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Trinity Term 1 > Fertilisation > Flashcards

Fertilisation Flashcards

1
Q

Methods of achieving fertility differ between organisms…

A
  • nutrient acquisition (materials of the egg must be used to drive the cell cycles)
  • differences in number and size of eggs
  • necessitates sex trade-offs within the breeding system
  • different parental response and behaviours
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2
Q

Describe Futus rhizoid fertility

A

attach their offspring to rocks to photosynthesise.

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3
Q

Describe Drosophila fertility

A

larvae

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4
Q

Describe frog fertility

A
  • tadpoles
  • rely on metamorphosis, and differing metabolism and different life stages
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5
Q

Describe chicken fertility

A

use an albuminous egg

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6
Q

Describe mouse fertility

A
  • such as Mus musculus
  • placental fertility system
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7
Q

Describe sea urchin fertilisation

A
  • external
  • easier to visualise
  • fourteen peptides released by the egg, creating a chemotactic gradient up which the small, flagellate and numerous sperm can navigate in its ambient marine environment
  • egg releases carbohydrate signals to initiate the calcium ion mediated exocytosis of the sperm’s acrosomal vesicle
  • sperm releases necessary enzymes for extracellular envelope penetration
  • stimulates the actin-dependent formation of the acrosome for delivery of the male nucleus
  • sperm and egg membrane fusion creates the fertilisation membrane
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8
Q

extracellular envelope

A

also termed the zona pellucida

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9
Q

How is species-specific recognition achieved in sea urchin fertilisation?

A

binding proteins on the acrosomal process

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10
Q

Describe fusion in mammals

A
  • sperm binds to the zona pellucida, so that its acrosome is anchored between cumulus cells of the cumulus cell layer
  • Izumo1 protein must bind to the egg receptor Juno
  • allows the acrosome to react and mature, and penetrate through the zona pellucida to the perivitelline space
  • Izumo-RFP protein localises at the acrosomal membrane
  • attachment of the opened acrosome to a cortical granule then allows fusion of the plasma membranes, and the deposition of the sperm nucleus and contents into the egg cytoplasm
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11
Q

polyspermy

A

the fusion of multiple sperms

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12
Q

‘block’ to polyspermy has two mechanisms

A

the fast and the slow.

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13
Q

Describe the fast block to polyspermy

A
  • achieved via a fertilisation-mediated membrane depolarisation event
  • small local calcium influx through calcium channels in the internal plasma membrane initiates a calcium wave
  • changes potential from -70 to +20mV
  • membrane gradually repolarises.
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14
Q

Describe one model for the fast block to polyspermy

A
  • enzyme-activated phosphorylation of PI to PIP to the PIP2 substrate in the plasma membrane
  • phospholipid-
  • activates PKC
  • hydrolyses IP3, releasing it into the cytoplasm
  • phosphorylation of the InsP3 stimulates other signalling events
  • allows calcium release from the ER at the IP3 receptor, which can go to PKC
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15
Q

PI

A
  • phosphatidyl inositol
  • 2 acyl chains, glycerol phosphate and sugar,
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16
Q

DAG

A
  • diacylglycerol
  • highly hydrophobic
  • usually acts as a landing site for phosphoprylating protein kinases
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17
Q

PKC

A

protein kinase calcium

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18
Q

InsP3

A

small, charged, diffuse and hydrophilic

19
Q

Describe the self-stimulation of calcium release in the fast block to polyspermy

A
  • further calcium on the cytoplasmic side primes the next IP3 receptors
  • stimulates further calcium release
  • non-equilbrium feedforward loop
20
Q

Describe ‘store operated calcium entry’

A
  • replace calcium so that it is not lost in the ER
  • cellular calcium influx at the STIMI, coupled to the plasma membrane at the ORA1 membrane receptor
  • allows calcium to transfer to the SERCA protein calcium pump embedded in the ER membrane
21
Q

How can the fast block to polyspermy be visualised?

A
  • rapid confocal imaging
  • Medaka fish eggs through luminescence after microinjection of aequorin
22
Q

aequorin

A
  • calcium ion photoprotein
  • isolated from Aequoria victoria
23
Q

Describe aequorin imagery of the fast block to polyspermy

A
  • multiple calcium waves spreading through the egg
  • triggered by a calcium ionophore
24
Q

Describe the visualisation of PIP2 hydrolysis

A
  • fluorescent tagging via microinjection of PIP2 with a chimera of GFP-PH of eggs of Lytechnius pictus
25
Q

PH

A

pleckstrin homology domain of PLC

26
Q

Describe the egg-receptor mediated polyspermy block model

A

sperm derived factor fertilin would activate an integrin egg receptor via a tyrosine kinase, thus activating PLC.

27
Q

Describe the sperm oocyte-activating factor-mediated polyspermy block model

A
  • IP3 is directly released by the sperm derived PLCz isoform, via the PIP2 already present in the cytoplasmic oocyte vesicles
  • results in the activation of an src-tyrosene kinases that activate the egg PLC
  • IP3 also releases calcium ions from the endoplasmic reticulum, via IP3R
28
Q

Describe the slow block to polyspermy

A
  • cortical granule fusion, mediated by the calcium wave
  • CICR
  • InsP3 enhances sensitivity of the InsP3-R, and cADPribose to calcium
  • elevated cytosolic calcium imitates further calcium release from either ER channel, in a non-equilibrium feedforward loop
  • cortical granule and vesicles can be exocytosed (gramnosecretion), resulting in calcium-protein and particle mediated-fusion
  • fertilisation envelope layer can form
29
Q

Describe calcium ‘puffs’ or ‘sparks’

A

calcium propagation via clusters of channels across the egg

30
Q

CICR

A
  • calcium-induced calcium-release
  • regenerative process where local calcium release spreads into a calcium wave or tide
  • activates a plethora of downstream targets
31
Q

cADPribose

A

natural ligand of the Ryanodine-receptor

32
Q

Describe the fertilisation envelope layer

A
  • polymer of mucopolysaccharides, cross-linked vitelline proteins and hyalin
  • hardens the zona pellucida, protecting the egg and preventing further sperm entry
  • the slow black to polyspermy
33
Q

How can the slow block to polyspermy be visualised?

A

using fluorescent reporters

34
Q

Summarise the polyspermy block

A
  • sperm fusion results in membrane depolarisation: the fast block to polyspermy
  • tyrosine kinase activation, which activates PLC
  • release of IP3 and DAG
  • CICR
  • activation of NAD+ kinase, producing NADPH, and exocytosis of the cortical granule
  • formation of the hyaline layer: slow block to polyspermy
35
Q

What moves an egg from quiescence?

A
  • CICR acts in combination with DAG, which activates PKC - results in sodium and potassium ion exchange for an increase in pH
  • stimulation of protein synthesis and DNA replication to activate the egg from quiescence and begin division
36
Q

Describe the first division of the embryo

A
  • male pro-nucleus must migrate to meet the female pro-nucleus
  • four molecules of calcium can bind co-operatively to the four EF-hand domains of calmodulin
  • CaM-kinase II activated by a transient calcium signal localised around the spindle
  • autophosphoryaltion
  • calcium can be released to recycle in its binding to calmodulin, leaving the CaM-kinase II calcium independent, but remaining active, at a 50-80% activity rate
  • Cam-kinase II can then go on to phosphorylate many other molecules
37
Q

Describe male pro-nuclear migration

A
  • indirect process of flow-mediated migration via motor drag on the microtubules
  • generation of cortical cytoplasmic flow sweeps the two nuceli together
  • visualised under mathematical modelling
38
Q

Describe the zygotic microtubules

A
  • derived from the astromicrotubules
  • extend from the centrosome, forming spindles at the site of sperm entry
  • reconstitution of the mitosis apparatus
39
Q

Describe calmodulin

A

4 domains separated by an alpha helix each

40
Q

Describe co-operative binding of calmodulin

A
  • induces major conformational change and preferential localisation at the mitotic apparatus during division
  • rapid transition to full binding on a single binding event (sigmoidal binding response)
41
Q

Describe the activation of CaM-kinase II

A
  • binding of calcium ions to calmodulin into a complex
  • exposure of its receptor protein for activation
  • allows allosteric binding onto the inhibitory domain of CaM-kinase I
  • winds catalytic domain inwards and blocked by an inhibitory molecule
42
Q

Describe the autophosphorylation of CaM-kinase II

A
  • produces ADP from ATP
  • phosphate binds to the allosteric inhibitory domain and allows the catalytic domain to unwind, and inhibitory molecule to dissociate
  • full activation
43
Q

Describe the inactivation of CaM-kinase II

A
  • protein phosophotase can dephosphorylate the calcium independent CaM-kinase II
  • produces a molecule of inorganic phopahte
  • compound becomes inactive once more
44
Q

Describe the downstream effects of CaM-kinase II activation by calcium

A

Activation of this CaM-kinase II by calcium results in the inactivation of cytostatic factor, which thus stimulates centrosome duplication, and activates anaphase promoting factor.

Trinity Term 1 (38 decks)

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