BIO 421 Exam 2: lysosome and autophagy

• hydrolytic enzymes​

• acid hydrolases

• ~5.0

• proton pumps​

Intro to the lysosome​:

• A membrane-enclosed compartment filled with _____

• ~40 types of _____ (proteases,nucleases, lipases, etc)​

• Used to digest a variety of macromolecules​

​

• Lysosomal hydrolases require an acid environment to function​

• this protects the cell in case the lysosome should leak or be damaged​

• lysosome maintains pH of ____ in interior​

• pH is maintained via the action of ____

Large scale trash dispsosal of the cell ​

Only have that enzymatic activity at a low pH ​

A neutral pH allows the lysosome to not digest anything... so harmless when they are released at a neural pH (pH of cytosol is 7.2) ​

Maintains a pH of about 5 internally via proton pumps ​

Requires ATP to keep the pH low ​

• shape and size​

• breakdown

• digestion

• recycling

• acid hydrolases​

• lysosome-specific

Lysosomes are heterogeneous

• Lysosomes vary widely in _____

This reflects the many different functions they perform:​

• ____ of extra/intra cellular debris,including defunct organelles​

• ___ of phagocytosed pathogens​

• production of nutrients via ___​

• Can be identified as a lysosome via:​

• presence of _____

• _____ proteins​

Come in many different forms: heterogeneous ​

Not a typical lysosome size... so hard to identify ​

Since they are not easily distinguishable... they are idenfitied through the prescence of acid hydrolases and lyososome specific proteins ​

• ER

• golgi

Transport of acid hydrolases and membrane proteins to the lysosome​

• lysosomal proteins such as hydrolases and membrane proteins are synthesized in the ___​

• They are then transported to the ___ for further modifications​

​

Syntheised in the ER co-translationally​

Goes to golgi for further modifications for vesicle transport ​

Answer: COPII coat ​

• Expect to see a COPII vesicle budding off the ER and then fusing with the golgi ​
  

What type of coat protein is used in the vesicles that the lysosomal proteins are packaged into for transport from the ER to the golgi? (clathrin, COPI coat, or COPII coat) ​

COPII

COPI

Clatherin

In vesicle transport, ___ is typical for ER to Golgi ​

____ is from golgi to ER ​

___ buds off golgi to go to secondary destinations ​

read and comprehend

Lysosomal acid hydrolases are recognized via a“signal patch” and tagged with Mannose-6-phosphate in the golgi​:

• In the golgi, many proteins are receiving modifications ​

• Signal patch... Area of the protien that is acting as a signal, but if you isolate the AA, they are not next to eachother linearlly... they are only next to eachother when they fold (why its called a patch) ​

• Phosphotransferase transfers Nano6 phosphate onto mannose ​

​

read and comprehend

M6P receptor binds M6P-tagged proteins tosort them into transport vesicles​:

• Vesicle cargo receptor M6P receptor on the trans face of the golgi bindsM6P-tagged proteins​

• M6P receptor is located on a budding clathrin-coated vesicle​

• clathrin is another type of vesicle coat protein​

• Vesicle buds off of golgi (same process we learned about last lecture!)​

Vesicle cargo receptor (M6P) receptor that binds any protein with the M6P tag ​

M6P then is on a budding clathrin-coated vesicle... M6P associated with clatherin and acid hydrolyases (putting them in as the vesicle forms) ​

Veiscle buds off the golgi (same process for inittation, budding and scission... just some of the proteins are different) ​

read and comprehend

pH-dependent dissociation of M6P receptor and hydrolase:

Catherin coated vesicle is going to bud and scission off the golgi, loosing the clathrin coat (becomingg a naked vesicle) ​

This vesicle travels to the endosome... going through teathering and docking ​

Golgi is more netural pH than endosome ​

So in slightly acidic environment, the acid hydrolases is released from lysosome and becomes inside the endosome ​

Removes phosphate but mannose remains (mature lyososome form) ​

Have not gotten to the lysosome yet! Is at the endosome right now ​

Answer: 1, 2, 3, and 5 ​

Sar1 GTPase is a specific protein associated with COPII proteins ​

Everything else is correct! ​

Teathering protein: resposible for picking up Rab protein on vesicle and to teather to membrane ​

Docking: where T snares and V snares engage to bring vesicle close; need to release teathering protein for this to occur ​

T snares are found on target membrane; v snares are on the vesicle ​

Fusion: T snares and V snares reverisbly bind together to smush membrane together... becoming one with target membrane

Which of the following would be necessary for fusion of the clathrin-coated vesicles containing the lyososomal acid hydrolyases with their target membrane? ​

1. t-SNAREs​

2. v-SNAREs​

3. Rab GTPase ​

4. Sar1 GTPase ​

5. teathering protein​​

read and comprehend

The endosome/lysosome system​:

• Endosomes are similar instructure to lysosomes butnot quite as acidic (pH~6.0)​

• They receive vesicle cargos from the plasma membrane as well as the golgi​

• They also recycle cargos back to the plasma membrane and/or golgi via vesicle transport​

• They are a sorting compartment and intermediary on the way to the lysosome​

​

Endosome and lyososome exist in a system, where the endosome becomes a lyososme at the end of its life ​

Can receive and sort cargos to put them in other locations through vesicle transport ​

The endososme is a sorting compartment and a middle man until its ready to go to the lyososome

• Endosomes ultimately fuse with lysosomes to deliver their cargo​

• can be classed as “early” or“late” endosome based on how close they are to fusing with a lysosome​

• Early and late endosomes can be distinguished based on particular proteins present/absent​

How long will it be for them to complete life cycle: early vs late ​

Can tell them apart based on the prescence or absence of certain proteins ​

Early endosomes contain materials that are not destined for the lyososome since they will go other places; late endosomes contain materials only destined for the lyososome ​

Autophagy

endosome

Phagocytosis

Lysosomes degrade material supplied by phagocytosis, endocytosis and autophagy​:

Endosomes can collect materials from different places to be degraded by the lyososome ​

_____: forms a membrane around the autophagosome (artificial endosome) to be targeted for degradadtion ​

Cell can uptake through vesicle transport to bring things from outside in... which fuses with _____, which then eventually fuses with lysosome ​

____: brings something big from outside (phagosome) to be trageted from degradation ​

• Autophagy

• Proteasome

• organelles

• build up

• recycle

Autophagy is the process of the cell ingesting entire parts of itself including parts of membrane bound organelles:

• ____ is used for larger things like organelles ​

• _____ is for smaller things like individual proteins ​

• Forms a membrane around that using phospholipids in cytosol to surround completely and form a compartment. This compartment is fused with the lyosoome to degrade ​​

Autophagy allows cells to:​

• dispose of ____ that are not functioning properly​

• prevent _____ of macromolecules such as lipids, protein aggregates,etc.​

• ____ cell components when nutrients are scarce​

If a mitochondira has been fissioned off, it will be degraded using autophagy ​

Also is used to get rid of build up of macromolecules ​

Allows the cell to reuse cell components as well ​

Answer: induce autophagy ​

• Bc it will use recycling process to gain nutrients that it is not getting from the environment

Would you expect starvation to: induce autophagy, supress autophagy, not affect autophagy​

1. isolation membrane; elongates

2. autophagosome

3. lysosome

4. degraded

Process of autophagy:​

1. ______ forms + _____ around autophagy targets​

2. complete membrane forms; the “______”​

3. autophagosome fuses with a ____​

4. contents are ____ by lysosomal hydrolases​

4 steps ​

1. Form the membrane around autohagy targets ​

2. Complete the membrane... becoming a completely enclosed compartment (called autophagosome when its completely closed) ​

3. Fuses with lyosome ​

4. This releases acid hydrolyases that degrades once they are fused ​

When they fuse, the proton pumps are still on, which makes the pH lower/closer to lyososome (pH of 5) ​

• Atg8/LC3

How do cell components get targeted for autophagy?​

• Process is slightly different for each organelle or typeof cell component​

• Generally, targets are tagged with a signal (often ubiquitin)​

• This signal is recognized bya receptor​

• Receptors interact with _____, a protein in the autophagosome membrane​

Multiple tagging pathways​

In general, they are tagged with a type of signal (some type of ubiqutination... but a different form that tags to the proteasome) ​

NOT THE ONLY SIGNAL ​

Different receptors for different signals... usually found in the cytosol. When it finds the ligand, the receptor binds and calls in a protein called Atg8/LC3 which helps form the autophagosome membrane ​

• The autophagosome then forms around the Atg8/receptor/target complex​

• A lysosome fuses with the autophagosome to form the autophagolysosome​

• This fusion also requires specialized SNAREs!​

• Hydrolases degrade targets​

Once this complex is formed, the membrane forms​

It has many copies of Atg8/LC3 on the outside and the inside ​

The autophagosome then fuses with the lysosome ​

Once they fuse, the pH drops, which activates the acid hydrolyases, which degrades what is inside the autophagosome ​

• neurological diseases

Defects in autophagy have been linked directly to neurological diseases

such as Parkinson’s Disease and Alzheimer’s Disease:

• Autophagy happens in most cells; its an important process for the maintiance in cell health ​

• However, the defects of this process is linked to _____​​

lifespan

Stimulating autophagy can increase lifespan!​

Most of the time, if you decrease something the cell does its unhelpful. However, the reverse (increasing = better) is not usually true ​

However, if you increase autophagy, it can increase the ___ by almost double ​

Pathways are conserved across evolution... not just in one animal but conserved in most! ​