Professor Alfred Cuschieri
Identify lysosomes in electron micrographs
State the characteristics and functions of lysosomes
Explain how lysosomes are formed
Name the properties of lysosomal membranes
Explain the mechanisms and importance of phagocytosis
Give examples of phagocytic cells
Explain the importance of receptor-mediated endocytosis
Describe how lysosomal enzyme deficiency cause serious disease
Identify peroxisomes and state their characteristic features
Explain the importance of peroxidase generation and degradation in lysosomes
Name some diseases caused by deficient functions of lysosomes
Distinguish between melanosomes and melanin synthesis
Distinguish between melanosomes and melanocytes
are membrane bound organelles
contain hydrolytic enzymes
have acidic contents (pH 4.5-5.5)
have electron-dense heterogeneous contents
digest ingested material and aged or damaged organelles
Lysosomal enzymes are hydrolases that catalyse the reaction: AB + H2O à AH + BOH
Over 60 lysosomal enzymes are known.
There is a hydrolyse for each type of biological molecule. These include:
Peptidases – hydrolyse proteins
RNAases – hydrolyse RNA
Lipases – hydrolyse lipids
Phosphatases – hydrolyse phosphates
Glucosidases – hydrolyse glycogen
Carboxylases - hydrolyse carboxyl groups
Sulphatases – hydrolyse sulphates
Esterases – hydrolyse esters
Acid phosphatase and b-glucuronidase have been used as histochemical markers for lysosomes.
1. an ATP driven proton [H+] pump to maintain a low pH (4.5-5.5) in the lysosomal compartment.
The proton pump:
Has a ‘lollipop’ structure similar to the F1 and Fo of mitochondria, and consisting of:
a head containing 6 polypeptide units
a stalk contains 5 polypeptide units
Is inhibited by n-ethylmaleimide
2. a glycoprotein coat, rich in carbohydrates, on its inner surface to protect it against hydrolysis by its own enzymes
3. transporter channels that transport break-down products such as amino acids, glucose, nucleotides and other small molecules out of the lysosome. These molecules may move out
a. by facilitated diffusion ;
b. by active transport;
c. by co-transport using the proton (H+) gradient to provide the energy for transport.
a. The hydrolytic enzymes are formed in the RER
b. Enzymes have a terminal mannose–6-phosphate group as a marker to be packaged in lysosomes
c. They are packaged at the trans face of the Gogi complex (termed the Trans Golgi Network -TGN)
d. The newly formed vesicles that bud off are termed early endosomes or primary lysosomes. They are small and have homogeneous, electron-dense contents.
e. They early endosomes (primary lysosomes) may fuse with phagocytic or other vesicle to form late endosomes or secondary lysosomes.
Functions of Lysosomes
Lysosomes may be involved in the following pathways:
3. Formation of endosomes
4. Receptor-mediated endocytosis
Phagocytosis is the process by which the cell engulfs particulate matter (>0.5 mm) from the extra cellular space, and digests it by lysosomal action. Three main cell types are capable of performing this function:
i. are widely distributed in the connective tissue around the body
ii. phagocytose and digest all particulate matter in the extracellular space, including micro-organisms, foreign particles and damaged cells.
iii. are derived from monocytes, circulating in the peripheral blood.
iv. may have different names in different tissues such as Kupffer cells in the liver, osteoclasts in bone, and microglia in the central nervous system.
b. Neutrophils in the peripheral blood, which phagocytose and digest microorganisms.
c. Eosinophils in the peripheral blood, which phagocytose and digest antigen-antibody complexes.
Macrophages have a gylcocalyx coat, rich in glycosaminoglycans, on their outer surface, which causes particulate matter to adhere to them. Neutrophils and eosinophils have specific receptors to recognise specific particles to be engulfed. (The specific molecules that bind to the receptors are called ligands).
a. Adhesion of the particle to the glycocalyx or specific receptor on the plasma membrane
b. Extrusion of pseudopodia to surround the particle. This is mediated by actin filaments.
c. Formation of a phagocytic vacuole containing the engulfed particle
d. Fusion with a primary lysosome
This is the process in which old or damaged organelles are broken down. It occurs in practically all cells as a recycling system. It is most marked in cells that are not replaced, such as neurons.
a. The organelle is surrounded by vesicles, which coalesce
b. The coalesced vesicles form a membrane surrounding the organelle. This is called an autophagic vacuole.
c. The autophagic vacuole fuses with one or more primary lysosomes to form secondary lysosomes.
d. Residual bodies are lysosomes with partially undigested material
Exocytosis is the reverse of pinocytosis, i.e. the extrusion of fluid contained in vesicles into the extra cellular space.
Endocytosis and exocytosis are important mostly for membrane flow. For example, exocytosis replaces the plasma membrane removed by phagocytosis. Similarly, exocytosis of secretion vesicles must be balanced by endocytosis.
Receptor-mediated endocytosis is the process whereby cells that have a specific receptor take up specific macromolecules.
This process is used for the uptake of hormones, growth factors, antibodies, lipoproteins etc
The receptors are integral membrane proteins
The molecule that binds to the receptor is termed the ligand.
The process of receptor-mediated endocytosis involves the following steps:
1 Binding of the ligand to the receptor
2 Lateral diffusion of the ligand-receptor complex
3 Accumulation of clathrin, adaptor protein and dynamin on the cytoplasmic surface of the plasma membrane at a particular site
4 Formation of a pit, and accumulation of the ligand receptor complex at the site of clathrin acculmulation
5 Deepening of the pit and formation of clathrin-coated vesicles containing the ligand-receptor complex
6 The vesicles lose their clathrin coat
7 The vesicles fuse with a primary lysosome (early endosome) and the ligand is cleaved from the receptor
Familial hypercholesterolaemia is a condition in which there is defective binding of low density lipoprotein (LDL) to its receptor. Receptor-mediated endocytosis of the LDL does not occur, and it accumulates as high levels in the blood.
The process of receptor-mediated endocytosis was first described through the study of LDL uptake by cultured fibroblasts.
Three molecules are involved in forming the coat on the cytoplasmic face of coated vesicles:
§ A molecule that has a triskelion (three-pronged) structure.
§ Under appropriate conditions it forms a hexagonal (geodesic) latticework on the cytoplasmic surface of the plasma membrane,
§ It causes the membrane to invaginate and form a coated pit and vesiclevesicle
2. Adaptor protein
§ Binds to the cytoplasmic end of the transmembrane receptor
§ Mediates the attachment of clathrin to the plasma membrane
§ Regulates clathrin assembly and disassembly
§ Is itself regulated by phosphorylation and dephosphorylation
§ Incorporates a GTPase that provides the energy for formation of the coated pit and vesicle
§ Undergoes a configuration change that brings about closure of the vesicle
Gangliosidosis is an example of a Lysosomal Storage Disease
Is caused by deficiency of the enzymes b-galactosidase.
Is associated with inability to break down the terminal galactose from GM1 ganglioside
Results in accumulation of GM1 ganglioside in lysosomes, which are consequently greatly enlarged.
Enlarged lysosomes accumulate in the cells of several tissues, the most important of which are:
Involvement of neurons of the central nervous system
-fits, psychomotor deterioration, mental retardation
Enlargement of liver & spleen
Widening of bones: deposition in marrow
Vacuolated lymphocytes deposition in lymphocytes
This disease is inherited as an autosomal recessive disorder.
There are several lysosomal storage disorders. All are associated with a deficiency of a particular lysosomal enzyme, resulting in accumulation of an undigested substrate within the lysosomes. The following are a few examples of lysosomal storage disease with the associated substrate:
Pompe à glycogen
Hunter disease à heparan & dermatan sulphates
Morquio’s disease à keratan& chondroitin sulphate
Tay Sachs disease à GM2 ganglioside
Niemann-Pick à sphingomyelin
Farberdisease à ceremide
Peroxisomes or Microbodies
The features of peroxisomes are:
Membrane-bound spherical organelles
Have a diameter of 0.2-2.0mm
Contain a granular matrix and an electron-dense crystalline core
Contain the enzymes catalase and urate oxidase
Have the ability to break down H2O2
Have a pH optimum of 7.5
RH2 + O2 à R + H2O2
- by conversion to water
2H2O2 à H2O
- by oxidation of another organic compound
H2O2 + AH2 à 2H2O2 + A
H2O2 is harmful to cells and has to be degraded almost as fast as it is produced.
1. b-Oxidation of fatty acids
very long are chain fatty acids are broken down to acetyl CoA with the production of H2O2
This reaction also occurs in mitochondria.
2. Transfer of NH3 (amino) groups from amino acids to ketoacids
- requires the enzyme aminotransferase
3. Oxidation of uric acid or urates to allantoin
requires the enzyme urate oxidase
All are associated with the production of H2O2
Peroxisomes also have synthetic functions including:
- synthesis of cholesterol and dolicholol (also occurs in SER)
- synthesis of plasmalogens (membrane components in brain & heart)
- synthesis of bile acids
Some diseases are caused by lack of specific peroxisome enzymes:
- a progressive neurological disorder
- a defect in b-oxidation of long chain fatty acid,
- results in accumulation of long chain fatty acids in neurons and other cells
Gout due to accumulation of uric acid
- failure of conversion of uric acid to allantoin
- causes deposition of uric acid in joints, and consequent painful arthritis
Zellweger syndrome caused by absence of peroxisomes
- causes severe neurological disorder, metabolic defects, and early death
Melanosomes are the organelles in which melanin synthesis occurs.
They are :
- Oval-shaped , membrane-bound bodies
- 0.3-1.3 mm in diameter
- found in melanocytes (pigment-producing cells)
- contain enzymes for the biosynthetic pathway of melanin
- are identified by the DOPA reaction
Biosynthetic pathway of melanin formation
A multi-step pathway in which tyrosine is DOPA by a series of reactions involving the enzyme tyrosinase as well other enzymes. DOPA is then converted to melanin.
In the DOPA reaction, cells are exposed to an excess of DOPA. Melanin pigment will be formed in the melanosomes, the organelles that have the necessary enzymes for the biosynthesis of melanin.
Melanosomes are formed from two sources:
From the SER – pre-melanosomes are formed by coalescence of vesicles, and appear to contain parallel filamentous contents.
In the RER and Golgi complex, tyrosinase and other enzymes are synthesised and packaged into vesicles. These fuse with the pre-melanosomes.
The plasma membrane of melanocyte has receptors for MSH. The MSH enters the cells by receptor-mediated endocytosis and the MSH-containing vesicles fuse with the melanosomes.
Melanocytes have numerous dendritic processes.
Melanin-containing vesicles are pinched off from the tips of these processes.
They are endocytosed by the keratinocytes, degraded by lysosomes and the melanin become dispersed within these cells.