1. Roles of RER and Golgi:
   • ER removes signal peptide: solubilises protein.
   • ER swaps C-terminal signal for membrane tether: attaches protein (loosely) to membrane.
   • ER adds glycosylation: glucose ‘fuse’ marks proteins as misfolded.
   • ER contains chaperonins and PDI: ensures proteins are correctly folded, crosslinked, and do not aggregate.
   • ER dislocates broken proteins via BiP: ensures terminally misfolded proteins are destroyed.
   • Golgi modifies glycosylation: prepares protein for ECM, by making it more acidic, hydrophilic and protease-resistant, and also able to bind appropriate receptors.
   • Golgi phosphorylates mannose: required for lysosomal transport.
2. Small hydrophobic molecules may enter the nucleus by diffusion. Proteins enter via the nuclear pores in the Ran cycle: these must have a suitable signal peptide such as PPKKKRKV.
3. Both mitochondria and chloroplast use TIM/TOM-like systems, with an amphipathic N-terminal signal. However, plastids require further signals for lumenal import, and the sequences must be distinguishable.
4. Both the nucleus and peroxisomes import their proteins pre-folded. However, peroxisomes import them through relatively small proteins complexes, whilst the nucleus has very large and complex pores.
5. Nuclear-directed proteins (including the nuclear pore proteins themselves) must be able to relocate into the nucleus after mitosis, because they are released into the cytoplasm during the breakdown of the nuclear envelope during prometaphase. Likewise the KDEL sequence for RER retention: proteins are continually re-trafficked from the CGN to the RER. However, the RER signal can be cleaved because proteins won’t be re-imported.
6. Having the RER import signal at the amino terminus allows proteins to be imported into the RER cisternae as they are translated: this saves energy, because the physical force of translation can be used to push the protein through. Note that all other import mechanisms require either GTP, ATP or a proton motive force.
7. The peroxisome is highly oxidising. Isomery of disulfide bonds would be difficult under such circumstances, so perhaps this is why proteins have to be imported prefolded and fully crosslinked.
8. Vesicular trafficking could be demonstrated using GFP fusion proteins. Cisternal maturation could be demonstrated by showing that the Golgi traffics molecules too big to fit in a vesicle.
9. Cells and their endosymbionts have plenty of machinery for translocating proteins, but the only machinery for translocating RNAs across membranes is the nuclear pore, which would be a rather more difficult system to subvert for use by endosymbionts.