In this article we will discuss about:- 1. Meaning of Endoplasmic Reticulum 2. Functions of Endoplasmic Reticulum.
Meaning of Endoplasmic Reticulum:
It is a double membrane system found throughout the eukaryotic cell. The membrane is continuous with the nuclear membrane and the plasma membrane. The endoplasmic reticulum (ER) is absent in the prokaryotic cell.
George Palade (1974) made electron microscope studies of the subcellular fractions in demonstrating endoplasmic reticulum and Golgi complex. In course of staining the cell with basic dyes, it has been observed that certain regions of the cytoplasm show intense staining. These regions were termed ergastoplasm in the late nineteenth century.
With the discovery, of electron microscope in 1940, K. ‘Porter, A. Claude and E. Fullam (1945) showed through electron microscope without making any sections that cells contain a lacelike network of strands throughout the cytoplasm.
Later, through fine sectioning, it has been established that the largest membranous network in eukaryotic cell is the Endoplasmic Reticulum (ER) consisting of tubules, vesicles and sacs. The endoplasmic reticulum is found in two forms—granular of Rough Endoplasmic Reticulum (RER.) and an ‘agranular’ or Smooth endoplasmic reticulum. (SER).
Ribosomes are attached with the Rough endoplasmic reticulum which causes certain regions of the cytoplasm to stain with basic dyes. Smooth endoplasmic reticulum lacks ribosomes. The cytoplasm of the cell was divided into two compartments, one the cell sap and the other cisternal space surrounded by the endoplasmic reticulum.
There is another space near the nucleus, termed the perinuclear space by the endoplasmic reticulum. There is a close relationship between the perinuclear space, cisternal space and the cell sap (Fig. 3.1). Soluble enzymes, transfer RNAs, free ribosomes and other factors for protein synthesis are present in the cell sap.
The smooth endoplasmic reticulum (SER) is the region where the synthesis and metabolism of fatty acids and phospholipids occur. Different cells may contain different amount of SER depending on their nature and function.
Enzymes that modify toxic chemicals or carcinogens to reduce its toxicity are present in SER. These toxic chemicals are secreted out of the body through the formation of vesicles by SER. Under the electron microscope, the SER consists of an interconnected series of convoluted tubules.
In addition to the presence of ribosomes in the rough endoplasmic reticulum (RER) it appears as large flattened sheets of membrane showing lumen or central cavity under the electron microscope after thin sectioning of the cell.
RER is present in cells in high frequency which participate in the synthesis of protein for export. The biochemical studies to note the functions of the endoplasmic reticulum are based on investigations on subcellular fractions isolated through centrifugation.
During the process of isolation, the endoplasmic reticulum is broken into smaller membrane fragments which sometimes reseal into vesicles called microsomes. Detailed biochemical studies on these membrane vesicles, or microsomes showed the presence of various enzymatic activities.
The main functions of the endoplasmic reticulum can be summarised as:
i. The synthesis and transport of protein molecules,
ii. The metabolism of carbohydrates and lipids and
iii. De-toxification and secreting out of drugs and toxic chemicals from the cell.
Functions of Endoplasmic Reticulum:
(i) Smooth Endoplasmic Reticulum (SER):
(a) Carbohydrate and Lipid Metabolism:
The enzyme Glucose-6-Phosphatase is present in the endoplasmic reticulum particularly in SER. It helps in the hydrolysis of Glucose- 6-Phosphate to glucose and phosphate. This enzyme has a key role in the metabolism of Glucose-6-Phosphate releasing from the glycogen granule near the SER.
Glycogen, the reserve food of many cells, is converted to Glucose-6-Phosphate through the formation of Glucose-1-Phosphate in the first step. The Glucose-6-Phosphate is then brought to the smooth endoplasmic reticulum (SER) where it is broken down to Glucose and Phosphate by the enzyme present in the SER.
The Glucose thus formed is then transported first into the cisternal space and finally to the extracellular space or to the blood stream for performing other cellular activities.
There are other enzymes of carbohydrate metabolism, such as Amylase and glucuronidase for the hydrolysis of starch and mucopolysaccharides. Other enzymes present in the membranes also help in converting lipids and other hydrophobic compounds to water soluble form for easy transport through the membrane.
Large number of fat droplets is found in the cisternal space of the endoplasmic reticulum. The enzymes of lipid metabolism present in the smooth endoplasmic reticulum are responsible for the synthesis of cholesterol and other steroid hormones in the endoplasmic reticulum.
As for example, smooth endoplasmic reticulum (SER) is well developed in the adrenal cortex and interstitial tissues of the testis because these tissues produce steroid hormones.
(b) Detoxification of Toxic Chemicals:
The main principle of detoxification of chemicals is by converting hydrophobic substances into water soluble ones. Most of the drugs, pesticides, toxins and other chemical pollutants are generally hydrophobic compounds, so they try to accumulate in body fats.
The reactions involved in this detoxification include hydrolysis, reduction or conjugation. G. Mueller and J. Miller first reported the presence of this type of enzyme system in liver cells.
They confirmed their view with an experiment in the laboratory showing oxidization of carcinogenic chemicals by liver homogenates in presence of NADPH. These oxidase enzyme systems for the detoxification of drugs and other chemicals, consist of an important element, termed cytochrome P 450.
This cytochrome P 450 has an important role in the detoxification of drugs. Cytochrome P450 is an iron containing compound having a special property of reducing its form when it binds with the substrate.
This reduced form again can absorb light at 450 nm. When any compound, say drug, is attached with the cytochrome P450, the iron (Ferric, Fe+++) present is reduced to Ferrous (Fe++) with the help of NADPH dependent cytochrome P450 reductase.
Now the reduced cytochrome P 450 will bind oxygen. One atom of oxygen is used to oxidize the substrate to convert the (Fe++) to (Fe+++) and the other atom of oxygen is used to oxidize the drug with the liberation of one molecule of water.
This enzyme system is also employed in the oxidation of steroids and fatty acids. The constant habitual user of drugs, for example, use of sedative phenobarbital, increases the synthesis of the enzyme oxidase present in the liver and SER resulting into the decrease in the effectiveness of other drugs like antibiotics, steroids, anticoagulants, narcotics etc.
In addition to cytochrome P 450, cytochrome P 448 and cytochrome b5 are also present in the endoplasmic reticulum. Cytochrome b5 has been employed for the desaturation of fatty acids. Cytochrome P448 tries to metabolize polycyclic hydrocarbons.
(ii) Rough Endoplasmic Reticulum (RER):
Transport of Protein Molecules:
Rough endoplasmic reticulum, i.e., portion of ER bound with Ribosomes takes part in the synthesis of certain specific proteins. These proteins are glycoproteins, lysosome proteins, membrane proteins and some organelle proteins.
Ribosomes may remain in the cell either in a bound form to ER or as free in the cytoplasm. This condition of ribosome depends on the physiological condition of the cell as well as on the cell type.
The meristematic cells or growing cells contain large number of free ribosomes and cells synthesizing membrane proteins or secreting proteins containing large amount of bound ribosomes. In other words, these cells have more Rough endoplasmic reticulum than Smooth endoplasmic reticulum.
These proteins, synthesised on RER, transport into the cisternae or space of the endoplasmic reticulum for exportation. The proteins synthesised on free ribosomes are utilised within the cell. For example, the protein (Ferritin) synthesised on the free cytoplasmic ribosomes of the liver cell, is retained within the liver.
The other proteins, albumin and gamma globulin, are synthesised on the membrane-bound ribosomes. These are then secreted into the blood. Hence there must be some sort of signal which dictates m RNA to synthesise protein either on free ribosomes or on membrane-bound ribosomes.
Again, ribosomes do not always remain attached with ER, only when membrane protein or secretory protein is needed by the cell, ribosomes will bound to the ER and the specific signal makes the mRNA to function.
(iii) Signal Sequences and Protein Processing in Endoplasmic Reticulum:
The association of mRNA ribosome complexes to the endoplasmic reticulum is done by the ‘signal’ sequences located near the N terminal end of the protein whose synthesis is to be directed. This is known as Signal hypothesis (Blobel and Dobberstein, 1975).
The mRNA ribosome complex having N terminal’ signal sequence with a short polypeptide binds with the endoplasmic reticulum with the help of a specific protein known as the Signal recognition particle (SRP).
This ‘SRP’ actually identifies the ribosome containing N terminal signal sequence from the pool of ribosomes present in the cytoplasm and then binds the proper ribosome—mRNA complex—to the endoplasmic reticulum. This SRP helps in the synthesis of secretary membrane and lysosomal proteins to the endoplasmic reticulum.
Now the ribosome—mRNA SRP complex—binds to the endoplasmic reticulum through further interaction between the SRP with the second receptor present in the ER, termed signal recognition particle receptor (SRPR) or signal sequence receptor (SSR) or ‘docking protein’. This ‘docking protein’ actually captures the ribosome portion of the complex (Savitz and Meyer, 1990).
Translocation of the protein starts and SRP helps in the insertion of the newly synthesised polypeptide into the ER lumen or cisternae probably through a proteinaceous aqueous channel. The structure of this channel or pore is not known.
SRP then dissociates from the complex to perform another cycle of targeting. The dissociation of SRP is catalysed by an enzyme, termed Signal peptidase, which is present on the inner surface of the endoplasmic reticulum facing the lumen (Fig. 3.2).
SRP consists of a single RNA molecule known as 7 SL RNA or SRP 7S which contains RNA and six different polypeptides. These polypeptides have two monomers, a 19KDa polypeptide, and a 54 K Da polypeptide, and two heterodimers, one composed of a 19 K Da and a 14 K Da polypeptide and the other comprised of a 68 K Da and a 72 K Da polypeptide.
A number of seed proteins and hydrolytic enzymes enter the endoplasmic reticulum after the synthesis. These proteins are then secreted either in the vacuoles or stored in other bodies. During transport of proteins, these are modified by the enzymes present in the endoplasmic reticulum.
The modifications of the protein was done by hydroxylation of proline and lysine residues, partial proteolysis of proteins and by glycosylation of specific amino acid residues. Most of the secretory proteins are glycoproteins—pepsin, ribonuclease, plasma proteins like immunoglobins, hormones (gonadotropins), mucous secretions etc.
Another important modification taking place in the lumen of ER is the attachment of carbohydrate to polypeptide chains by the enzyme glycosyltransferases. The glycosylation reaction involves certain steps. In the first step, a carrier of oligosaccharide, dolichol phosphate, remains attached in the lumen side of endoplasmic reticulum.
The carbohydrates like mannose, glucose and N acetyl glucosamine are attached with the dolichol phosphate to form a core oligosaccharide chain in presence of an enzyme Glycosyltransferase. This same enzyme, again, performs another reaction to attach this core oligosaccharide chain to the asparagine residue of the growing polypeptide chain in the lumen of the ER which is still hanging from the end of the ribosome.
When the protein synthesis is complete, the final protein product is released into the lumen of the ER which is finally modified in the Golgi complex. The binding of the ribosome to the ER is stabilized by an interaction between ribosome and ribophorins, the integral membrane proteins.
Actual mechanism of the signal sequences and protein processing in the ER has been studied in case of certain seed storage proteins such as Zeins of Maize and the Vicilins and legumins of Pea which accumulate in membrane-bound vesicles. It has been possible to know the mechanism in detail with the development of the experiment in cell free system.
The three maize proteins synthesised from cDNA sequences have relatively short signal sequences of 18-21 amino acids. The general characteristics of the signal sequence Eire almost similar for most proteins that transport through endoplasmic reticulum.
Most of the proteins are glycosylated before transportation. But there are some exceptions such as legumin and concavalin A which are not glycosylated. An unusual rearrangement of amino acids occur in the lectin, concavalin A (Con A), which consists of four sub-units of 237 amino acids.