What is the difference between cytosolic IF and lamins with regard to protofilaments? In which compartment are lamins?
IF Protofilaments are arranged antiparallel
Lamin Protofilaments are arranged with an overlap of head-head with tail-tail
Lamins are exclusively located in the nucleus (nuclear lamina)
Please write down the following cytoskeletal proteins in an order with increasing filament thickness: microtubule, actin, vimentin, lamin A
lamin A (3,5 nm), Actin (6 nm), Vimentin (11 nm), Microtuble (24 nm)
What do actin drugs lactrunculin and cytochalasin do?
and what are other drugs for the same structure
Lactrunculin A binds to actin monomers to inhibit their polymerisation, promotes filament disassembly
cytochalasin: binds to barbed end to prevent actin filament assembly and disassembly
phalloidins inhibit actin filaments dissasembly by locking adjacent actin subunits together
Jasplakinolides: stabilize actin monomer: enhancing nucleation and assembly
Which end of microtubule is located a) at centromeres and b) at centrosomes?
a) plus end (Centromer = center of Chromosomes)
b) minus end → elongates via plus end (Centrosomes with mother and daughter)
What cell component does the drug colchicine affect and what part of cell division is affected by that? // Which part of cell division is disturbed by treatment of cells with Nocodazole or Colchicine and exactly which process is inhibited by the toxin?
Tubulin-specific-drug (Microtubuli)
→ binds subunits and prevents their polymerization
Microtubules required for spindle formation and, thus, chromosome segregation
Which structure in animal cells is shared by centrosomes and cilia and which protein component is the main constituent of this structure? // What do centrosomes and cilia have in common and where do they originate from?
Centrosomes and cilia both have microtubule-based structures
MTs based structures originate from the centriole
Name a major gamma-tubulin-binding scaffolding protein within the pericentriolar matrix
gamma-TuRC: nucleates assembly and remains associated with minus end
a) Where in epithelial cell do you find hemidesmosomes &
b) which type of cell adhesion proteins are employed to establish contact to the outside world?
a) basal surface of epithelial cells
b) integrins
Integrins are transmembrane proteins that serve as receptors for components of the extracellular matrix, such as fibronectin and laminin.
Function of retromer complex
receptor recycling
from MVB (multivesicular body, early endosome) to TGN
Name of complex which is required for vacuolar degradation of membrane proteins coming from the plasma membrane
ESCRT complex
What is the function of the multi-vesicular body/ late endosome (MVB/LE)
it sorts ubiquinated membrane proteins incorporating them into vesicles
degradation transmembrane proteins
Endosome fuses with MVB/PVC/LE then protein membrane gets invaginated
→ ESCRT complex generates intraluminar vesicles
How are membrane proteins degraded? Which complex is involved? // which complex is involved in the degradation of membrane proteins (in plants)? Briefly explain how it works.
endosome with membrane protein fuses with MVB → then invagination
invagination of membrane proteins (ubiquinated) through ESCRT complex
vesicle gets transported from MVB to vacuole and degraded
involved complex: ESCRT complex
What is the function of an autophagosome and what is its function?
double membrane vesicle → newly formed during autophagy
engulfs wide rage of intracellular material and transports it in vacuoles
degradation of large molecules
Which coat protein is typical for vesicles transporting mannose 6-phosphate receptors back from the early endosome to the Golgi apparatus?
Retromer coat. Retromer coated vesicles retrieve receptors from EE back to TGN
Why are phosphoinositides (PIPs) important for correct protein sorting?
Each membrane uses specific PIPs.
By binding their corresponding protein, PIPs assure that the protein is transported to the correct cell membrane
Name the general, common mechanism function of BIN2 and CTR1 proteins in their hormonal pathway. What common molecular activity of function do BIN2 and CTR1 proteins have in this pathway // What are BIN2 and CTR1 doing in their corresponding signal pathway? Which exact mechanism are the performing?
BIN2 (= brassinoid intensive 2, Brassinosteroid signal transduction)
negative regulators → in absence of hormone (BIN2 brassinosteroid, CTR1 Ethylene) act as a repressor kinase
→ is a kinase, that, when active, phosphorylates its target molecules (for BIN2: transcription factors BES1 and BZR1 )
→ TFs get inactivated and transported out of the nucleus
→ inhibit downstream signaling
When and where are PIN proteins relocalized during cell division?
When
PIN transporter regulate Auxin efflux → coordinate polar positioning
PIN recycling is first established directly after cell division
Where
At the apical end of the mother cell.
Then, when the cell plate starts to develop, PIN proteins are found in the cell plate as well. After cytokinesis, one daughter cell shows PIN proteins at the apical AND the basal cell, while the other daughter cell shows PIN proteins only at the apical cell. The PIN proteins at the basal side of the first daughter cell are vacuolarly degraded.
a) Which process does the PLEIADE/MAP65-3 protein of Arabidopsis thaliana contribute to? and
(b) how is this impressively reflected by the subcellular phenotype of pleiade/map65-3 loss-of-function mutants?
a) microtubules associated proteins of the MAP65 family crosslink microtubules
b) fail of differentiation → eventual destroyed
→ show no crosslinked microtubules but only parallel microtubules
Commonalities of different plant signal transduction pathways: name function that the following protein mechanistically have in common in plant signal transduction
a) common, general, mechanistic function of AUX/ IAA, JAZ and DELLA proteins in respective pathway b) common, general, mechanistic function of SCF^TIR1, SCF^COI1 and GID1 protein in respective pathway
SCF is receptor complex, TIR1 is the F-box protein
a) are repressor proteins of the transcription factors of Auxin, Jasmonate, Giberellin
→ inhibit transcription factors ARF, MYCZ, PIF3/4 when hormones are absent or at low concentration
Auxin
SCF TIR1 - receptor
AUX/IAA - repressor
ARF - transcription factor
Jasmonate
SCF COI1 - receptor
JAZ - repressor
MYC2 - transcription factor
Giberellin
GID1 // SCF SLY1 - receptor
DELLA - repressor
PIF3/4 - transcription factor
b) hormone receptor that, upon binding hormone, act as ubiquitin E3 ligase and ubiquitinate the respective repressor protein
→ ubiquinate repressor proteins so that it gets degraded by 26S proteasome and TF gets activated
What is the casparian strip and what is its function in plant cells?
diffusion barrier in cell wall of root endodermal cells
→ forces nutrients to pass into symplast cells (apoplastic barrier)
made out of lignins (polymerization of lignins in the cell wall region)
Why are the factors that cause planar polarity so different in plants and animals?
During evolution, multicellularity arose independently in the animal and plant kingdom (convergent evolution).
The last common ancestor was most likely unicellular.
Which factor builds up a tissue gradient and instructs planar polarity in plants?
hormone Auxin - key regulator of plant growth and development, and its asymmetric distribution can create tissue gradients and instruct planar polarity.
The root epidermis of Arabidopsis thaliana and many other plants displays a simple tissue polarity termed planar polarity: All root hairs polarily initiate at the ends of epidermal cells that are oriented towards the root tip. How was it experimentally demonstrated that a local auxin concentration gradient can instruct the coordination of polar hair initiation in the root epidermis of Arabidopsis thaliana?
By reintroducing a local auxin source in the form of auxin soaked sephadex beads.
Function of transmembrane proteins
a) Flippase b) Scramblase
a) Flippase = P-type ATPase → moves PE and PS from outer to cytosolic leaflet
b) Scramblase → moves lipids in either direction towards equilibrium
You have a concentration of K+ = 100nM and Na+ = 10 nM in the cytosol and K+ = 10 nM and Na+ = 100nM in the extracellular matrix. a) What is the membrane potential when only K+-channels are open? Multiple choice: -6 mV, -60 mV, -600 mV, -100 mV, 0 mV, 6 mV, 60 mV, 600 mV b) What is the membrane potential if K+ and Na+ channels are open?
a) answer: -60mV
b) answer: 0 mV
Name two methods to measure the activity of V-type ATPase
patch clamp technique
ATPase Assay
Name two modes/mechanisms for the regulation of V-type ATPases. How do they work in general?
“Regulated assembly” (reversible dissociation of the V-ATPase complex into its component V1 and V0 domains)
Regulated expression of V-ATPase subunits
Disulfide bond formation at the catalytic site of subunit A (reversible inactivation)
General working mechanism:
V-type ATPases are large, multisubunit complexes composed of two main sectors: V1 (cytoplasmic) and V0 (membrane-integrated).
The V1 sector contains catalytic subunits responsible for ATP hydrolysis, while the V0 sector forms a proton-conducting channel.
V-type ATPases utilize the energy derived from ATP hydrolysis to pump protons across membranes, creating an electrochemical gradient.
They are rotary motors.
How do V-ATPases work? Name two basic functions of V-ATPase in the plasma membrane of animal cells
It is a proton pump (establishing pH gradient) in endosomal and lysosomal membranes of animal cells
= H+-ATPase → pumps H+ from cytosol to extracellular space → acidify intracellular compartments
It is required in the plasma membrane of osteoclasts (=cells that degrade bone to initiate normal bone remodeling) and some kidney tubule cells.
a) Cytosol (unstimulated cells)
b) mitochondria (unstimulated cells)
c) ER (unstimulated cells)
d) extracellular space
a) 10^-4 mM
b) 0.1-0.8 mücroM (??)
c) 0.1 - 1 mM
d) 1-2 mM
Gap junction multiple choice – what can go through and structure
Pore Ø ~1.4 nm
Passage of molecules ≤1000 Da (Anorganic ions, incl. Ca2+; H2O; Sugars; AA; Metabolites; Nucleotides incl. ATP & cAMP; IP3)
No passage of proteins
Can be closed (regulated by e.g. Ca2+)
Gap junctions = Connection between 2 membranes (distance very small)
Connexin = Integral membrane proteins of the gap junctions
A gap junction channel consists of 2 Connexons.
Hemichannel = 6x Connexin (=Connexon)
Each connexon consists of 6 connexins, which are special membrane proteins.
What is the patch clamp technique?
used to study the electrical currents flowing through individual ion channels in cell membranes
What is the name of the pattern depicted here?
Left: Amplitude-modulated oscillating response
Right: Frequency modulated oscillating response
Left: Phasic response
Middle: Tonic response
Right: Phasic-tonic response
There are stimulus- and cell-specific differences in intracellular Ca2+-signals. The following two differences can be observed between various intracellular Ca2+-signals:
Spatial localization
Amplitude & Duration
Name two characteristics of protein kinase A
cyclic AMP-dependent protein kinase
2 catalytic + 2 regulatory domains
cAMP → catalytic subunit dissociates → subunits are activated
Name two characteristics of G-coupled proteins
3 subunits (alpha, beta, gamma)
attached to membrane via fatty acyl chain or alpha and gamma
alpha subunit is GDP bound and interacts with GPCRs
alpha subunit = GTPase
A cell that is able to respond to an inducing signal is considered competent. Which 2 mechanisms allow cells to acquire competence? // Competence may be required during development. Name two molecular mechanisms by which a cell can acquire such competence.
formation of gap junctions
expression of receptors
The heart is an electrical organ. Prior to cardiomyocyte contraction, action potentials result in release of intracellular Ca2+ from the sarcoplasmatic reticulum. Name those cellular junctions between cardiomyocytes that are involved in electrical coupling of cardiomyocytes
gap junctions
adherens junction
What 3 hormones are produced within endocrine glands of pancreatic islands of Langerhans?
Glucagone (alpha-cells)
Insulin (beta-cells)
Somatostatin (D-cells)
Name 2 principles of how morphogens work. What are 2 characteristics of morphogens?
Principles
induce different cell fates at different threshold concentrations
diffusion over several cell diameters
Characteristics
Cells can acquire an identity based on positional information
Must have concentration-dependent effect on its target
Must exert a direct action at a distance
TGFß
= transformin growth factor
TGFß binds Typ II receptor , Typ II receptor phosphorylates Typ I receptor
phosphorylated Typ I receptor recruits and phosphorylates SMAD2 or SMAD3
SMAD2/3 dissociates from receptor and oligomerizes with SMAD4
oligomer translocates to nucleus → recruits gen regulators
→ activation of target gene
Wnt pathway
presence of Wnt
Wnt binds Frizzled
recruitment of Dishevelled, Axin and LRP6
ß-catenin degradation inhibited
ß-catenin accumulates to transcription factor → gene transcription active
absence of Wnt
ß-catenin gets degraded → no gene transcription
Hedgehog pathway
with hedgehog
internalisation of patches → degradation
Smoothened phosphorylated by PKA and CK1 → moves to plasma membrane
recruitment of protein complex → inhibition of Ci proteolyse
Ci released → transcription activated
Within cardiomyocytes binding of Myosin to Actin filaments is controlled by a regulatory protein complex compromised of several proteins. Name 4 regulatory proteins of this complex // What are the proteins involved in regulating the interaction between actin and myosin?
referring to the Actin-Troponin-Tropomyosin complex which consists of actin, troponin, tropomyosin (blocks myosin binding site on actin) and myosin
Regulatory proteins are:
TroponinC (TnC, Ca2+ binding site)
TroponinI (TnI, inhibition site)
TroponinT (TnT, tropomyosin binding site)
Tropomyosin (blocks myosin binding site on actin)
Select the correct quote on how diffusion from the capillary system is mediated to the interstitium (multiple choice) a) Colloid osmotic pressure in capillary, hydrostatic pressure in interstitium b) hydrostatic pressure in capillary, colloid osmotic pressure in interstitium c) hydrostatic pressure in capillary, hydrostatic pressure in interstitium d) Colloid osmotic pressure in capillary, hydrostatic pressure in capillary
b is correct → hydrostatic pressure in capillary, colloid osmotic pressure in interstitium
capillary efflux/ influx
arterial side: hydrostatic pressure pretty high (in capillary) compared to colloid osmotic pressure
→ gradient becomes less
→ Filtration
venous side: colloid osmotic pressure higher than hydrostatic pressure to regain liquids from the loss (90%) 10% from lymphatics
Multiple choice about Osmotic pressure
colloid osmotic pressure in interstitium
higher at venous side
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