Exam 13: Intracellular Membrane Traffic

Which endocytic process is best depicted in the following schematic diagram?

In Drosophila melanogaster, loss-of-function mutations in either Pink1 or Parkin show similar phenotypes including impaired ability to fly, male sterility, and degeneration of dopaminergic neurons. Transgenic overexpression of Parkin in mutants lacking Pink1 significantly ameliorates the loss-of-function phenotype, but overexpression of Pink1 cannot rescue the Parkin loss-of-function phenotype. According to these findings, which protein is more likely to act upstream of the other one? Assuming that the Parkin loss-of-function phenotype is merely due to defects in autophagy, would you expect Parkin overexpression to also rescue an ATG9 loss-of-function phenotype?

The Golgi apparatus is made up of an ordered series of compartments. To process N-linked oligosaccharides, different Golgi compartments carry different enzymes that alter the sugar chains sequentially. Which Golgi cisternae are normally responsible for the addition of galactose and sialic acid, respectively, in complex oligosaccharides?

Reconstitution of vesicle transport in cell-free systems was historically carried out in the presence of isolated "donor" and "acceptor" Golgi stacks. The donor stacks are isolated from cells expressing a viral protein and lacking a processing enzyme. The protein can be transported to the acceptor stacks (that do contain the processing enzyme) and be processed only in the presence of an energy source (such as ATP) and a cytosolic fraction. The transport vesicles that mediate this process are visualized by microscopy. Various compounds can be added to this system to help understand the mechanism of transport. Two such compounds were added in early experiments in the 1980s, both of which blocked transport of the viral protein and resulted in the accumulation of transport vesicles. However, the accumulated vesicles following treatment with one of these compounds (A) appeared to be coated, while those resulting from treatment with the other compound (B) did not. If one of these compounds is a nonhydrolyzable GTP analog and the other one is N-ethylmaleimide, which compound (A or B) do you think represents the GTP analog? Write down A or B as your answer.

An animal cell has been wounded and has a small rupture in its plasma membrane. Which of the following is more likely to happen next?

You have engineered a fusion protein composed of a cis Golgi resident protein and the green fluorescent protein. Similarly, you have made a fusion protein composed of a trans Golgi protein and the red fluorescent protein. You express these proteins in cells and follow individual Golgi cisternae using fluorescence microscopy. You observe that each individual cisterna emits a green, faint yellow, or red fluorescent signal at different times: it initially shows green fluorescence, but the green fluorescent signal fades away with time, and instead red fluorescence appears in the same cisterna. Additionally, you do not observe any red fluorescent cisterna that becomes green over time. Do these observations agree better with the cisternal maturation model (C) or the vesicle transport model (V) for the organization of Golgi stacks? Write down C or V as your answer.

In mammals, colostrum (or "first milk") is produced by the mother in late pregnancy and shortly after giving birth to feed the newborn. In addition to nutrients, it is particularly rich in antibodies that are absorbed through the intestinal epithelium and support the weak immune system of the infant. Occasionally, some infants suffer from either respiratory acidosis or alkalosis. In acidosis, the blood pH becomes acidic due to lack of sufficient ventilation, while the opposite happens in alkalosis due to hyperventilation. Considering the molecular mechanism of transcytosis in intestinal epithelia, which of these conditions-acidosis (C) or alkalosis (L)-would you expect to interfere more with antibody absorption by transcytosis in these infants? Write down C or L as your answer.

Aspartic acid residues resemble phosphorylated serine residues in proteins and also carry a similar negative charge. As a result, a protein in which serine residues are mutated to aspartic acid residues may mimic the function of the corresponding wild-type protein when those serines are phosphorylated. Given that phosphorylation on serine residues in a number of Golgi matrix proteins occurs during mitosis, would you expect that mutation of these serines to aspartic acids would prevent Golgi stack formation in interphase (1) or prevent Golgi dispersion in mitosis (2)? Write down 1 or 2 as your answer.

Rab5 and Rab7 constitute a Rab cascade in the process of endosome maturation. One of the Rab5 effectors is a Rab7-GEF, while one of the Rab7 effectors is a Rab5-GAP. Which of these proteins would you expect to find in early endosomes? Write down Rab5 or Rab7 as your answer.

Neurotransmitters are retrieved from the synaptic cleft soon after their release from the presynaptic axon terminal. Do you think treatment of neurons with the drug dynasore, a dynamin inhibitor, would affect this retrieval positively (P) or negatively (N)? Write down P or N as your answer.

What is the effect of defective or missing N-acetylglucosamine phosphotransferase on lysosomal protein sorting?

Indicate true (T) and false (F) statements below regarding secretory vesicles. Your answer would be a four-letter string composed of letters T and F only, e.g. TTTF. ( ) Proteins destined for secretory vesicles (for regulated secretion) often aggregate in the lumen of the TGN. ( ) The lumen of secretory vesicles is generally less acidic than the lumen of the TGN from which they originate. ( ) A secretory vesicle starts recycling Golgi components only after it has budded from the TGN and its coat proteins have disassembled. ( ) Secretory vesicles recycle Golgi components to the TGN in small clathrin-coated vesicles.

In the following schematic diagram depicting the formation of vesicular tubular clusters between the ER and the CGN, what major coat proteins are indicated by 1 and 2, respectively?

Indicate whether each of the following descriptions better applies to receptor-mediated endocytosis (R), phagocytosis (F), pinocytosis (P), or macropinocytosis (M). Your answer would be a four-letter string composed of letters R, F, P, and M only, e.g. MMPF. ( ) This pathway depends on caveolin and cavin proteins. ( ) Pathogenic particles such as SV40, papillomavirus, and cholera toxin enter the cell via this pathway. ( ) In animals, this pathway is normally limited to professional cells such as neutrophils. ( ) This pathway proceeds by the formation of highly ruffled regions in the plasma membrane which then collapse, resulting in fluid uptake.