A variety of proteins synthesized by rough endoplas­mic reticulum pass through and is processed by the Golgi apparatus on their way to their final cellular destinations.

Best understood among these are pro­teins destined to be secreted from the cell, plasma membrane proteins, and lysosomal proteins.

Secretion:

Although the Golgi apparatus is involved in many dif­ferent cellular processes, its principal role in many cells is in secretion. Two sets of experiments bear on the role of the Golgi apparatus in secretion. In 1964, L. Caro and G. Palade showed that Golgi bodies in the acinar cells of pancreas tissue are involved in the packaging of enzyme precursors into zymogen gran­ules prior to secretion.

Caro and Palade injected ra­dioactive amino acids into rats and followed the move­ments of the “label” using autoradiography. This type of experiment is called a “pulse-chase” because the initial short-term application of labeled amino acids (i.e., the “pulse”) is immediately followed by the more prolonged application of unlabeled forms (i.e., the “chase”).

Although amino acid metabolism and protein synthesis are not interrupted, the meta­bolic fate of the labeled amino acids can be traced through the cell with time. As might be expected, af­ter a 3-minute pulse the label appeared almost exclu­sively in the rough endoplasmic reticulum, as this is the region of protein synthesis. Following the 3-minute pulse, nonlabeled amino acids were added for 17 minutes (e.g., a total of 20 minutes from the begin­ning of the pulse). Although some of the radioactive la­bel remained in the rough endoplasmic reticulum, most of the label had shifted to the Golgi apparatus.

When the chase was continued for an additional 100 minutes (120 totals), almost the entire label had left the endoplasmic reticulum and the Golgi apparatus and was now found in the zymogen granules and in the extra­cellular space (as a result of the discharge of the con­tents of the vesicles at the plasma membrane). These experiments showed that the path of the amino acids is first into proteins in the rough ER and that these proteins are then transferred into the cisternae of the Golgi apparatus and then into the zymogen granules.

Two models have dominated thought concerning the manner in which proteins pass from the cis face of the Golgi apparatus to the trans face. According to the cisternal progression model, new cisternae are con­tinuously formed as vesicles containing proteins to be processed by the Golgi are dispatched from the endo­plasmic reticulum and coalesce at the cis face. Each cisterna then progresses through the stack toward the trans face. On reaching the trans face, the cisternae are broken down to form small vesicles that carry their entrained proteins to various cellular destina­tions.

The weight of recent scientific evidence argues against the cisternal progression model. For example, it is now known that the different cisternae of the Golgi possess different biochemical properties (e.g., different enzymes; see below), and it is difficult to ex­plain how a cisterna having one set of properties could change into another. Consequently, the cisternal pro­gression model is giving way to the cisternal transfer model, according to which the proteins proceed from one cisterna to the next via small vesicles that are re­leased from one cisterna, move forward, and then fuse with the next cisterna.

Posttranslational Processing of Proteins:

In 1966 using similar autoradiographic techniques to those employed by Caro and Palade, M. Neutra and C. P. Leblond studied the secretion of mucus by the goblet cells of intestinal epithelium. Mucus is a glyco­protein in which glucose and glucose derivatives are linked together to form polysaccharide side chains of the protein molecules (Fig. 18-10).

Mucous Glycoprotein  Glucose labeled with tritium (3H) was used to follow the assembly and fate of the glycoproteins. Fifteen minutes after injec­tion of the radioactive sugar, the label was most con­centrated in the cisternae of the Golgi apparatus. This label did not enter or associate with the rough endo­plasmic reticulum first.

After a 20-minute chase, the label appeared in the mucous vesicles, and after 4 hours, most had been released through the plasma membrane into the intestinal lumen. This experiment not only revealed the path of glucose through the cell but also demonstrated that the final stages of assem­bly of the glycoprotein occur inside the Golgi appa­ratus.

Using the goblet cell as an example, Figure 18- 11 depicts the central role of the Golgi apparatus in the packaging of newly synthesized proteins into vesi­cles for secretion. The assembly and processing of large molecules in the Golgi apparatus is not unique to goblet cells. Cartilage cells assemble glycoproteins in the cisternae of their Golgi bodies and sulfate groups have been shown to be added as well.

Recent studies indicate that the enzymes employed in the processing reactions are selectively localized in the sequential cis­ternae of the Golgi apparatus in the order of their use. In plant cells, pectins and cellulose are assembled in the Golgi bodies prior to deposition onto the forming cell plate or cell wall.

J. E. Rothman, A. Kornfeld, H. Schachter, and oth­ers have shown that glycosylation of proteins begins in the rough endoplasmic reticulum and continues in the Golgi cisternae (Figs. 18-12 and 18-13). Prelimi­nary glycosylation in the endoplasmic reticulum is called core glycosylation.

Incorporation of Amino Acids into Secretory Proteins by the Goblet Cells of the Intestine

In the endoplasmic reticu­lum, a specific branched oligosaccharide is added to each glycoprotein being synthesized. The branched oligosaccharide consists of 14 sugar units: 2 molecules of N-acetylglucosamine (Nag), 9 molecules of mannose (man), and 3 molecules of glucose (glc) (Fig. 18-13).

Functions of the Golgi Apparatus

The oligosaccharide is covalently linked to the R- groups of asparagine (asn) residues in the proteins. Shortly after addition of the oligosaccharide, and while the protein is still in the endoplasmic reticulum, one of the mannose units and all three glucose units are removed (Fig. 18-13). Following this, the proteins are transferred to the cis face of the Golgi apparatus.

Division of Labour among the cis, Medal and Trans Cisternae of the Golgi Apparatus and the Endoplasmic Reticulum

All of the glycoproteins arriving at the cis face of the Golgi apparatus from the endoplasmic reticulum have the same oligosaccharide chains. Before passing from one Golgi cisterna to the next, the proteins un­dergo chemical modification (i.e., processing). For ex­ample, in the cis cisterna, phosphate groups are added to the ends of the oligosaccharide chains of proteins destined for lysosomes, Secretory proteins and pro­teins destined for the plasma membrane experience more extensive processing.

Five of the mannose units of their oligosaccharide chains are removed in the cis cisterna and are replaced in the medial cisternae by two molecules of N-acetylglucosamine (Fig. 18-13). After transfer to the trans cisterna, disaccharides consisting of galactose and AT-acetylneuraminic acid (also known as sialic acid) are added to the ends of each branch of the oligosaccharide.

Proliferation of Cellular Membranes:

In addition to its role in secretion, the Golgi apparatus plays a role in the preparation of membranous ele­ments for organelles such as lysosomes and the plasma membrane (Fig. 18-12). Proteins destined to be incorporated into lysosomes or the plasma membranes are synthesized by ribosomes attached to the endoplasmic reticulum (i.e., rough ER).

Some of these proteins are released into the intracisternal phase of the ER and others remain anchored in the ER mem­branes, within minutes of their synthesis, these pro­teins appear in the cis face of the Golgi apparatus. The mechanism by which intracisternal phase and membrane-associated ER proteins reach the Golgi ap­paratus is uncertain but is generally believed to in­volve one or a combination of the following processes.

Transfer to the Golgi apparatus may be mediated by dispatchment of small vesicles from the ER that mi­grate to and fuse with the cisterna that comprises the cis face. Newly synthesized proteins discharged into the intracisternal phase of the ER would be enclosed within these vesicles, and proteins that were left an­chored in the ER membrane would be constituents of the vesicle wall. Alternatively, proteins in the intracis­ternal phase of the ER reach the cisterna of the cis face by diffusion along transient connections between the channels, whereas ER membrane-anchored pro­teins reach the Golgi apparatus by lateral flow within the membranes that form these connections.

As glycoprotein processing progresses, the proteins are successively transferred from one Golgi cisterna to the next, ultimately reaching the trans face. The transfer may take the form of small intermediary vesi­cles, or diffusion (or lateral membrane flow) through transient continuities between adjacent cisternae.

Proteins that are destined to be components of lyso­somal membranes or the plasma membrane and that are anchored in the ER membranes at the time of syn­thesis are presumed to move from the ER to the cis face and from the cis face through the medial cister­nae to the trans face as membrane components. The membranes of the vesicles discharged from the trans face contain these proteins (see Fig. 18-12). Soluble lysosomal proteins (as well as proteins destined for se­cretion) move from the lumenal phase of the ER through the cisternal space and are enclosed by the membranes of discharged vesicles.

Vesicles containing membrane-bound and soluble ly­sosomal enzymes are called primary lysosomes.  Vesicles containing secre­tory proteins fuse with the plasma membrane and empty their contents outside the cell. Vesicle membranes that are studded with pre­sumptive plasma membrane proteins also fuse with the plasma membrane.

As seen in Figure 18-12, those portions of the protein that faced the lumenal phase of the ER and that were glycosylated (or sulfated, etc.) in the Golgi cisternae face the exterior of the cell once they are incorporated into the plasma membrane. One of the characteristic fea­tures of many plant tissues is the presence of one or more large vacuoles. Some of these vacuoles have been shown to contain hydrolytic enzymes comparable to those present in lysosomes. For this reason, it has been suggested that the Golgi bodies of plant cells may give rise to some, if not all, of these vacuoles.

Sorting Problems Faced by the Golgi Apparatus:

It is clear from the preceding discussion that proteins destined for secretion granules, lysosomes, and the plasma membrane enter the cis face of a Golgi body from a common source—the rough endoplasmic retic­ulum. These proteins are delivered to the cis face of the Golgi along with a vast excess of ER membrane proteins, yet the vesicles dispatched from the trans face are essentially free of ER proteins. Thus, the Golgi apparatus also acts to progressively sort these proteins. ER proteins are believed to be returned to the endoplasmic reticulum by some of the small vesi­cles released from the margins of the cis cisternae (Figs. 18-12 and 18-13).

As noted above, there ap­pears to be an enzymatic compartmentation of the Golgi cisternae in that certain enzymes are selectively localized in specific cisternae of the stack. Although one might have expected that processed proteins with different destinations would exit the stack at different levels, this appears not to be the case. Instead, irre­spective of their final destination, secretory and lyso­somal proteins pass together through the stack of cis­ternae and are not sorted until they reach the trans face. In 1981, J. E. Rothman suggested that a Golgi body may actually be two organelles in tandem: the cis Golgi and the trans Golgi.

Beginning at the form­ing face, the cis Golgi consists of all but the last one or two cisternae and its role is to sort out ER proteins so that they may be returned to the ER and to process proteins having other destinations. The trans Golgi consists of the last one or two cisternae of the matur­ing face and it acts to receive the refined proteins and distribute them through vesicles to their specific loca­tions throughout the cell.

More recently, Rothman and others have moderated this view and now suggest that the Golgi apparatus consists instead of three compartments: the cis com­partment, the medial compartment, and the trans compartment. The cis compartment sorts out and dis­patches the ER proteins and also adds phosphate to the terminal sugars of lysosomal proteins. The medial compartment (which consists of cisternae at the cen­ter of the stack) is where N-acetylglucosamine is added.

The addition of the terminal galactose and N- acetylneuraminic acid units occurs in the trans com­partment and this is also where the various proteins are sorted according to final destination. The addition of phosphate to the terminal sugars of lysosomal pro­teins in the cis compartment apparently serves to preclude the addition of N-acetylglucosamine in the medial compartment and galactose and N-acetylneuraminic acid in the trans compartment.

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