PHSL233 Term Flashcards
Which processes are required for establishing epithelial polarity?
a) Neighboring cells interacting with each other through adherens junctions
b) Connections between epithelial cells and their basement membrane
c) Nectin proteins from neighboring cells connecting with each other
d) All of the above
d) All of the above
Which is the correct order for entry of secretory pathway proteins into the endoplasmic reticulum (ER)?
a) signal peptide cleaved - protein enters via translocon - binding of ribosome to ER
b) protein enters via translocon - signal peptide cleaved - SRP (signal recognition particle) binds signal sequence
c) SRP binds signal sequence - ribosome docks on ER - protein enters via translocon
c) SRP binds signal sequence - ribosome docks on ER - protein enters via translocon
Choose the CORRECT statement about polarity protein complexes:
a) aPKC phosphorylates Scrib polarity proteins to keep them basolateral
b) Both the Par and Crb complexes are basolateral
c) The Scrib complex is basolateral
d) A and C are correct
d) A and C are correct
Which order is correct for trafficking of a protein in the secretory pathway?a) Endoplasmic reticulum (ER)-endosome-Goldi-cell surfaceb) endoplasmic reticulum (ER)-Golgi-cell surfacec) nucleus-lysosome-cell surface
b) endoplasmic reticulum (ER)-Golgi-cell surface
Which is the CORRECT order for entry of secretory pathway proteins into the ER?a) signal peptide cleaved - protein enters via translocon - binding of ribosome to ERb) protein enters via translocon - signal peptide cleaved - SRP (signal recognition particle) binds signal sequencec) SRP binds signal sequence - ribosome docks on ER - protein enters via translocon
c) SRP binds signal sequence - ribosome docks on ER - protein enters via translocon
“Which statement about protein topology is CORRECT?
a) ““stop-transfer”” sequences remain in the ER membrane as transmembrane domains
b) In the ER, the region of a transmembrane protein in the cytosol will end up outside the cell
c) The N-terminal end of a protein always enters the lumen of the ER.”
“a) ““stop-transfer”” sequences remain in the ER membrane as transmembrane domains”
Which post-translational modification occurs in the ER?a) transcription; N-linked glycosylation; degradation
b) assembly of protein complexes; disulphide bridges; O-linked glycosylation
c) GIP anchors added; N-linked glycosylation; chaperone-mediated folding
c) GIP anchors added; N-linked glycosylation; chaperone-mediated folding
Endocytosis of transmembrane proteins occurs through interaction with the AP2 compex BECAUSE the transmembrane proteins contain endocytosis motifs such as YXXφ.
a) Both statements are true and causal
b) Both statements are true but not causal
c) First statement true, second false
d) First statement false, second true
e) Both statements false.
a) Both statements are true and causal
Choose the CORRECT statement about protein sorting and targeting in epithelia
a) The exocyst is the site for delivery of proteins to the basolateral membrane
b) Experimental evidence shows that the trans-Golgi network (TGN) can sort apical and basolateral proteins into separate vesicle populations
c) In the indirect sorting pathway, all proteins are first delivered to the same membrane domain.
d) All of the above are correct.
d) All of the above are correct.
Lysosomes only degrade proteins BECAUSE monoubiquitin is used as a tag for degradation in the endocytosis pathway.
a) Both statement true and causal
b) Both statement true but not causal
c) First statement true, second false
d) First statement false, second true
e) Both statements false
d) First statement false, second true
functions of the kidney
-regulation of water and ion balance, body pH-excretion of metabolic wastes-excretion of foreign chemicals-endocrine functions (calcitriol, erytropoietin, renin-gluconeogenesis
Define polarity
A difference in structure, composition or function between the two poles of a cell, such as, apical/basolateral in an epithelial cell.In epithelial cells this also means location of a protein in a specific location in the cell membrane
Which statement is CORRECT about Kidney?
a) The kidney filters about 200L of plasma /day
b) The kidneys do not produce hormones
c) The urinary system has 3 ureters
d) The kidneys excrete foreign chemicals
e) The kidneys receive 50% of the blood flow
d) The kidneys excrete foreign chemicals
how much plasma do the kidneys filter each day?
~180L
Why is polarity important in epithelia?
Depending on the exact transporters and channels present in each membranedomain this will dictate whether that epithelia primarily absorbs or secretessubstances
how much of the total filtrate is reabsorbed?
99%
How are epithelia ‘made’?
If single epithelial cells are placed on a substrate that resembles the basementmembrane, the cells attach to that substrate and start showing signs of somepolarity. If the cells are allowed to divide so that all cells are close to their neighbours, aswell as linking with the basement membrane, cell-cell contacts such as tightjunctions now develop, and the cytoskeleton starts forming under thebasolateral membrane. Ca2+ is needed for cell-cell contacts to form. Upon Ca2+ addition adherens junctions and then tight junctionsformed and the Na+K+ATPase now localised to the basolateral domain. However,full epithelial polarity is not observed until 36 hours after tight junctions areformed
renal blood flow
-kidney supplied by renal artery-renal artery divides to form glomerular and peritubular capillaires, and vasa recta-renal veins tale blood away from kidneys
What are the three protein polarity complexes?
Par, Crb and Scrib
types of nephrons
-superficial (cortical)-juxtameduallary
How is epithelial cell polarity established?
-Formation of adherens junction (AJ, nectin, E>cadherin) and interactions between cells and basement membrane-Small GTP proteins (cdc42) activated-Cdc42 activates aPKC, Par complex associates with apical domain-Crb complex associates with Par complex-aPKC phosphorylates proteins causing change in location e.g. scrib complex to BL region-TJ proteins start forming TJ apical to AJ-Full mutual exclusion of polarity complexes: Par/Crb complex in apical region; scrib in basolateral region-TJ fully formed, apical-basolateral polarity established
3 types of processes of the nephron
-glomerular filtration-tubular secretion-tubular reabsorption
What is the trafficking pathway that plasma membrane proteins follow?
CytosolEndoplasmic reticulumGolgiCell surface
Which statement is INCORRECT about Nephron?
a) The nephron is the smallest functional unit of the kidney
b) The afferent arteriole branches to make the glomerulus
c) Each kidney contains more than 1 million nephrons
d) Reabsorption of material is from the blood to the lumen of the nephron
e) Juxtamedullary nephrons are located within the medulla of the kidney
d) Reabsorption of material is from the blood to the lumen of the nephron
How do proteins enter the endoplasmic reticulum
Proteins contain an address label called a signal sequence that guides a ribosome translating a cell membrane protein to theendoplasmic reticulum. The signal sequence is bound by the signal recognition particle and translation stops until the ribosome docks onto the endoplasmic reticulum. Translation starts again and the protein is threaded through a ‘pipe’ or protein translocation channel called the translocon into the lumen of the endoplasmic reticulum.
What is glomerular filtration?
-bulk flow of protein-free plasma into the bowman’s space (forms filtrate)-the filtrate contains everything in plasma except RBC and protein
Which statement is FALSE about Renal Clearance and GFR?
a) The typical GFR for a healthy person is 125ml/min
b) RC is the volume of plasma from which a substance is cleared by the kidneys per unit time
c) GFR is not a useful indicator of renal function
d) Creatinine is a substance that can be used to determine GFR
e) GFR is the amount of filtrate produced per unit of time
c) GFR is not a useful indicator of renal function
Which statement is TRUE about Body water?
a) 65% of a male’s body is composed of water
b) Normally, a person has 4 liters of plasma in their blood
c) A person can live without water for 3 months
d) The Ascending limb of the loop of Henle is water permeable
e) The net water intake and water output of person per day is 0.
e) The net water intake and water output of person per day is 0.
Which statement is TRUE about Water handling by the nephron?
a) AQPs are only found in animals
b) AQP2 is located in the apical membrane of proximal tubule cells
c) An osmotic gradient is NOT required for water movement
d) The proximal tubules reabsorb 50% of the filtered water
e) AQP3 and AQP4 are located in the basolateral membrane of collecting duct cells
e) AQP3 and AQP4 are located in the basolateral membrane of collecting duct cells
Which statement is FALSE about Renal Corpuscle?
a) The renal corpuscle contains the glomerulus and the Bowman’s capsule
b) Podocytes do not aid with filtration
c) The basement membrane is (-) charged to assist filtration
d) Jonah Laomu died from nephrotic syndrome
e) The endothelium of the glomerulus is fenestrated
b) Podocytes do not aid with filtration
renal corpuscle structure
-glomerulus-bowmans capsule (and space)-podocytes
How is correct protein topology achieved?
Hydrophobic and positive stretches of amino acids within a protein facilitate the orientation or ‘topology’of a protein
filtration barrier
-fenestrated endothelium of the glomerular capillaries (freely permeable to water, ions and small solutes)-basement membrane (matrix of negatively charged proteins, charge based barrier)-podocytes (gaps between foot processes, prevents protein and large macromolcule filtration)
Post-translational modifications at the ER
-glycosylation (N-linked)-GPI anchors added-protein folding (mediated by molecular chaperone proteins)-assembly of protein complexes
Nephrotic syndrome
increased permeability of filtration barrier to proteins
Post-translational modifications at the golgi
-N-linked glycosylation-O-linked glycoslyation-Sulfation of sugars and some tyrosines-protein sorting
“Which statement is TRUE about Reabsorption/Secretion?
a) 100% of the filtered urea is reabsorbed by the nephron b) Reabsorption is the only renal process that ““recovers”” filtered substances and water
c) Secretion is not an energy intensive process
d) Normally, 95% of the filtered glucose is reabsorbed by your kidneys, unless you are a diabetic
e) Secretion is the only renal process that leads to excretion of materials and fluids”
“b) Reabsorption is the only renal process that ““recovers”” filtered substances and water”
glomerular ultrafiltarion pressures and net equation
-hydrostatic-oncotic (osmotic)-PUF = PGC-PBS-πGC
ER assisted degradation system
- A protein (or substrate) is recognised as being non-functional ormisfolded by the same molecular chaperones that are helping to fold thatprotein.2. Next, the protein is sent out of the ER backwards through a proteintranslocation channel (translocon). This is called retrotranslocation ordislocation.3. The protein needs to be marked for degradation. This occurs throughubiquitin pathway enzymes tagging the protein with ubiquitin.4. Any sugars that were added to the protein in the ER are removed from theproteins (deglycosylation) and recycled.5. Finally, the marked proteins are threaded into a large barrel-likestructure called the proteasome, and the chemical bonds between aminoacids are broken (cleaved by enzyme proteases) releasing amino acidsand short peptides for reuse in new proteins.
pressures favouring filtration
-glomerular capillary hydrostaic pressure (PGC)
The process of membrane vesicle budding and fusion
Clathrin coated vesicles move proteins from the Golgi to the plasma membrane. vSNARE protein on vesicle binds tSNARE protein on target membrane. With Rab they force vesicle fusion with cell membrane
pressures opposing filtration
-Bowman’s space hydrostatic pressure (PBS)-glomerular capillary oncotic pressure (πGC)
trans golgi network
Uses COP1, COP11 and clarathin vesicles. The trans-Golgi network is specialised for the purpose of partitioning offareas containing proteins for different destinations. ENaC and Na+K+ATPasecomplexes move to plasma membrane via constitutive pathway.
epithelium in the proximal tubule
-leaky absorptive epithelia
fluid phase endocytosis
-molecule begins in solution-particles not bound to receptors are endocytosed-a clathrin-coated endocytotic vesicle carries the endocytosed material
epithelium in distal tubule and collecting duct
-tight absorptive epithelia-(under hormonal control)
receptor-mediated endocytosis
-molecule begins in solution-the ligand attaches to receptors-a clathrin-coated endocytotic vesicle carries the endocytosed material
% water reabsorbed per day
99%
caveolae endocytosis
small indentations in the plasma membrane called caveolae can mediate clathrin-independent endocytosis
Apical and basolateral sorting signals
The sequences YXXϕ,NPXY, LL, and monoubiquitin are known endocytosis motifs in plasmamembrane proteins (ϕ=hydrophobic amino acid). These endocytosis motifs arerecognised by clathrin-binding adaptor protein complexes (called adaptins). The specific adaptin in clathrin-coated pits developing at the plasma membrane for endocytosis is adaptin complex 2 (AP2). AP2 is made up of 4 subunits: α and β adaptins (bind to clathrin), μ2 adaptin (binds YXXϕ) and σ2subunits.
% sodium reabsorbed per day
99.5%
% glucose reabsorbed per day
100%
possible protein trafficking pathways for sorting ofapical and basolateral proteins in epithelial cells
- Direct sorting: apical and basolateral targeted proteins are sorted atthe trans-Golgi network into separate apical and basolateral destined tubularextensions or vesicles. 2. Indirect sorting: all plasma membrane proteins are packaged intovesicles at the trans-Golgi network and the vesicles ALL fuse at the basolateralmembrane. Then all the apical proteins are retrieved and targeted to the apicalmembrane.3. Random sorting: all plasma membrane proteins are packaged intovesicles at the trans-Golgi network and the vesicles randomly fuse with plasmamembrane all over the cell. Proteins in the wrong location are endocytosed andtrafficked to their correct location
reabsorption pathway
tubule to blood
experimental evidence for the direct protein sorting route
-Epithelial cells in culture, expressing basolateral and apical proteins tagged with different coloured fluorescent tags (green and blue).-Experiment: cool to 20oC to accumulate proteins at the trans Golgi network, warm up cells to 37oC and use live cell imaging to ‘watch’ the proteins.- VSV-G-blue and green-GPI were sorted into distinct vesicular carriers at the TGN.
secretion pathway
blood to tubule (from peritubular capillaries)
what compartments do the transferrin and immunogolbin receptors move through in epithelial cells?
After endocytosis from the basolateral membrane the transferrinreceptor moves tothe basolateral early endosome (where transferrin and iron arereleased), common endosome, and back to the basolateral membrane.IgA receptor is endocytosed from the basolateral membrane, moves to the basolateral early endosome, common endosome,then to the apical recycling endosome and from there it is delivered to the apical membrane. This is called transcytosis.
glomerular filtration rate (GFR)
-amount of filtrate produced per unit of time-normal value 125ml/min (180L/day)-a usual indicator of renal functioning
compartments for apical membrane proteins to move through in epithelial cells
Apical membrane proteins (ENaC) are traffickedto the apical early endosome. Here they will be degraded or recycled.If degraded, it will move to the late endosome,multivesicular body and then to the lysosome. If recycled it will move from the apical early endosome to the common endosome where it may mix with basolateral proteins, then it moves to the apical recycling endosome and finally back to the apical membrane
renal clearance (RC)
-volume of plasma from which a substance is completely cleared by the kidneys per unit of time-conc of substance in urine and plasma, and rate of urine produced must be known- C=Us x V/Ps (for substance ‘s’)
define protein half life
Protein half-life describes the time that half of the original population of aprotein remains.
short lived protein example
cell cycle regulators (destroyed within minutes)
how can GFR be estimated using RC?
-RC of inulin or creatinine = GFR-these substances are not reabsored from tubule, not secreted into tubule and not metabolised
daily filtered load
DFL = GFR x[substance]plasma
long lived protein example
proteins in the lens of the eye(may last a lifetime)
transport maximum (Tm)
-there is a limit to the amount of a substance transported per unit time-the transport protein can saturate-the binding sites on transport proteins become staurated when the conc of the substance increases beyond a certain limit (the Tm)
role of lysosomes in protein degradation
- Degrade transmembrane proteins that are targeted to lysosomes as anendpoint in the endocytosis pathway,- Degrade other proteins/sugars/nucleic acids and also organelles that aretoo old or not functioning correctly.- Contain acid hydrolases - enzymes that work at acidic (low) pH to breakapart proteins, sugars and nucleic acids. Low pH is maintained by H+ andCl- being pumped into the lysosome
an increase in body water leads to ?
increased excretion of water by the kidneys
autophagy process
- A specialised form of lysosomal degradation where a double membranestructure called an autophagosome develops around an organelle or areaof cytosol that needs to be destroyed.- The autophagosome fuses with the lysosome and contents are degraded,with breakdown products released into the cytosol for recycling.- Pathway switched on during starvation.
a decrease body water leads to ?
increased reabsorption of water by the kidneys
enzymes of the ubiquitin system
-E1 activating enzyme, E2 conjugating enzyme, E3 ligase- The first enzyme (E1) activates ubiquitin using ATP and covalently links to ubiquitin. - The E1 transfers the ubiquitin to a second enzyme (E2 conjugating enzyme). - The E2 conjugating enzyme transfers the ubiquitin to a third enzyme (E3) that chooses the substrate that the ubiquitin will be attached to. This E3 ubiquitin ligase may first bind to ubiquitin and then transfer it to the protein to be degraded
water gains
-fluid and food (diet)-metabolically produced
single ubiquitin tag…
Targets protein for endocytosis
water losses
-sweat, faeces, urine
string of ubiquitins…
Targets protein to the proteosome (where it is threaded into the barrel of the proteasome and chopped up into small peptides that can be recycled by deubiquitinating enzyme on the proteasome
which segments of the nephron are permeable to water?
-proximal tubule-thin descending limb of LoH-Late distal tubule and collecting duct (under ADH influence)
Lysosome mediated degradation
-Acid hydrolases-Breaks down proteins, sugars, DNA/RNA-Somewhat random, but ubiquitin tagging needed for some substrates.-Mainly cell surface and extracellular proteins (except autophagy). Molecules endocytosed
what are aquaporins?
a family of plasma membrane proteins that form channels permitting a high rate of water flow
Proteosome mediated degradation
-Peptidases-Breaks down proteins, ubiquitin recycled-Highly regulated-Requires a series of three enzymes: E1, E2 and E3-Cytosolic, nuclear, ER proteins (ERAD)
how much water do the proximal tubules receive per day?
180L
