Question

Consider cargo proteins that are destined for secretion to the extracellular space. Please predict where the cargo will end up (ex. in the cytoplasm, a specific organelle, transport vesicles, multiple places, extracellular space) for: (i) & (ii) Each individual condition; & (iii) The combination of both conditions.

C.

(i) v-SNAREs on COPII coated vesicles are mutated to be unable to bind t-SNAREs

(ii) A transmembrane domain is added to the cargo protein.

(iii) Both
D.

(i) Receptors in the Golgi are mutated so they are unable to bind to cargo proteins.

(ii) COPI coat proteins are mutated so they are unable to bind to membranes.

(iii) Both

Answer 1

Transport vesicles must ocuur in a highly selective manner so that they recognize the correct target membrane with which to fuse. Owing to large diversity of membrane systems, a vesicle may likely encounter many potential target membranes before it finds the correct one. SNARE proteins seem to have a central role both in providing specificity and in catalyzing the fusion of vesicles with the target membrane. There are at least 20 different SNAREs in an animal cell and they exist as complementary sets vesicle membrane SNAREs, called v-SNAREs, and target membrane SNAREs, called t-SNAREs which when interact with each other form stable trans-SNARE complexes, which lock the two membranes (both of the vesicle and the target memebrane) together. On the other hand, transport vesicles bud off as coated vesicles having distinctive proteins covering their cytosolic surface. Before the vesicle fuses with a target membrane, the coat is discarded, as is required to allow the two cytosolic membrane surfaces to interact directly and fuse. There are three well-characterized types of coated vesicles, which differ in their coat proteins: clathrin-coated, COPI-coated, and COPII-coated vesicles. Each type is used for different transport steps, different cargo delivery route in the cell. COPI-coated packages bud from pre-Golgi compartments and Golgi cisternae and COPII-coated vesicles bud from the ER and transport proteins to the golgi apparatus. Therefore, if

C. i) the v-SNAREs on COPII coated vesicles are mutated to be unable to bind t-SNAREs then vesicles with the cargo proteins will bud off from the ER but will never be able to recognize the cognate t-SNARE on the golgi apparatus. Cargo will remain inside the vesicles.

ii) a transmembrane domain is added to the cargo protein, then it will remain embedded in the membrane of of ER. Once the transport vesicles bud off they will remain associated with the membrane till they reach Golgi apparatus. Thereafter it will be transferred once the appropriate trans-SNARE complex forms.

iii) both occurs then the cargo proteins will remain embedded in the membrane of transport vesicles as the vesicles will not fuse with their target membranes.    

Therefore, if

D. i) the receptors in the Golgi are mutated so they are unable to bind to cargo proteins then cargo proteins will be released but not enter into the golgi apparatus. So, they may get diffuse in the cytoplasm.

ii)  COPI coat proteins are mutated so they are unable to bind to membranes then the cargo proteins will remain in the vesicles. COPI proteins mediate a retrograde transport pathway that selectively recycles proteins from the cis-Golgi complex to the ER. So, the vesicles will remain in and around the periphery of intra-golgi network.

iii) both occurs, then cargo proteins will not be transported back to the ER, so it will remain in the cytoplasm.