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Chapter 5 Protein Synthesis, Processing, and Trafficking 55

The Unfolded Protein Response monitoring/regulating (1) transport competence of nascent proteins,
(2) capture of cargo in transport vesicles, and (3) protein retention/
The ER monitors the amount of unfolded protein in its lumen. retrieval for ER-localized proteins.
When that number exceeds a certain threshold, ER sensors activate
a signal transduction pathway. The set of responses activated by this
pathway is called the unfolded protein response (UPR). A number INTRAGOLGI TRANSPORT AND PROTEIN PROCESSING
of cellular insults disrupt protein folding and cause unfolded protein
accumulation in the ER lumen. The UPR is an adaptive response Organization of the Golgi Apparatus
signaled through three ER-localized transmembrane proteins PERK,
IRE1, and ATF6. These proteins function as sensors through the The Golgi complex comprises a stack of flattened, membrane-bound
properties of their ER-luminal domains and trigger a concerted cisternae that is highly dependent on microtubules for structural
response through the function of their cytosolic domains. The activa- integrity. The stack of cisternae can be subdivided into three parts
tion of the sensors results in a complex response aimed to (i) limit referred to as cis, medial, and trans with the cis and trans sides facing
accumulation of unfolded protein through reducing protein synthesis, the ER and the plasma membrane, respectively (see Fig. 5.4). Both
(ii) increasing the degradation of unfolded protein, and (iii) increas- the cis and trans faces are associated with tubulovesicular bundles of
ing the ER protein folding capacity. membranes. The ERGIC comprises the bundle on the cis side of the
IRE1 is conserved in all eukaryotic cells and has protein kinase Golgi stack and is the site where incoming proteins from the ER are
and endoribonuclease activities that, upon activation, mediate sorted into those directed for anterograde or for retrograde transport.
unconventional splicing of a 26-base intron from the XBP1 mRNA The tubulovesicular bundle at the trans side is the trans-Golgi network
to produce a basic Leucine Zipper (bZip) potent transcription factor. (TGN, see Fig. 5.4).
ATF6, upon accumulation of unfolded protein in the ER lumen, is A major feature of the Golgi is polarity. The processing events are
transported to the Golgi compartment where it is cleaved by two temporally and spatially ordered because the processing enzymes have
proteases, S1P and S2P. These enzymes release a cytosolic fragment a characteristic distribution across the Golgi stack. In the Golgi,
of ATF6 containing a bZip-transcription factor that migrates to the different types of modifications take place as for example proteolytic
nucleus to activate gene transcription. S1P and S2P are two important processing, protein O-glycosylation and elaboration of N-linked
Golgi proteases as they are also involved in the regulation of choles- chains, phosphorylation or sulfation of oligosaccharides, and sulfation
terol metabolism. Finally, PERK-mediated phosphorylation of eIF2α of tyrosines.
attenuates general mRNA translation; however, paradoxically, it The importance of protein glycosylation for human biology is
increases translation of the transcription factor ATF4 mRNA to also underlined by the identification of many inherited human disorders
induce transcription of UPR genes. If the UPR adaptive response is that are caused by defects in these processes and cause clinical mani-
not sufficient to correct the protein folding defect, the cells enter festations in members of families as described in Box 5.2.
apoptotic death.
ER is now regarded as a sensor of perturbations of cell homeosta-
6,7
sis. Activation of the UPR and defects in UPR are known to be Retention of Resident Golgi Proteins
important factors that contribute to many disease processes ranging
from metabolic disease, neurologic disease, infectious disease, and Extensive analysis has failed to reveal a clear retention motif enabling
cancer (reviews on cancer and on UPR and diseases under the section subdomain-specific retention of resident Golgi proteins. Two possible
Suggested Readings). models have been proposed. One model is retention by preferential
interaction with membranes of optimal thickness. It is based on the
finding that the transmembrane domains of Golgi proteins are shorter
Control of Exit From the Endoplasmic Reticulum than transmembrane domains of plasma membrane proteins. These
differences should allow a preferential interaction with the Golgi
On achieving transport competence, proteins are granted access to membrane lipid bilayer that is thinner than that of plasma membrane.
higher-ordered membrane domains termed ER exit sites. At ER exit The other model is kin-recognition/oligomerization. It postulates that
sites, membrane-bound and soluble proteins are concentrated into proteins of a given subdomain of the Golgi membrane can aggregate
transport vesicles for trafficking to a network of smooth membranes
called the ER-Golgi Intermediate Compartment (ERGIC, see Fig. 5.4).
COPII complex, composed of coat proteins, concentrate and package
the protein cargo into vesicles. COPII binds to cargo molecules either BOX 5.2 Human Glycosylation Disorders
directly, if they span the membrane, or through intermediate cargo In humans, the three main glycosylation pathways are the N- and
receptors and then provides some of the force that causes vesicle O-glycosylation and the glycosylphosphatidylinositol (GPI) anchoring.
budding, thereby linking cargo acquisition to vesiculation. About 2% of the human genome encodes glycosylation reactions.
ER resident proteins are selectively sequestered in the ER both for Moreover, glycosylation pathways intersect with glucose, lipid, and
the absence of export signals and to the presence of ER retention isoprenoid metabolism, expanding the number of players involved in
signals. Soluble luminal ER resident are retained through a C-terminal these key protein modifications. Nearly 70 inherited glycosylation dis-
ER tetrapeptide retention motif KDEL. Frequently, transmembrane orders have been identified so far and this number is steadily increasing
proteins have either a C-terminal dilysine motif KKXX or an because of the progress in the technology of DNA sequencing and in
8
N-terminal diarginine motif XXRR, or variants thereof for trans- mapping mutations. The characterized mutations combined to the
biochemical lesion and to the clinical manifestations are classified in
membrane proteins. However, it is more accurate to indicate ER the CDG (congenital disorders of glycosylation) database. Mutations
localization signals as “retrieval motifs” because proteins bearing these affect almost every organ and some proved to block embryo develop-
signals can transiently escape from the ER into the ERGIC, from ment in animal models of disease. Abnormalities in N-glycosylation
which they are returned to the ER through the retrograde vesicular cause severe myasthenic syndromes caused by hypoglycosylation of
transport (see Fig. 5.4). the acetylcholine receptor that affects the signal transmission at the
For the KDEL motif of luminal ER proteins, a specific retrieval neuromuscular plaque. Other cause neurologic disorders. Complica-
receptor has been identified, first in yeast and then in mammals. The tions also arise from secondary effects caused by ER stress consequent
KKXX motif has been shown to interact directly with the COPI coat to poor glycosylation. O-Glycosylation defects are associated mainly to
protein complex that is involved in retrograde transport from the ER severe muscular dystrophy (Walker-Warburg syndrome) whereas lack
of the first step of GPI synthesis provokes paroxysmal nocturnal
to the Golgi. Retrograde transport also serves to replenish the vesicle hemoglobinuria, a well-known hematologic disorder that results in
components lost as a result of anterograde (forward) transport. In erythrocyte lysis.
conclusion, selective protein exit from the ER is achieved by