mother cell’s vacuole move into the bud where they fuse with one another to form a new vacuole. These vesicles carry both v-SNARES and t-SNARES. Are both types of SNARES essential for this homotypic fusion event? To test this point, you have developed an ingenious assay for fusion of vacuolar vesicles. You prepare vesicles from two different mutant strains of yeast: strain B has a defective gene for vacuolar alkaline phosphatase (Pase); strain A is defective for the protease that converts the pre- cursor of alkaline phosphatase (pro-Pase) into its active form (Pase) (Figure Q13-2A). Neither strain has active alkaline phosphatase, but when extracts of the strains are mixed, vesicle fusion generates active alkaline phospha- tase, which can be easily measured (Figure Q13-2). Now you delete the genes for the vacuolar v-SNARE, t-SNARE, or both in each of the two yeast strains. You prepare vacuolar vesicles from each and test them for their ability to fuse, as measured by the alkaline phospha- tase assay (Figure Q13-2B). What do these data say about the requirements for v-SNARES and t-SNAREs in the fusion of vacuolar vesicles? Does it matter which kind of SNARE is on which vesicle? Please note the two questions in the last paragraph of this are rhetorical (not meant to be answered directly). This is a true/false question: v-SNARES interact weakly with each other. O True O False (B) 100 alkaline phosphatase (% maximum) 75 50 25 0 t strain A strain B experiment 1 2 vt vt t t vt 3 vt V 7 vt V V t V vt 4 5 6 SNARE combinations t V V t vt 8 9 10 11 vt Figure Q13-2 SNARE requirements for vesicle fusion (Problem 13-7). (A) Scheme for measuring the fusion of vacuolar vesicles. (B) Results of fusions of vesicles with different combinations of v-SNARES and t-SNARES. The SNARES present on the vesicles of the two strains are indicated as v (v-SNARE) and t (t-SNARE). (Adapted from B.J. Nichols et al, Nature 387: 199-202, 1997. With permission from Macmillan Publishers Ltd.)
The Correct Answer and Explanation is :
The correct answer to the true/false question is:
False.
Explanation:
v-SNAREs and t-SNAREs are essential for vesicle fusion, and the interaction between v-SNAREs and t-SNAREs is specific and strong. They form a stable, complementary pair of SNARE complexes that drive the fusion of vesicles, such as in the homotypic fusion of vacuolar vesicles.
The key point in the experimental design is that vacuolar vesicles from two different yeast strains, A and B, can fuse to activate alkaline phosphatase when mixed. This process relies on vesicle fusion, which involves the interaction between SNARE proteins on the vesicles. Strain A lacks the protease to convert pro-Pase into active alkaline phosphatase, while strain B has a defective alkaline phosphatase gene. When the two strains’ vesicles fuse, the alkaline phosphatase enzyme becomes active, signaling successful vesicle fusion.
The experiment then tests the importance of specific SNAREs—v-SNAREs (on the vesicle membrane) and t-SNAREs (on the target membrane)—for this fusion event by deleting specific SNARE genes. The results show that vesicle fusion does not occur when either v-SNARE or t-SNARE is missing. When both SNARE types are absent from the vesicles of both strains, no fusion or activation of alkaline phosphatase occurs. However, fusion and enzyme activation do occur when either v-SNARE or t-SNARE is present on the appropriate vesicle, confirming the essential role of both types of SNAREs.
Moreover, the data indicate that it doesn’t matter which vesicle contains which SNARE type (v or t). The key requirement is that both types of SNAREs are present on the vesicles, regardless of which vesicle holds the v-SNARE or t-SNARE. The SNARE proteins must be on complementary vesicles to interact and enable fusion, and this interaction is strong, not weak. Therefore, v-SNAREs do not interact weakly with each other, which refutes the statement in the true/false question.