Membrane Fusion

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Membrane Fusion
Chapter 8 of Yeagle. Majority of work is taken
from papers by B. Lentz at UNCCH (Biochemistry)

• Membrane fusion is ubiquitous and critical
  process in biology
• Cellular trafficking and compartmentalization
• Import of food
• Export of waste
• Intracellular communication
• Sexual reproduction
• Cell division
• Viral fusion

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Protein Mediated
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Fission/fusion
tubes
Lipid segregation
Lipid mixing
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Mechanisms of Membrane Fusion Fusion Pore is
made of (A) protein and lipid, (B) lipid only
(B) Is the accepted pathway, this does not imply
proteins are NOT involved. Proteins are involved,
but they do not line the fusion Pore
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Detailed molecular mechanism is still unclear for
fusion. Secretory Vesicles and Viral Fusion

• Today Focus just on the lipids and bilayer
  fusion without the aid of proteins.
• Research of the last 15 years has shown a close
  association between membrane fusion and inverted
  phase formation.
• Both TH and fusion can be affected by the
  physical properties of the lipids and the system
• Temp
• pH
• Lipid composition, lipids of negative or positive
  spontaneous curvature
• Cation binding to lipids

Fusion Mechanisms are believed to proceed though
intermediate structures that are also observed in
the La to HII phase transition
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Stalk Hypothesis
Relationship between Fusion and Inverted Phase
Formation
TMC trans monolayer contact ETMC extended
trans monolayer contact
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Fusion is a necessary step in the lamellar to
inverted phase transition
So, if this is true, the physical properties that
affected the phase transition should also affect
fusion similarly
What are the 2 competing forces?
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• Two processes occur during fusion
• Lipid mixing of different leaflets
• Content Mixing

Content Mixing is true Fusion
1997 model

• If Stalks form, lipids can mix.
• Contents only mix upon FP (Fusion Pore) formation

2 competing forces curvature energy, Interstitial
chain packing
Spontaneous curvature of a mixture of lipids is
the mole-fraction weighted average of the
spontaneous curvatures
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Lipid Effects.
Negative Spontaneous Curvature PE, anything
that lowers TH also facilitates fusion
Positive Spontaneous Curvature LPC, anything
that raises TH also inhibits fusion
Phospholipase C can induce membrane fusion
because its in vivo reaction is to produce DAG.
DAG has small head group compared to chain,
hence, negative spontaneous curvature. Conversely
, production of LPC, lysophosphatidylcholine will
inhibit fusion. Additionally, sphingomylenase
action of sphingomyelin to produce ceremide can
also affect fusion. Ceremide seems to induce
leakage instead of fusion unless cholesterol is
present. Arachidonic acid will also promote
fusion
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All of this work is done with PEG induced fusion

• PEG will force lipid bilayers together in close
  proximity to induce fusion. Factors that can be
  affected to induce fusion
• Bilayer dehydration
• Imperfect lipid packing
• Local alterations in bilayer curvature
• Outer leaflet packing defects
• Elastic free energy
• Changes in membrane fluidity
• Locally induced non-bilayer phases

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Assays to monitor lipid mixing versus content
mixing Fluorescence
HTPS at pH 8.0 (8-hydroxy-pyrene-1,3,6-trisulfonic
acid) Make other vessicle with pH 5.0, A
decrease in fluorescence of HTPS indicates
movement of protons, hence a small pore formed.
Tb3 - DPA fluorescence increases upon
interaction with DPA. Here an increase indicates
content mixing.
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