Lecture #10 Flashcards
what are lysosomes?
ubiquitous organelles located in the cytoplasm
what are the functions of lysosomes?
involved in autophagy, secretion, in the repair of the pm and in the energy control metabolism in the cell
what are some examples of diseases associated to lysosomal dysfunction?
Alzheimers, Parkinsons, MS
what is an example of a cell type that is more sensitive to lysosomal dysfunction?
neurons
what type of organelles are lysosomes?
intracellular organelles with a very acidic lumen - lysosomal enzymes work in an acidic pH
where are the lysosomes located?
in the perinuclear space - there are different patterns depending on the cell stage (can either be perinuclear or very far from the nucleus)
describe the shape of lysosomes:
either spherical or tubular → size and number are quite heterogeneous depending on the stage of the cell cycle and the type of cell
describe lysosome biogenesis:
lysosomal enzymes follow the normal protein synthesis in the cell, then they are packaged into vesicles in the golgi, are modified, and delivered to the lysosomes via a specific tag
what is lysosomal biogenesis regulated by?
many intracellular and extracellular signals → the state of the cell impacts biogenesis
describe the membrane of the lysosome:
a phospholipid bilayer membrane rich in carbohydrates
describe the lumen of the lysosome:
there are many soluble hydrolyses as well as activators
describe the other membrane formations in the lysosome:
interfaces between the intra-lysosomal vesicles are important because some enzymes are bound to these intramembrane formations
describe the lysosomal proteins:
these are not only hydrolyses and activators, but also intramembrane or membrane-associated proteins such as LYNUS
all proteins are classified according to their function (structural proteins, channels, protein that regulate fusion, catabolic enzymes, transporters, etc)
what is the primary function of the lysosomes?
the degradation of substrates
where does the primary function of lysosomes occur?
on the surface of the intra-lysosomal vesicles catalyzed by hydrolyses
about how many hydrolyses are located in the lumen?
around 70
describe the cycle of lysosomes:
- entry of the material into the lysosomes either from the inside or the outside of the cell
- degradation of the material resulting in the formation of building blocks
- building blocks are recycled
*must be tightly regulated in order to maintain the correct energy metabolism in the cell
describe the organization of lysosomes:
they are part of a membrane system (sometimes it is difficult to see the boundaries between different membrane compartments such as the golgi, vesicles, endosomes, etc)
how are hydrolyses driven to the lysosomes?
they have specific tags → in the golgi they are tagged with specific molecules in particular mannose 6 phosphate (M6P) which is recognized by M6P receptors
do all enzymes get delivered directly to the lysosomes?
no - a fraction of the enzymes escape and are secreted directly outside the cell
what happens to the enzymes that escape?
since they are tagged with M6P they can be re-uptaken by neighboring cells that express M6P receptors → cross correction
what is enzyme replacement therapy?
based off of the cross correction system: if there is an enzyme deficiency, the functional copy can be provided by healthy cells or by gene therapy → the secreted enzymes can be recaptured by the deficient cells that do not produce the enzyme themselves
describe the endocytic route:
plasma membrane - early endosomes - lysosome - recycled back to plasma membrane
what is the pH of the lysosome?
5
what is an example of a lysosomal product where transporters are necessary?
cholesterol
how is cholesterol formed?
the lysosome degrades lipoprotein complexes (proteins and lipids) releasing cholesterol to be transported out of the organelle into the cytosol
in what types of cells do lysosomes exert exocytosis function in order to get rid of toxic material or aggregates?
secreting cells such as hematopoietic cells, osteoclasts, melanocytes, or in non-physiological conditions
how does the lysosome function when the cells are in a state of starvation?
they can sense the lack of nutrients and activate a response that provides the cells with nutrients through lysosomal-mediated degradation and autophagy
how does the lysosome function when the cells are in an anabolic state?
they can synthesize proteins and lipids to shut down autophagy
how many different forms of hydrolyses does the lysosome contain?
70
how is the pH of the lysosome maintained?
there is a hydrogen pump that carries out active transport, and imports H+ ins inside the lysosome against the concentration gradient
describe the proteins that are embedded in the membrane of the lysosomes:
complexed with sugars which face the organelle lumen to protect the membrane from the acidic environment
what is the function of LYNUS?
huge complex of proteins that serves as the lysosomal nutrient sensing machinery
what are the two types of channels in lysosomes?
CIC7 (chloride) and MCOLN1 (muco-lipin)
describe the function of CIC7:
helps the hydrogen pumps make the environment more acidic
describe the function of MCOLN1?
a calcium channel which regulates the efflux of the calcium
what other channels are important for calcium import?
TPC1-2 : two-pore channels that assist the entry and exit of calcium
what is the category of trafficking and fusion machinery?
SNARES
in order for a vesicle to fuse with the lysosome, what must be true?
their membranes must be compatible in terms of composition and membrane identity
what is the player of recognition between vesicles, endosomes, or autophagosomes which fuse with the lysosome?
RAB7
what is the name of the group of signaling lipids necessary, and what specifically is the key master regulator of fusion with the lysosome?
phosphoinositides ; phosphatidylinositol-3,5-bisphosphate
which are the three SNARE proteins that allows for fusion?
vSNARE, tSNARE, and VAMP
what type of enzymes are found on the surface and in the lumen that are devoted to degradation and catabolism?
catabolic enzymes
what is the function of LAMP2A?
transporter that mediates chaperone-mediated autophagy
what are the two different routes of autophagy?
membrane (macroautophagy) and chaperone-mediated
how do proteins brought by chaperone-mediated autophagy interact with the lysosome?
pass from the cytosol, and are incorporated into the lysosome directly without fusion
what must proteins contain in order to go through chaperone-mediated autophagy?
KFREQ specific pentapeptide
what chaperone recognizes proteins that need to be degraded by this pathway, and brings them to the lysosome?
HSPA8
what specific transporter is involved in cholesterol export into the cytosol?
NCP1
how can LAMP1 be used in the laboratory?
it is a protein that can be used to recognize lysosomes became it is an integral structural protein embedded into the membrane
what is the fusion of the lysosomes and the pm dependent on?
calcium - dependent
what channels (among others) release calcium in relation to lysosomal-pm binding?
MCLON1 and TRPML1
in regards to the fusion of the lysosome and the pm, which snare proteins are present on the lysosome that mediate this interaction?
V-SNARE and VAMP7
in regards to the fusion of the lysosome and the pm, which snare proteins are present on the pm that mediate this interaction?
T-SNARE, SNAP23, and syntaxin4
in what type of cells is it common for membrane repair to be needed?
muscle cells - constantly exposed to mechanical stress
what is the interaction between the lysosome and membrane repair regulated by?
calcium and the protein Dysferlin → must have both
what are dysferinopathies?
muscular dystrophies or other atrophies of muscle cells due to the apoptosis as a result of continuous damage (often pediatric onset)
what are calpains?
proteases that digest the actin cortical cytoskeleton temporarily to allow for vesicle fusion
what stage of endoscope interacts with the lysosome?
late endosome → forms the endolysosome
what occurs after digestion in the endolysosome?
the lysosome buds off again from the endolysosome to form a new lysosome
what occurs after the autophagosome and the lysosome fuse?
they become the autolysosome and degrade the material due to the acidic pH, then there is lysosomal reformation
what process is extracellular material digested by?
formation of multi-vesicular bodies forming the late endosome and then finally joining the lysosome to become the endolysosome
what is chaperone-mediated autophagy recognized by on the lysosome?
LAMP2A
where do lysosomal hydrolyses come from?
- synthesized in the ER
- phosphorylated in the cis gogli to attach a mannose and phosphate forming the m6p tag
- go through several cisterns to reach the trans golgi
- bud off in clathrin-coated vesicles
what two reasons is Claritin coating fundamental?
to interact with actin (light chain) and Bar proteins that help the pm curve, and to select the cargo that needs to be transported
what does clathrin work with in order to be able to perform its function?
adaptors like AP2 which recognize receptors that are embedded in the vesicle
what happens after the clathrin-coated vesicle is delivered?
clathrin / COPI / COPII are recycled back to the ER
describe the concept of cross-correction:
if you put two cells close to each other, the WT can cross correct the deficient cell through the exocytosis of the enzyme which is recognized by the receptor of the deficient cell
when treating lysosomal deficiency disorders, what cells are used for engineering,?
hematopoietic cells → precursors of monocytes. macrophages which are able to colonize every tissue of the body
why are hematopoietic cells used as the vessel?
in a condition where there are not as many lysosomal enzymes present, there is a lot of inflammation so the environment favors the recruitment of monocytes and macrophages
what are caveolae?
patches in the pm of high density proteins and lipids. that different from clathrin, are not formed dynamically when needed but are always present
what are some examples of selective autophagy?
mitophagy, xerophagy, pexophagy, aggrephagy
where are the two main areas where membranes are sourced for autophagy?
the pm or ER
name some of the positive regulators of autophagy:
Beclin 1, Vps34 (a subunit that regulates PA trikinase class 3), UVRAG
without these proteins the autophagosome cannot be formed and engulfment doesn’t occur
which ATG proteins are important for autophagosome elongation, and are needed to select cargo, mature, and go through the lysosome?
ATG 5, 7, 12
what is the anterograde movement mediated by?
kinesics that move by hydrolyzing ATP
what is used for transport from the periphery to the perinuclear region?
dyneins
what is basal autophagy and what is an example?
when the ER is too big, so part of it needs to be degraded
how are the different autophagic states regulated?
SLCA1 pathway
