Transcript Lecture 10

Module 632
Lecture 10
JCS
Unconventional
myosins and cell
biology
MODULE - 632
Lecture 8
Muscle Contraction
Lecture outcomes:
At the end of this lecture a student will be aware that :
1) there are many different types of motility within cells
2)
many movements require directed polymerisation of
the actin cytoskeleton as well as cell growth (axons)
3) most movements are driven by molecular motors – myosin
(actin -based) and kinesin/dyneins (microtubular based).
•
5) there is a large family of myosins in eukaryotes
•
6) myosins are important for cellular transport, and changes in
cell shape
•
7) myosins are important in how we hear
•
8) mutations in myosins can cause human genetic diseases –
deafness, cardiomyopathy, sk. muscle myopathy.
Cell motility is driven by different motors:
• Cytoskeleton polymerisation/depolymerisation
– Actin filaments
• Pseudopodia, lamellipodia, phagocytosis,
acrosomal process extension – listeria hijacks this
system, pollen tubes.
– Tubulin (forms microtubules)
• Molecular motors:
– Actin-based motors
• MYOSIN FAMILY
– Microtubule-based motors
• KINESIN family
• DYNEIN family.
Fibroblast
Fish keratocyte
ACTIN FILAMENT DYNAMICS
Minus end - pointed end
Plus end - barbed end
Acrosomal process of sea urchin sperm:
Many transport functions require
molecular motors:
e.g.
Actin-based Myosin family
MT-based Kinesin & Dynein families
Many cell motilities require several
different motors or motors + filament
dynamics!
Locomotion of cells:
 Actin filament polymerisation
 Actin filament “stress fibres”
 Focal Adhesions
  Myosin I
  Myosin II
Axon growth - Neuronal vesicle transport:
MYOSIN V
Other myosins
found here
include Myo
VI, Is,
dynein
kinesin
microtubule
The myosin family :
II
VII
XI
XI I
IV
VI
VI
I II
XI
I
VII
X
I
I
Most myosins have been identified using the “Walker” P- motif (J.E. Walker).
The sequence is GESGAGKT = Gly-Glu-Ser-Gly-Ala-Gly-Lys-Thr
Cellular functions of the myosin family
From Mermall et al. (1998)
Science 279, 527-533
Plant cells also exhibit vesicular transport:
(called cytoplasmic streaming)
But driven by myosin!
Especially the giant cells of the pond algae, Nitella and Chara
100mm
Up to
60mm.sec-1
Myosin motor polarity:
Direction determined by actin filament polarity:
All myosins, except myosin VI, move towards the plus (+) or
barbed end of actin.
Note:
The + end of actin is defined at the most rapidly polymerising end. It is also
called the barbed end because of the arrow-head pattern when decorated
with myosin S1 heads. The + end usually points towards the cell periphery
The other end is called the minus (–) or pointed end
- e.g. most cytoskeletal myosins walk away from the nucleus
- fits with the muscle thick filaments moving along the thin filaments towards
the Z-line (where the thin filament F-actin barbed ends are)
Velocity is determined by the myosin isotype.
Many unconventional myosins
are associated with hearing.
Myosin-I, II, III, VI, VIIa
Slides courtesy of Chris Batters (NIMR Mill Hill)
The Human Ear
Sound enters the ear canal and causes the tympanic membrane to vibrate. This
vibration is changed from a low force large movement to a high force small movement
by the three bones in the inner ear. The stapes uses this higher force to vibrate the oval
window, which separates the air filled outer ear to the fluid filled inner ear. The
vibration causes the fluid in the ear to accelerate around the cochlea deflecting arrays
of stereocilia. When they are deflected mechanosensitive ion channels open and action
potentials are created allowing us to hear (Very similar action in the vestibular organ,
allowing us to balance).
Stereocilia
Single hair cell –
note stereocilia
Stereocilia
Outer hair cells
Inner hair cells
Variety of myosins in
hair cells –
From website of Dr
Sutherland MacIver –
Univ. Edinburgh
Stereocilia
Scanning electron
micrograph
(x 2,500) hair
bundles in the
bullfrog saccule.
Transmission electron
micrograph (x 50,000)
of the tip of the
stereocilia and its
adjacent, taller
neighbour.
Hair Cell
Tip-Link
Jeffrey R Holt, PNAS 2000.
Myo1c
Physiological
significance
“Tip Link”
tensioning motor
Gillespie and Walker Nature, 2001. 413:194-202
The stereocilia are linked together by
tip links ‘Cadherin 23’ (May 2004,
Nature). The tip links allow the
coordinated opening of the ion
channels ‘very important so that the
action potential threshold is reached’.
*******************************************
The tension in the tip link is
maintained by an array of
unconventional myosin-I (myo1c)
D.Corey
Slow adaptation within the ear
Calcium marker showing the
entry of calcium into the
stereocilia upon deflection
An array of 20-100 myo1c molecules maintains the
mechanosensitive ion channel in a ‘just off’ position by
climbing up the stereocilia or slipping down.
Myosin mutations and disease:
Myosin II mutations: Human familial hypertrophic cardiomyopathy
(HCM) mutants (“Sudden death syndrome”). Human cardiac myosin
is encoded by two genes (V1 and V3 isoforms) which are expressed
at low levels in skeletal muscle.
HCM mutations occur in at least 9 different cardiac sarcomeric protein
genes including myosin heavy chain, tropomyosin, actin, MyBPC,
troponin genes.
Myosin VII mutations: Usher’s deaf/blind syndrome (myosin VII)
Myosin V mutations: Dilute mutation (in mice) mild phenotype =
altered coat colour : severe phenotype = convulsions and early death.
Familial hypertrophic cardiomyopathy – FHC
‘Sudden death syndrome’
Most mutations which cause
this are in the myosin heavy
chain – others occur in
troponins, TM, actin, MyBPC
etc.
Skeletal myopathies –
Nemaline myopathies
Nema =
rod/thread (Gk)
Mutations identified in:
 &  tropomyosin
troponin
nebulin
-skeletal actin
Nebulin mutants most
common – large gene –
not many sequenced.
Actin mutants (ACTA1)
>105 mutants. Most
dominant.
-tropomyosin mutant
1
Actin
(ACTA1)
gene mutants
2a
2x
CFTD –congenital fibre
type disproportion
Classical nemaline