• Introduction
• The Lysosome: Structure
• The Formation of Lysosomes
• Lysosomal Enzymes Digest Everything
• Functions of Lysosomes
• Why doesn't the lysosome digest itself?
• Lysosome & Digit Formation
• Lysosomes in the Disease Process
• Occupational Diseases: Silicosis
• Abnormalities of Mucopolysaccharide Metabolism
• Background to Tay Sachs: Glycosphingolipids
• Tay Sachs Disease

Based upon appearance alone, the lysosome is one of the simplest cellular organelles. Basically it is a bag of digestive enzymes. But looks can be deceiving. This group of lysosomal enzymes is capable of digesting essentially every type of biological molecule. For this reason, the lysosome was originally considered to be only involved in digesting materials that the cell ingested through phagocytosis or pinocytosis. With more research it became clear the lysosome has many more cellular responsibilities. This was dramatically emphasized when it was shown that the absence of a single lysosomal enzyme in humans can lead to serious abnormalities, dementia and death.

Lysosomal enzymes are synthesized on the rough endoplasmic reticulum (rer) and packaged into prelysosomal vesicles by the Golgi. The enzymes are glycosylated in the rough endoplasmic reticulum and a mannose group is phosphorylated in the Golgi to target them to lysosomal vesicles as discussed in a future lecture. In the classic view of lysosomal biogenesis, prelysosomal vesicles that bud directly from the Golgi fuse to form mature, primary lysosomes. Recently it has been shown there are different routes to forming lysosomes. For example, the formation of late endosomes by the fusion of vesicles (early endosomes) from the cell membrane with vesicles from the trans-Golgi network can lead to lysosome formation. It is clear our understanding of the details of endosomal events and lysosome biogenesis are still in their infancy (Mullins & Bonifacino, 2001. BioEssays 23: 333-343). The topics of endosome formation and lysosomal digestion are again addressed in our lecture on receptor-mediated endocytosis.

The following picture shows the diversity of lysosomes enzymes that are capable of digesting almost all biological molecules.

• The amounts and types of each lysosomal enzyme will vary with the cell type and with its physiological state. For example, the human sperm contains a unique protease called acrosin in a single, specialized lysosome called the acrosome. This and other acrosomal enzymes are used to penetrate the zona pellucida of the egg during fertilization.
• It is likely that different lysosomes contain varying amounts of the different lysosomal enzymes in any cell type.

The next diagram is a composite showing essentially all of the known functions of lysosomes. We've divided these up into 6 functional groups that are summarized below. These are not the only functions of lysosomes as new responsibilities for this organelle are being discovered including metal ion homeostasis and cell membrane repair.

1. Digestion of Ingested Materials - - Cells ingest materials by various endocytotic means including the classic phagocytosis ("cell eating") and pinocytosis ("cell drinking"). Inside the cell, the material that is taken up is enclosed in an endosome (phagosome or pinosome, respectively). Inside the cell the endosome fuses with a primary lysosome to form a digestive vacuole. In the digestive vacuole the hydrolases of the lysosome will act on the ingested material to break it down. After digestion is complete, the vacuole is called a residual vacuole because it is full of residual, indigestible components. The contents of the residual vacuole are released outside of the cell by exocytosis.
2. Cell Death - Lysosomes mediate events in the controlled or programmed death of cells called apoptosis. This is discussed below. They also come into play during necrosis, the pathologic death of cells and tissues. For example, meat becomes tenderized after the death of the animal because the lysosomes break down releasing their enzymes into the muscle causing the digestion of the contractile and other muscle proteins.
3. Autophagy - The survival of cells requires that cellular constituents are constantly turning over. New molecules and structures are made while old unnecessary or worn out components are removed. During starvation, cells use autophagy to break down cellular components to provide energy for their survival. In the case of organelles, the mitochondrion, for example, is separated from other cellular constituents by an isolation membrane to become an autophagosome. The autophagosome fuses with primary lysosomes to form an autophagic vacuole within which the mitochondrion is digested. The resulting residual vacuole is exocytosed. The following picture shows the sequence of events in digesting a mitochodria.
   
   It is still not known whether the double-membrane isolation membrane that forms the autophagic vacuole is formed de novo or from existing membrane structures such as the endoplasmic reticulum (for more see: Juhasz & Neufeld, 2006. Autophagy: A 40 year search for a missing membrane source. PLoS Biology 4: 0161-0164.)
4. Molecular Turnover - In this situation, molecules are digested by lysosomal enzymes. The exact ways in which the different types of molecular turnover occur are under active investigation. But this process removes old, abnormal or unnecessary molecules allowing cells to alter their physiology or behaviour. Some of the molecules enter the digestive pathway via receptor mediated endocytosis as mentioned below and discussed in detail in a future lecture.
5. Extracellular Functions - Lysosomal enzymes have responsibilities that lie outside of the cell as well. For example they can digest extracellular components or modify the cell surface. For example, high levels of secretion of glycosidases are linked to some of the changes in cell adhesion molecules that underlie the behaviour of some cancer cells.
6. Receptor-Mediated Endocytosis - Lysosomes play an important role in the uptake and modification of critical molecules such as cholesterol. They also mediate events of receptor recycling and the shutting down of events of cell communication. This sequence of events involves a receptor binding to its ligand followed by their uptake into coated vesicles. The coated vesicle is uncoated prior to fusing with lysosomes. The events that unfold vary with the type of endocytotic event and are detailed in the lecture on receptor mediated endocytosis.

Since the lysosome is full of digestive enzymes that can digest essentially all cellular components, why doesn't the lysosome digest itself? This is because the inner leaflet of the lysosomal membrane is coated with an extensive glycocalyx (like that present in the intestinal epithelium to prevent its digestion; see lecture on "The Cell Membrane"). The integral and peripheral membrane proteins on the inner surface are highly N-glycosylated glycoproteins containing poly-lactosamine which prevents access by the digestive enzymes. For a review on how lyosomes digest other membranes but not their own see: Kolter & Sandhoff, 2005. Principles of membrane digestion...Ann. Rev. Cell Dev. Biol. 21: 81-103. The figure below shows the expression of GFP-LC3 (LC3 is a marker protein for autophagy) in mouse heart muscle cells revealing the presence of autophagosomes.

Apoptosis is dealt with in more detail in several other courses including "Human Development, BIO380". Here's the URL for the lecture in which apoptosis is discussed: http://www.utm.utoronto.ca/~w3bio380/lecture3.htm

• Cell death is an important phenomenon
• Normal event in development of brain (neurons), ovaries (ova), hands
• Mediated by specific signalling pathways that lead to activation of "Death Genes"
• Specific characteristics are evident: blebbing of membrane, laddering of DNA, etc.

The controlled killing of cells is an essential process that occurs during embryonic development as well as during our day to day lives. One classic example of programmed cell death is digit formation.

• Lysosomes "kill" cells between digits
• Process of controlled cell death is called "Apoptosis"
• Ducks have no lysosomal activity between digits of toes = webbed feet; chickens have activity = no web
• Lack of apoptosis in humans can lead to webbed fingers called syndactyly as shown in the next figure

• Silicosis & Asbestosis
• Cancer (secreted lysosomal enzymes alter cell surface)
• Abnormalities of Mucopolysaccharide Metabolism
• Rheumatoid Arthritis (secretion of high levels of lysosomal enzymes digest extracellular matrix)

• Material made of Silica: Rose quartz, glass, digital watches, onyx, porcelain, beach sand, agate
• Inhaled silica (silicon dioxide) dust enters lungs
• Macrophage ingest & dust enter 2o lysosomes
• Can't be digested
• Lysis & release of enzymes
• Sets up inflammatory response in lung tissue
• Can lead to Tuberculosis and failure of respiratory system

• Over 2 dozen types
• Genetic Defect = Absence of 1 Enzyme (e.g., alpha-fucosidase, alpha-mannosidase, etc.)
• Tay Sachs Disease, Hurler's Syndrome, Gargoylism, etc.
• Often called Glycosphingolipid (GSL) lysosomal storage diseases: because they involve problems with digestion of GSLs
• Detection: Amniocentesis & enzyme analysis or genetic screening
• Possible Medical Intervention: Genetic Engineering; Pharmacological; Enzyme replacement therapy

Glycosphingolipids are commonly found at the surfaces of eukaryotic cells. They are comprised of a ceramide moiety that inserts in the cell membrane plus an oligosaccharide chain. As can be seen in the picture below the ceramide portion consists of fatty acid chains like the phospholipids discussed in the lecture on cell membrane structure. Attached to these are sugar moieties (sialic acid residues) that orient to the outside of the cell. When the cell membrane components are recycled, normal digestion occurs by the stepwise removal of monosaccharides producing fatty acid chains that can be released from the lysosome.

• Occurrence in Jewish People of Ashkenazic (Central European) Descent
• Due to missing Hexosaminidase A (also called N-Acetylglucosaminidase A)-an enzyme that removes acetylglucosamine residues from polysaccharides
• Neurologic Disease: Build up in secondary lysosomes constrict nerve axons

• Leads to blindness, dementia & paralysis
• Evident by 6 mo.; Death 2-5 years of age
• Quebec: Lineage with similar disease; different gene defect

Let's look in a bit more detail about how the loss of the single Hexosaminidase A enzyme can have such a devastating effect. In normal cells the turnover of ganglioside GM2 occurs regularly. Once inside digestive vacuoles the normal complement of enzymes breaks it down and the contents of the residual vacuole are exocytosed. In Tay Sachs, the absence of Hexosaminidase A prevents complete digestion of the ganglioside GM2 because acetylglucosamine residues cannot be cleaved off. This results in the inability of the residual vacuoles to be exocytosed. Thus they continue to accumulate in the cytoplasm of the cell causing it to swell up. We'll look at this issue in more detail when we discuss the topic of protein targetting in cells.