Practice exam

• Which of the following statements is FALSE?

  • The cytosol is always viscoelastic and mechanically heterogeneous
  • Mechanical stretch to the cell results in deformation of the whole cell
  • The nucleus is the least deformable organelle in the cell
  • None of the above

  None of the above
• For one particular cell type, it has been estimated that a filament persistence length of at least 5 μm is necessary for the cells to create stable membrane protrusions. When a compound X is added to

  the culture medium, this threshold persistence length becomes 10 μm. Which of the following is a possible mechanism by which compound X could affect the cell?

  • It doubles the cell cortical tension
  • It doubles the polymerization rate of actin filaments.
  • It doubles the rate of Cdc42 activation
  • It doubles the bundling size of actin filaments
  • It doubles the cell cortical tension
• Actin treadmilling results from...

  • Disturbance to the steady state equilibrium of actin polymerization
  • The force balance at the membrane that resist further actin polymerization
  • The difference in the availability of monomeric actin at the two ends of the filament
  • The difference in the polymerization rates at the two ends of the filament


  The difference in the polymerization rates at the two ends of the filament
• An actin filament is growing normally when it encounters a small network of bundled actin filaments held together by depletion interactions. Considering the magnitude of forces involved in actin

  dynamics and interactions, what is the most likely scenario?

  • The single actin filament buckles and depolymerizes
  • The single actin filament stops polymerizing and gets recruited into the network
  • The single actin filament unzips the actin bundles and destabilizes the network
  • The single actin filament gets pushed back by the network as it continues to grow

  The single actin filament unzips actin...
• The component of ECM is most responsible for its interaction with cells is...

  • Glycoproteins
  • Collagens
  • Proteoglycans
  • Glycosaminoglycans

  Glycoproteins
• When fibrin concentration is unchanges but the number of protofibrils within fibrin fibers increases, ...

  • Both the fiber stiffness and network stiffness will increase proportionately
  • The network will deform less affinely
  • The network will exhibit more plasticity
  • Temperature will become more influential in determining the nework mechanics

  The network will deform less affinely
• Which of the following is NOT part of the adhesion complex?

  • Focal adhesion kinase
  • Alpha-actinin
  • Vimentin
  • Vinculin

  Vimentin
• Durotaxis...

  • Requires force transmission through the focal adhesion
  • Is induced by a spatial gradient in substrate stiffness
  • Is always directionally anisotropic
  • All of the above

  All of the above
• Cells can sense the viscoelasticity of the ECM because...

  • The matrix promotes cell polarization and regulation of GTPases
  • The matrix can store mechanical stresses exerted by the cells
  • The matrix can recognize and sustain more cellular forces
  • The matrix can allow sustained actin threadmilling

  The matrix can recognize and sustain more cellular forces
• Talin is a cellular mechanosensory because...

  • It physically connects integrin to actin filaments and allows force transmission
  • It can change conformation when mechanically loaded, promoting binding to vinculin
  • Its binding to vinculin and actin filaments becomes stronger when mechanically loaded
  • It is required in the formation and maturation of the focal adhesions

  It can change conformation when mechanically loaded, promoting binding to vinculin
• What is the essential principle behind the operation of optical stretcher?

  • Use of real-time morphology analysis to determine the particle mechanical properties
  • Use of two diverging laser beams to trap a particle
  • All of the above
  • Use of microfluidic system to manipulate particle

  Use of two diverging laser beams to trap a particle
• You are using microrheology to measure the viscoelasticity of your cell cytoplasm. You find out that the displacement of the probes you use is too small to track accurately. What can you do to solve this issue?

  • Use an improved algorithm for particle localization
  • Use higher imaging resolution
  • Use camer with better sensitivity
  • Use larger probes

  Use higher imaging resolution
• What is an advantage of using microtissue gauges as a method to measure cell forces?

  • It allows simultaneous application of external forces
  • It allows the use of viscoelastic ECM
  • It allows tuning of the anisotropy of the 3D cell environment
  • All of the above
  • All of the above
• What is NOT an advantage of using micropillar arrays as a method to measure cell forces?

  • It allows simultaneous application of external strain
  • It allows the use of substrates with a wide range of stiffnesses
  • It provides subcellular measurement resolution
  • It allows simultaneous application of haptotactic cues

  It allows the use of substrates with a wide range of stiffnesses
• Which mutation will result in decrease in tissue size?

  • Mutation in Ajuba that inhibits alpha-catenin binding
  • Mutation in alpha-catenin that inhibits actin-binding
  • Mutation in myosin that decreases its catalytic activity
  • All of the above

  All of the above
• Which of the events described below are accompanied by an increase in tensile forces on E-cadherin adhesions?

  • An epithelial layer increasing in cell density
  • An epithelial cell proceeding through mitosis
  • A cell extruding from the epithelial layer
  • All of the above

  An epithelial cell proceeding through mitosis
• Traction force microscopy uses beads...

  • As a way to tune the elasticity of the substrate
  • To ensure that the cells can adhere to the substrate
  • As a fiduciary marker to track the deformation of the substrate
  • To increase the resolution of the measurement

  As a fiduciary marker to track the deformation of the substrate
• The mechanosensory Piezo plays an important role in epithelial homeostasis and is activated when epithelial cells become compressed at high cell density. Which cellular process is induced by Piezo under this condition?

  • Extrusion
  • Apoptosis
  • Quiescence
  • Cell cycle exit

  Extrusion
• Which of the following statements about cell type and cell ID in the Cellular Potts Model (CPM) is TRUE?

  • A pixel can switch between both cell type and cell IDs
  • Every cell has a unique cell ID and cell type
  • The contact energy is only evaluated between cells of different cell types
  • Every cell has a unique cell type

  A pixel can switch between both cell types and cell IDs
• Pixel copy attempts are accepted according to an acceptance probability distribution P, which is designed to minimize the energy function H. Which of the following statements ... is true?

  • The acceptance probability P decays exponentially for H<0
  • For every high temperature T, the P is effectively zero for pixel copy attempts that increase H
  • Because of the quadratic functional form of the constraints encoded in H it possesses a unique minimum
  • For very low temperature T, the P is effectively zero for pixel copy attempts that increase H

  For very low temperature T, the P is effectively zero..
• What is NOT an example of post-translational control in bacterial gene expression?

  • Induced protein degradation
  • Attenuation
  • Riboswitches
  • Allosteric RNA polymerase activation

  Allosteric RNA polymerase activation
• Which of the following statements about CAR-T immunotherapy is FALSE?

  • T-cells are engineered to express the clustered antigen receptor (CAR)
  • Engineered T-cells are re-transfused into the patient again
  • Engineered T-cells can recognize, bind and kill the cancer cells
  • T-cells are isolated from the blood of a patient

  T-cells are engineered to express the clustered antigen receptor (CAR)
• Which enzyme is NOT part of the Gibson assembly mastermix?

  • DNA polymerase
  • DNA ligase
  • 5' exonuclease
  • Alkaline phosphatase

  Alkaline phosphatase
• Which of the techniques below allows the generation of cell substrates for studying haptotaxis?

  • Photopatterning
  • Rapid prototyping
  • Microcontact printing
  • All of the above

  All of the above
• You intend to use a 3D scaffold in your study. To achieve the goals of your project, two critical criteria must be met for the scaffold: (i) the range of material used to fabricate the scaffold must be as wide as possible and (ii) the distribution of local mechanical stresses in the scaffold must be as homogeneous as possible. Which of the following techniques will be most appropriate for creating your scaffold?

  • 3D printing
  • Electrospinning
  • Selective laser sintering
  • Salt leaching

  3D printing
• Main cytoskeletal components

  • Actin filaments: soft, (semi-)flexible polymers. Persistence length 10-20 um. Highly dynamic, can polymerize and depolymerize to allow cell dynamics and motility. Involved in cell contraction (together with motor protein myosin)
  • Microtubules: stiff filaments. Persistence length in mm-cm range. Plays a role in cell division and molecule transport (together with motor proteins dynein and kinesin)
  • Intermediate filaments: mechanical properties between actin and microtubules. Protects the nucleus. Cross-links with actin and microtubules
• Explain the role of Cdc42, Rac and Rho in regulating the cytoskeleton organization.

  • Cdc42 regulates membrane protrusions, filopodia
  • Rac regulates lamellipodia
  • Rho regulates stress fibers
• Describe the cell morphology and actin organization in these images.

  • Flask 1: actin mostly at the cell edges forming lamellipodia -> Rac activation
  • Flask 2: plenty of actin stress fibers throughout the cell body -> Rho activation
  • Flask 3: cell forms a lot of finger-like protrusions (filopodia) -> Cdc42 activation
  • Flask 4: cell has diverse actin organization -> control cells
• Molecular tension probes

  • Concept: single-molecule probe for sensing single-molecule forces
  • Create molecule in which you can look into the distance such that if the molecule is deformed it can tell you how much it is deformed. If you know deformation and mechanical property then you should be able to determine force
  • If cell applies some force through the integrin, that will be translated at molecule level. If molecule is stretched, then the distance between fluorophore and quencher becomes larger. So quantify how much molecule is stretched by looking at fluorophore-quencher interaction
• DNA hairpin force sensor

  • Concept: use DNA hairpin instead of proteins/polymers as molecular spring
  • When cells binds to ligand and applies force -> DNA hairpin unzips, the distance between fluorophore and quencher becomes higher -> you get a signal
• Explain for both Vinculin and Ajuba which cellular process is regulated by their force-induced recruitment to E-cadherin adhesions.

  • Vinculin: strengthens cell-cell junctions by providing additional linkage of the cadherin complex to the actin cytoskeleton, therby enabling cell-cell junctions to withstand forces without breaking.
  • Ajuba: relieves the inhibition of the nuclear translocation of YAP (by sequestering its inhibitory kinase LATS), leading to nuclear accumulation of YAP and regulating cell cycle entry/proliferation
• Explain how forces on the E-cadherin complex can induce transcription via beta-catenin/TCF.

  • Tensile forces on the cadherin complex result in beta-catenin being phosphorylated (by Src)
  • This decreases its affinity for E-cadherin, resulting in their dissociation and allowing beta-catenin to translocate into the nucleus (and bind to TCF)
• In eukaryotic cells, DNA is first transcribed into a primary mRNA transcript (pre-mRNA) and then processed into a mature mRNA transcript. Describe the steps that are needed to turn a pre-mRNA into a mature mRNA and explain why they are important.

  1. Splicing
  2. 5' capping
  3. Polyadenylation
• Splicing

  Not all the RNA nucleotides in the primary transcript are translated into proteins. The sequences that do not code for a protein are called introns and are actively removed by a large complex called the spliceosome in a process called splicing. Exons are sequences that appear in the final mRNA and are translated by proteins. From one pre-mRNA multiple mature mRNA can arise due to a process known as alternative splicing, this is another way to regulate gene expression in eukaryotes.
• 5'Capping

  mRNA in eukaryotic cells are also modified at the 5 and 3 prime ends. Modification of the 5'-ends of eukaryotic mRNA is called capping. The cap consists of a methylated GTP linked to the rest of the mRNA y a 5' to 5' triphosphate 'bridge'. (Capping occurs very early during the synthesis of eukaryotic mRNAs, even before mRNA molecules are finished being made by RNA polymerase II). The cap is important in the recognition by ribosomes to induce translation. Capped mRNAs are very efficiently translated by ribosomes to make proteins.
• Polyadenylation

  Modification of the 3'-ends of eukaryotic mRNAs is called polyadenylation. Polyadenylation is the addition of several hundred A nucleotides to the 3' ends of mRNAs. The poly(A) tail protects the mRNA molecule from enzymatic degradation in the cytoplasm and aids in transcription termination, export of the mRNA from the nucleus, and translation.
• Name all components involved in the CRISPR-Cas9 based genome editing. Briefly discuss their function and describe how DNA is recognized in this system.

  • Cas9
  • crRNA
  • tracrRNA: contains a complementary sequence to the crRNA and results in a conformational change in the Cas9 protein by binding of a RNA hairpin at the 3' end to the Cas9 protein.
  • PAM site is a 2-6 basepair DNA sequence immediately following the DNA sequence targeted by the Cas9 nuclease in the CRISPR bacterial adaptive immune system.
  • DNA is recognized via Watson-crick base pairing by crRNA (or gRNA)
• Briefly describe the working principle of the methods for producing microfabricated model systems.

  • Soft lithography
  • Microcontact printing
  • Photopatterning
• Soft lithography

  • You have a mold, often made of silicon or glass
  • Pour PDMS on top of mold
  • PDMS solidifies or polymerizes
  • Peel off PDMS from mold
  • PDMS will now have specific texture or shape
• Microcontact printing

  • With microfabricated topography you always have a third dimension that comes out of the plane of the substrate, but this is not always desirable as it complicates the interpretation of the results.
  • Microfabricated substrate used as a stamp to create 2D substrate that defines adhesion areas for the cells.
  • Less physiological as it is 2D, but makes interpretation easier.