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BL3303 > Willin > Flashcards

Willin Flashcards

1
Q

When does myelination occur?

A

In late embreyonic gestation and post-natal development in a caudal-rostral gradient in the CNS before developing in the PNs

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2
Q

What is an internode made from?

A

An oligodendrocyte (CNS) or Schwann cell (PNS)

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3
Q

What is located in the internode, paranode and node?

A

Intenode - K+ chanels
Paranode - CASPR and Coactin (where myelin adheres)
Node - Neurofascin p186 and Ankyrin G

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4
Q

When comparing the mRNA of schwann/oligodendrocytes from pre and post myelination what was found?

A

2 different types of Neurofascin (NF)
NF155 - found in glial cells
NF186 - found in neuronal cells

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5
Q

What was found when NF155 was immunolabelled?

A
  • Found in cell bodies of oligodendrocytes
  • Concentrated at the paranode
  • co-localised with CASPR at neuron-glial junction
  • Responsible for myelin adhesion to neuron
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6
Q

How is NF155 proposed to adhere myelin to neurons?

A
  • Anchored in the glial paranodal loop

- Binds with CASPR and contactin which are anchored in the axon membrane

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7
Q

What method has shown the binding of neurodascin?

A
  • Use yeast two hybrid with Gal4-UAS system expressing cytplasmic tail of neurofascin (bait)
  • optic and sciatic nerve yeast-two libraries (prey)
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8
Q

What does the cytoplasmic tail of neruofascin bind to?

A
  • Ankyrin isoforms
  • Ezrin
  • A novel protein (willin)
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9
Q

What is ezrin?

A
  • Part of 4.1 superfamily
  • Contains FERM domain which can bind phospholipids and proteins, coil-coil and tail
  • Can link transmembrane proteins to the cytoskeleton (actin)
  • Controlled by signalling pathways
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10
Q

Where do neurofascin and ezrin co-localise?

A

At interdigitating Schwann cells

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11
Q

What are the properties of Willin?

A
  • Contains FERM domain
  • Binds phospholipids
  • Seen in cell-cell interactions
  • Stimulated by growth factors
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12
Q

What are the roles of merlin and expanded?

A
  • Have a role in cell adhesion and structure and control activation of the hippo complex
  • Regulate cell growth and proliferation
  • When knocked down cause uncontrolled proliferation
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13
Q

What is the hippo pathway?

A
  • Kinase cascade first identified in Drosophila
  • Mutants have wing hyperplasia and eye defects
  • Components have different names but process is conserved in humans
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14
Q

What are the steps of the hippo pathway?

A
  1. Receptor ‘fat’ activates merlin/expanded
  2. This phosphorylates Hpo/Sav which phosphorylates wts/mats
  3. Phosphorylation of yorki transcription factor then stays in cytoplasm and signals apoptosis, if not enters nucleus and regulates survival
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15
Q

How is the hippo pathway linked to cancer?

A
  1. YAP (human yorki homolog) is an oncoprotein

2. Has a role in organ size homeostasis which is inhibited by hippo pathway

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16
Q

What is the TRex method?

A
  • Express TetR (tetracycline repressor) in cells, these bond to TetO2 (operator) sequences repressing transcription of gene of interest
  • Addition of tetracycline binds to TeR allowing transcription of gene of interest
17
Q

How have Willin and the hippo pathway been linked?

A

Using TRex method (Willin is very toxic)

  • Upon aplication of Tet there is activation of hippo components
  • Decrease in ammounts of nuclear YAP
18
Q

Does willin behave similarly to expanded?

A
  • When expressed in drosophila goes to the same place

- However slightly different effect on wing structure, but not suprising as it is a smaller protein

19
Q

What are MCF10A cells?

A
  • Breast cancer line used as they show signalling components of epithelial mesenchymal transition
20
Q

What is epithelial mesenchymal transition?

A
  • Epithelial cells are usually stationary, show apical-basal polarity, tight junction and expression of cell-cell adhesions (cadherin)
  • When undergo EMT loose polarity and adhesion, gain migratory and invasive properties to become mesenchymal stem cells (metastatic cancers)
  • Caused by YAP expression
21
Q

What is the role of Willin in MCF10A cells?

A
  • Willin knockout leads to loss of epithelial markers (E-cadherin and occludin) and gain of mesenchymal markers (N-cadherin and vimentin)
  • However dependent on YAP phosphorylation (effect removed if YAP phosphorylation site is removed)
  • Only willin FERM domain needed to reduce YAP induced genes

BL3303 (15 decks)

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