Ribosome structure and function

Function - protein synthesis

Structure -

• Two subunits (large and small), composed rRNA and multiple proteins 
• Subunits come together by binding an mRNA strand, which exhibits basophilic staining  
• Polysomes (polyribosomes)–groups of  ribosomes that are translating the same  strand of mRNA and can make many copies of the same protein

Structure and Function of rER?

Structure:

•  membrane-limited flattened sacs called cisternae  
•  rER may be continuous with nuclear envelope
•  mRNA assoc w/ ribosomes exhibits basophilic staining
•   “Ergastoplasm”–portion of the cytoplasm that is basophilic staining–in secretory cells, e.g. pancreas the ergastoplasm is the rough ER

Function:
synthesis of secreted and membrane proteins

Ribosomes produce proteins that are injected into the lumen of the cisternae where they are modified, concentrated, or carried to another part of the cell w/in the ER

Structure and Function of Smooth ER?

Structure of sER
•  Similar to rER, but lacks SRP receptors so exhibits eosinophilic staining 

•  Tubular rather than sheet-like
•  Can be separate from rER or continuous with it  

Functions of sER

1. Synthesis of steroids & lipids

2. Detoxifying  enzymes of harmful compounds
3. Sequestering and releasing Ca++ in a controlled manner, specialized form in muscle cells called sarcoplasmic reticulum

Structure and Functions of Golgi Apparatus

Structure of Golgi
•  A series of cisternae–stacked flattened membrane-lined sacs with tubular extensions embedded in a network of microtubules  
•  Associated with small vesicles  
•  Abundant in secretory cells, and in cells producing large amounts of membrane and membrane-associated proteins
•  Apparent in L.M. as non-staining area

Function of Golgi
•  Modification, sorting, packaging proteins
•  During passage through Golgi, proteins and lipids undergo remodeling of N-linked oligosaccharides, which were added in rER:

• Glycoproteins and glycolipids have oligosaccharides trimmed and translocated
• Enzymes add, remove or modify sugar moieties
• M-6-P is added to proteins destined for late endosomes or lysosomes
• Glycoproteins are phosphorylated and sulfated
• Proteolytic cleavage of some proteins

Strcture and Function of Lysosomes

Function: Vesicles where macromolecules are digested

Structure of Lysosomes

•Contain hydrolytic enzymes targeted to lysosome by M-6-P: proteases, nucleases, glycosidases, lipases, phospholipases

•Have a specialized membrane resistant to hydrolytic digestion occurring in the lumen

• specialized proteins are glycosylated on the luminal surface of the membrane, protecting the membrane from the hydrolytic enzymes
• proton pumps to acidify the interior (pH ~ 4.7)
• During digestion nutrient diffuse through the membrane into the cytoplasm, in some cases transport proteins move nutrients across the membrane

What is the difference between lysosome and proteosome?

•  Degrade individual protein molecules (as opposed to lysosomes that digest bulk material)
•  Composed of 7 proteins including proteases and an ATPase
•  Recognize and degrade proteins with ubiquitin attached. 
•  Proteins that are denatured or oxidized proteins are recognized and tagged for degradation by enzymes that conjugate ubiquitin to lysine residues within the protein.

Describe the contents and function of the peroxisomes and how these relate to physiological conditions that affect the number of peroxisomes in the cell. 

•  Spherical, single membrane-bound organelles that contain oxidative enzymes: (1) catalase (breaks down H₂O₂ which forms during oxidation reactions), (2) peroxidases
•  Oxidative enzymes such as D-amino acid oxidases, b-oxidation enzymes, are especially abundant in hepatocytes where they are involved in detoxification:

• Alcohol in converted to acetaldehyde
• β -oxidation of long chain fatty acids 

•  Proteins destined for peroxisomes are formed by cytoplasmic ribosomes and tagged with peroxisomal targeting signal attached to C terminus
•  Number of peroxisomes increases in response to diet, drugs and hormonal changes

How does the ER interact with ribosomes and how do those proteins (membrane and secreted) differ from the ones produced for the cytoplasm?

rER synthesize membrane and secretory proteins

•  The portion of the protein inside the cisterna is modified by enzymes:

•  Glycosylation
•  Disulfide and internal hydrogen bond formation
•  Folding
•  Partial subunit assembly

•  If not properly modified, they can't leave ER

Free ribosomes synthesize cytoplasmic proteins:

• hemoglobin – in RBCs 
• actin, myosin – in muscle cells
• neurofilaments – in nerve cells
• keratin – in keratinocytes
• mitochondrial enzymes

Compare and contrast the role of the signal sequence in the production of membrane and secreted proteins.

For both membrane and secretory proteins:

• Protein synthesis begin on ribosomes in the cytoplasm.
• The ER signal sequence becomes attached to a signal recognition particle (SRP) that binds a SRP receptor on the ER membrane targeting the ribosome to the ER.
• The protein then gets cotranslationally inserted into and passes through the membrane into the cisterna as it is translated 

secretory proteins will pass completely into the cisterna–signal sequence is cleaved from secreted proteins in the lumen of the ER  
membrane proteins will pass back and forth through the ER membrane–as new hydrophobic signal domain is added, which stops the threading process and permanently anchors the protein in the membrane at this site

Characterize the polarization of the Golgi and relate this to its function. 

•Polarized functionally and morphologically

cis-Golgi network (CGN)–area nearest rER is forming face 
trans-Golgi network (TGN)–area farthest from rER is maturing face     
medial Golgi–area in middle 

•Vesicles covered with coatemer (also called COPs) mediate bi-directional transport of proteins between rER and Golgi.

• retrograde transport (Golgi -> ER) –vesicles coated in COP-I mediate transfer of proteins
• antegrade transport (ER -> Golgi) –vesicles coated in COP-II mediate transfer

Compare and contrast 3 different pathways by which materials destined for degradation reach the lysosome with respect to size of particles and origin of particles (intracellular versus extracellular origin).

1. phagosome–forms as material is phaogcytosed and fuses with the lysosome to create a phagolysosome. (large extracellular particles) 
2. endosomes–complexes internalized follow the endocytic pathway from early to late endosomes and finally are delivered to the   lysosome for degradation.  (small extracellular particles)
3. autophagy–(intracellular particles both large and small)

Identify, describe, and distinguish between 3 types of autophagy.

1. macroautophagy–nonspecific

• A portion of cytoplasm or an organelle is surrounded by a membrane from ER to form an autophagosome
• The autophagosome fuses with a lysosome forming an autophagolysosome, in which degradation takes place
• A mechanism for recycling raw materials to maintain essential life process when nutrients for the cell are scarce (e.g. occurs in liver during early stages of starvation)   

2. microautophagy–nonspecific

• Degradation of cytoplasmic proteins in a slow continuous process under normal physiologic conditions
• Small cytoplasmic soluble proteins internalized into lysosomes by invagination

3. chaperone-mediated direct transport

• A special chaperone protein, hsc73, binds to specific proteins to be degraded, and transports them through the lysosomal membrane