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Cell signalling > Gap Junctions > Flashcards

Gap Junctions Flashcards

1
Q

The connexin family

A 
21 family members, 21 genes
Number gives molecular weight
Little splicing
Present in all chordates
Not present in pre chordates
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2
Q

Innexin/pannexin family

A

Gap junctions in pre chordates
8 genes in drosophila, 25 in c.elegans.
Splicing can produce different variants e.g. ShakB lethal, neural and neural +16
Other family members named numerically
Pannexins- innexins that were kept in chordates

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3
Q

Original members found in the innexin/pannexin families

A

DmshakB- Defective escape
Dm-ogre- small optic lobes
Ce-unc7 and unc9- uncoordinated movement
Ce-eat5- feeding defect

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4
Q

Membrane topology

A

4 TM domains
Two extra cellular loops
Single intracellular loop
No sequence homology- example of convergent evolution

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5
Q

Structure of gap junction

A

6 protein subunits
TM3 lines the pore
Extra cellular fixed in conformation by disulphide bonds
Interdigitation

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6
Q

Gap junction TM3

A

Channel narrows from 40A to 15A

This is because TM3 tilts

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7
Q

Opening and closing of gap junction channel

A

Rotation and tilting of TM3
Movement of either small or bulky AAs in the pore
N terminus may possibly swing and block the pore

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8
Q

Biochemical regulation

A

Ph- protonation of histidine residues in c terminus or I loop
Ca- Ca:H transporter makes the cell more acidic.
Activates phospholipase C and A2 -> DAG and arachidonic acid
These integrate into membrane around pores
Calmodulin can bind to gap junction proteins and prevent them entering membranes
Different connexins have different c-termini so can respond differently to Ca changes

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9
Q

How would you tell if a region is calcium sensitive?

A

Removed region and measure sensitivity

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10
Q

Phosphorylation

A

Ser, thr or tyr kinases on c tail
This can alter signalling between channels
Kinases activated by growth factors and neurotransmitters

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11
Q

Voltage

A

N terminus AAs sense membrane potential

Conserved Pro in TM2 gives a 15-20 link and can block pore

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12
Q

Transcriptional

A

The gap junction proteins have a very short half life

3-4 hour turnover

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13
Q

Size and charge selectivity

A

Different sequences in pore lining regions
Different permeability- prefer anions or cations
Max 2kDa but some only 500Da

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14
Q

Assembly

Not all combinations work together

A

Homomeric or heteromeric hemichannel
Homotypic or hetrotypic typic channel
Cx43 and 40 are incompatible- can be used to create compartments in embryos

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15
Q

Analysing channel function

A

Dye transfer- cascade blue injected
Voltage clamp- micro electrodes in both cell can send and receive voltage
Patch clamp inserted into membrane can measure single channel

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16
Q

Chemical vs electrical synapses

A

Electrical is x5 faster (0.1ms)
Electrical can be uni or bi directional
Electrical are generally excitatory

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17
Q

Neuronal gap junctions- types of synapse

A

Cx36 is major, also 45 and 57
Bidirectional/self rectifying- either direction, sync neutrons for memory
Unidirectional/non rectifying- one direction, when lots of neutrons fire. Escape responses

18
Q

Giant fibre physiology

A

Two GFs from each side of brain connected by GCI (giant commissural interneuron)
GF synapses
TTMn -> tergotrochanteral muscle motor neuron
PSI peripherally synapsing neuron -> DLMn dorsal longitudinal flight muscle

19
Q

Experiment to prove shakb in gap junctions

A

Shakb RNA injected

Cell pairs voltage clamped, those with shakb showed voltage transmission

20
Q

Alternative splicing of shakb

A

Shakb neural- in the developing CNS not GFS
Shakb neural +16- primarily GFS
Shakb lethal- CNS inc GFS and gut and heart

21
Q

Where is shakb neural + 16 found?

A

Found in the GF system

22
Q

Is rectification due to heterotypic gap junctions?

A

Rectifying electrical synapse is a heterotypic junction
Composed of neural 16 (pre) -> lethal (post)
Found in TTMn and PSI
Asymmetrically hated by voltage-
N+16 is relatively positive and lethal negative
Supports a positive to negative flow
Shakb (n+16) Homotypic channels show little voltage sensitivity, likely to be gated in post synaptic lethal

23
Q

Gap junctions mutations in the ocular lens

A

Lens fibres no organelles depend on epithelial
NaKATPase in epithelial gives a translenticular potential and current flow
Cx 43 + 50- epithelium
46 50 - lens fibres

46 and 50 give different phenotypes although same place
50 deletion gives slow growth and microphthalmia
Both give nuclear cataracts

24
Q

Female infertility

A

Cx37
No ovulation
Follicle stops growing as Antrum formed
Growth promoting factors from follicular cells per ate through gap junctions to make ooycytes mature

25
Q

Charcot Marie tooth

A

10-20% are cx32 X linked
Autosomal- peripheral myelin protein 22 or p0
Weak distal muscles, reduced touch, CNS defects less severe
X100 faster from peri nuclear to periaxonal
Myelin dies

26
Q

Recycling of K+

Role of cx

A

1 epithelial network- organ of corti
2 connective tissue network- spiral ligament and stria vascularis

Cx26 might couple cells or supply glucose for survival

27
Q

Cx in hearing loss

Two types

A

Non syndromic and syndromic
Cx26 -> non syndromic but can cause skin alone, 50% of cases
Most cx26 non syndromic are recessive but all syndromic cx26 are dominant
May be more redundant in the skin and compensated for

28
Q

Role of Cx in the skin

A

Expression of different connexin markers as cells move up from the stratum basal
No junctions are found in the stratum corneum
Cx26 may control keratinocytes differentiation, if a mutation causes over proliferation this gives flaking skin
Communication departments in cell proliferation

29
Q

Vonwinkel syndrome

A 
Cx26
Dominant
Palmoplanter hyperkeratosis
Auto amputation 
Hearing loss
30
Q

Keratitis ichthyosis deafness (Kid)

A 
Mutations in NT of ECL1 of cx26
Determines pore size and sensitivity
Keratitis (inflamed cornea) blindness
Ichthyosis (dry thick scaly skin) prone to infection
Widespread skin
Hair and hearing loss
31
Q

Connexin expression in the heart

Speed of the connexins

A 
SA- 45,40
AV- 45,40 (0.05m/s)
Atria- 40,43
His- 40,43
Purjunke- 40,43 (5/ms)
Bundle- 45
Ventricle- 43

40- large channels (0.8-1 across atria)
43- medium channels
45- low conductance channels

32
Q

Atrial fibrillation

A

Atria don’t contract, blood pooling
Thrombosis risk of embolic stroke
Can be acquired after hypertension, arterial disease
Idiopathic is autosomal dominant mutations in cx40, 10-20%

33
Q

Ventricular fibrillation

A

Causes pump failure
Ventricles twitch instead of contracting
Cx43

34
Q

Atrial standstill

A

Loss of electrical and contraction in atria

Polymorphism in cx40 and mutation in SCN5A for Na+ channel

35
Q

Sudden infant death syndrome

A

Cx43
Lone AF
Ventricular tachyarrythmias (fast rate)

36
Q

Non junctional functions of connexins

A

Can act independent
Open hemichannels can release ATP to act on receptors on other cells
Regulation stops leak of molecules

37
Q

Types of taste buds

A

1- glial like
2- receptor, sweet bitter umami
3- sour and salty, release serotonin onto neuron

38
Q

Pannexin in taste buds

A

T1R and T2R GPCRs on type 2 cells activated
Activates PLCB2 raising Ca
TRPM5 calcium gatedchannels open, NA+ entry
Depolarisation opens more NA channels
Pannexin 1 channels open- regulated by voltage and Ca
ATP released
ATP acts on P2X on type 3 to depolarise
Type 3 release serotonin

39
Q

Why not conventional transmission in taste buds?

A

Receptor cells continuously replaced

Release of ATP means that synaptic contacts aren’t made and broken

40
Q

TTMN
GCI
PSI
DLMN

A

Tergotrochanteral
Commissural interneuron
Peripherally synapsing
Dorsal longitudinal

Cell signalling (5 decks)

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