - J. Cell Biol. (2003)

i-SNAREs for the trans-Golgi fusion reaction. (A) Increasing concentrations of candidate i-SNAREs were incorporated into acceptor liposomes containing the ttrans complex. (B) Proteoliposomes were analyzed by SDS-PAGE and Coomassie blue staining. The position of each candidate i-SNARE is indicated by asterisks. (C) The resulting acceptor liposomes were incubated with donor liposomes containing the v-SNARE Sft1, and relative fusion activities are plotted as a function of the molar ratio of the candidate i-SNARE to the t-SNARE.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171956&req=5

fig3: i-SNAREs for the trans-Golgi fusion reaction. (A) Increasing concentrations of candidate i-SNAREs were incorporated into acceptor liposomes containing the ttrans complex. (B) Proteoliposomes were analyzed by SDS-PAGE and Coomassie blue staining. The position of each candidate i-SNARE is indicated by asterisks. (C) The resulting acceptor liposomes were incubated with donor liposomes containing the v-SNARE Sft1, and relative fusion activities are plotted as a function of the molar ratio of the candidate i-SNARE to the t-SNARE.
Mentions: The trans-Golgi fusion reaction (ttrans = Sed5/Ykt6, Gos1; vtrans = Sft1) is strongly inhibited by the vcis Bet1 (K50 = 0.4) and Tlg1 (K50 = 2.5), and is slightly inhibited by the tcis light chains Bos1 (K50 = 7.5) and Sec22 (K50 = 10; Fig. 3). In contrast, Vti1 and Snc1 had no significant effect (Fig. 3). During the course of our work, we noticed that a high concentration of Tlg1 inhibits incorporation of the t-SNAREs into liposomes. However, the lower concentrations of Tlg1 used for calculation of K50 did not have a significant effect on t-SNARE reconstitution, yet resulted in at least 50% inhibition of fusion (Fig. 2 B and Fig. 3 B; Tlg1, lanes 2 and 3).

Bottom Line: A new functional class of SNAREs, designated inhibitory SNAREs (i-SNAREs), is described here.A striking pattern emerges in which certain subunits of the cis-Golgi SNAREpin function as i-SNAREs that inhibit fusion mediated by the trans-Golgi SNAREpin, and vice versa.Although the opposing distributions of the cis- and trans-Golgi SNAREs themselves could provide for a countercurrent fusion pattern in the Golgi stack, the gradients involved would be strongly sharpened by the complementary countercurrent distributions of the i-SNAREs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular Biochemistry and Biophysics, Memorial Sloan-Kettering Cancer Center, 1275 York Ave., Box 251, New York, NY 10021, USA.

ABSTRACT
A new functional class of SNAREs, designated inhibitory SNAREs (i-SNAREs), is described here. An i-SNARE inhibits fusion by substituting for or binding to a subunit of a fusogenic SNAREpin to form a nonfusogenic complex. Golgi-localized SNAREs were tested for i-SNARE activity by adding them as a fifth SNARE together with four other SNAREs that mediate Golgi fusion reactions. A striking pattern emerges in which certain subunits of the cis-Golgi SNAREpin function as i-SNAREs that inhibit fusion mediated by the trans-Golgi SNAREpin, and vice versa. Although the opposing distributions of the cis- and trans-Golgi SNAREs themselves could provide for a countercurrent fusion pattern in the Golgi stack, the gradients involved would be strongly sharpened by the complementary countercurrent distributions of the i-SNAREs.

Show MeSH
Related in: MedlinePlus