Cilia and Ciliopathies

Question 1

What are cilia

Answer 1

• microtubule-supported structures
• protrude from apical surface of all ccells
• in epithelia they have specialised functions
• membrane-bounded and contain a thin layer of cytoplasm
• length of single cilium is 1-10uM and width is less than 1uM
• all cilia show active transport of proteins, ions and nutrients across ciliary membrane and from cell cytoplasm to shaft
• receptors are moved to and from ciliary stem

Question 2

Specialised functions of cilia

Answer 2

• particle and pathogen clearance (e.g. mucociliary clearance in airways)
• monitoring (e.g. pressure/volume sensing)
• sensory perception (e.g. rod structures of retina; hair cells of cochlea)

Question 3

Loss of ciliary function

Answer 3

results in group of epithelial diseases known as ciliopathies

Question 4

Motile cilia

Answer 4

• found in airway of lungs, middle ear, fallopian tube, and brain cisterns (spaces)
• e.g. nasal turbinate cells (motile): mucociliary clearance of inhaled particles and pathogens
• e.g. hair cells of cochlea: mechanical transduction of tectorial membrane/tympanum movement to auditory sound

Question 5

Non-motile (primary) cilia

Answer 5

• found in a wide variety of organs
• e.g. kidney collecting duct: senses changes in urine flow and volume by shear on cilium
• e.g. retinal cells: photoreception

Question 6

Structure of hair cell cilia

Answer 6

• afferent and efferent nerve endings
• rich in neurotransmitters
• depolarisation is necessary for sound transduction
• each stereocilia are tethered by ‘top-link’
• as cilia move relative to one another, top link terminates on potassium channel which opens with each movement of the cilia
• flickering opening of potassium channels causes release of neurotransmitter, allowing us to hear

Question 7

Structure of kidney collecting duct cilia

Answer 7

• single structure
• like radio anteni that stick upwards into tubular lumen
• sense rate of flow and composition of fluid flowing over apical surface of cells
• monitors urine osmolarity and transduces information back to cell through gap junctions
• modification of ion transport fine tunes composition of urine

Question 8

Structure of retinal ciliary

Answer 8

• highly specialised
• cytoplasmic extensions that form plates
• extended structures (rods) allow you to see black and white
• afferent and efferent nerve fibres

Question 9

Structure of primary cilia

Answer 9

• coated in thin layer of cell membrane
• thin layer of cytoplasm covers the inner microtubular structure
• microtubules exist in 9 pairs with no central pair
• axoneme extends into cilium

Question 10

Structure of primary cilia

Answer 10

• coated in thin layer of cell membrane
• thin layer of cytoplasm covers the inner microtubular structure
• microtubules exist in 9 pairs with no central pair
• axoneme extends into cilium

Question 11

Intrafraglabellar transport (IFT)

Answer 11

• allows for transport of proteins up length of cilia (proteins deposited either within cytoplasm or within membrane surrounding the cilium)
• kinesin can bind to MT and ciliary package, and moves substances into the cilium
• IFT dyenin is used to cargo substances out of cilium by offloading at tip of cilium
• upward process known as anterograde movement
• downward process known as retrograde

Question 12

Basal body

Answer 12

• composed of centriole
• anchors cilium in place
• anchors important cell signalling proteins
• loss of structure causes loss of organisation of epithelial structure due to lack of sequestering effect

Question 13

Role of Nexin cross-links

Answer 13

bind microtubule pairs together, must be broken/reduced to facilitate movement

Question 14

Formation of cilia: G0 phase

Answer 14

• for epithelial cell to gain apical and basal polarity, cell must enter G0 phase
• movement out of G0 to cell cycle associated with cell growth (good for immature epithelial cells, but can lead to cancer if gets out of hand)

Question 15

Formation of cilia: M phase

Answer 15

• centrioles anchor spindles during mitotic division
• single ‘maternal’ centriole is retained which can anchor to cell membrane in G1 or G2 and S phase
• can either use maternal centriole as template for growing second centriole, or (in an epithelial growing sheet) maternal centriole can act as baseplate that extends upwards to form cilia
• latter locks cell into G0 phase and is now fully polarised and functional
• this cell can only divide further if ciliary structure or junctions holding cell together are disrupted

Question 16

Steps involved in outer segment formation of cilia

Answer 16

• morphogenesis begins when mother centriole contacts ciliary vesicle
• axonemal extension from centriole causes cailiary vesicle to invaginate and form ciliary sheath
• fusion with plasma membrane externalises developing outer segment and transforms outer sheath to perciliary membrane
• disc formation and outer segment extension

Question 17

How do motile cilia move

Answer 17

• microtubule structures have arms with dyenin protein attached to them
• arms held together by nexin proteins
• dyenin reaches on shaft of neighbouring microtubules
• hydrolyses ATP and can then spin around and bind way along length of microtubule (goofy feet)
• rapid (nasal cilia can beat 60 times a second)

Question 18

Beating of motile cilia

Answer 18

• directional but not symmetric
• waves 1-9 collapse forward and is termed the recovery stroke
• ## as cilia turns, we get stiffening waves that produce movement, shown in 10-12

Question 19

Steps of Anterograde and Retrograde

Answer 19

1) IFT complex assembly in cytoplasm next to basal body
2) IFT complex consists of IFT particles which bind cargo dynein or kinesin
3) kinesin 2 is molecular motor that moves IFT complex along each MT pair towards top of cilium (anterograde)
4) cargo is offloaded at tip of cilium
5) turnover products for removal are loaded onto IFT complex
6) dynein is molcular motor that moves IFT complex along MT pair towards ciliary base place (retrograde)
7) IFT complex dis-assembly, turnover products proken down and IFT particles (dyenin and kinesin 2) are recycled

Question 20

Growth factor signalling

Answer 20

• e.g. platelet derived growth factor
• require ciliary localisation to activate downstream targets (e.g. ERK signalling)
• regulates cell growth and proliferation
• ERK drives cell growth in response to attachment to PDGF receptor found in primary cilia
• endocrine signal giving cell early warning that message is coming

Question 21

Maintenance of cell polarity

Answer 21

• cilium can alter model of signalling through same receptor pathway
• cilium promotes Wnt signalling to planar cell polarity (PCP)
• pathway (promotes cell polarisation) and inhibits canonical Wnt singalling to nucleus
• beta catenin acts as loss of polarisation stimulus (e.g. cancer stimulus)

Question 22

Cell differentiation

Answer 22

• SHH signalling stabilises cell against loss of differentiation and maintains characteristics of the epithelial cell
• SHH binds to the patched receptor (Ptc)
• relieves inhibition of smoothened (smo) and allows TF Gli to migrate to nucleus
• influences embryonic development
• drives loss of polarisation during EMT in cancer

Question 23

What are ciliopathies

Answer 23

• rare diverse range of diseases that affect multiple organ systems
• two classifications: motile (e.g. CF) and primary
• may involve multiple genes which affect ciliary strucutre/function
• defined by shared clinical features and proteins which are found at or near cilia
• arise due to disruptions in signalling processes, caused by loss of cilia

Question 24

Polycystic kidney disease

Answer 24

• primary ciliopathy
• cysts form along nephron and collecting ducts
• caused by gene mutation of Polycystin (PKD1…3)
• these genes regulate Ca2+ uptake in endoplasmic reticulum and mutations are linked to loss of Ca2+ signalling
• genes are localised to basal body of renal primary cilium
• mTOR is potent activator of cell growth and protein synthesis -> loss of mTOR repression by polycysteins results in formation of cysts
• when cilium is absorbed and cell polarisation is lost = cancer

Question 25

Cilia Dyskinesia

Answer 25

• motile ciliopathy
• recessive disorder that arises from mutations in genes involved in axoneme assembly
• affects any organ with motile cilia
• missing dyenin arms or central tubules