Gravity Signaling

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Gravity Signaling

• Dr. Stanley Roux
• Thomas Bushart
• tbushart_at_mail.utexas.edu
• Bio Labs room 6 9

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Gravity

• A constant, (mostly) inescapable, and directional
  force
• Consistency and directionality make it useful an
  information source in addition to, or in lack of,
  other cues
• Orientation
• Gravitaxis (geotaxis)
• Differentiation
• Differential growth

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Sensing Response Animals

• Specialized cells to sense the movements of
  fluids (endolymph) and solids (otoliths,
  statocyst)
• Movement/pressure translated by nervous system
  into signals for balance and orientation

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End Results Responses of Plants and Single Cells

• Arabidopsis a multicellular weed
• Shoot (-) and root () growth
• Ceratopteris an aquatic fern
• Asymmetric cell division and downward rhizoid
  growth
• Euglena a photosynthetic flagellate
• (-) gravitaxis
• Chara a non-motile, multicellular algae
• Cytoplasmic streaming in internodal cells

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Reasoning Backwards

• Root curving
• Asymmetric cell divisions
• Directional movement of a single cell
• Asymmetric cytoplasmic streaming
• Growth, cell divisions, movement, and streaming
  all require signaling, therefore polarized
  responses would require polarized signaling

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The Great Unknown

• How is the directional force of gravity sensed by
  a single cell? (PE KE response)
• Statolith Model
• The internal downward movement of a dense
  particle
• Pressure Model
• The sagging of the cell due to buoyancy
  differences

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Starch-Statolith Theory

• Conceived from examining the cells required for
  gravity response
• Root columella cells are required for gravity
  perception in roots
• Ablate columella cells and root will still grow
• Gravistimulate the root (turn the plant on its
  side) and root curvature is impared
• Endodermal cells in shoots are similarly
  necessary
• Since not the responding cells, likely to be the
  sensing cells

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Columella Cells

• This special root cell type contains large,
  dense, membrane-bound starch grains called
  amyloplasts (present in shoot cells as well)
• Reorientation of the root can result in movement
  of these amyloplasts in the new downward direction

Chen et al. 120(2)343 Plant Phys
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Further Implications for Statoliths

• Mutants with reduced or absent starch production
  would have smaller or absent amyloplasts
• Less starch less response to gravity
• Microgravity experiments on Arabidopsis root
  curvature
• WT and starch mutants similar in micro g
• 1-g centrifuge WT reorient, starch mutants are
  impaired

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Pressure Theory

• Mass of the protoplast as a whole is affected by
  the gravity vector
• Chara internodal cells and Euglena
• Lack amyloplasts
• Manipulations of environment can affect
  cytoplasmic streaming response and directional
  movment

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Pressure and Density

• An elastic object more dense than its
  surroundings will sag such that there is
  compression forces on the bottom and tensile
  forces on the top
• A balloon filled with water will flatten out when
  placed on a table (water gt air)
• Place the water filled balloon in water and it
  will take the natural shape of the balloon
  (water water)
• Anything inside the water filled balloon will
  behave independently of the media surrounding the
  balloon (i.e. a marble would still sink to the
  bottom)
• Cells are like a balloon, organelles like marbles

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Change Environment

• Cytoplasm is naturally more dense than water (and
  of course air)
• Density of media density of cytoplasm
• Euglena remain motile, but loose directionality
• Chara streaming will become equal
• Density of media gt density of cytoplasm
• Euglena swim downward
• Chara stream in the opposite direction

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Further Evidence for Pressure Model

• Hydrostatic pressures applied externally to ends
  of horizontal Chara cells effects streaming rates
• Positive pressures streaming away from
  stimulated end
• Negative pressures streaming towards stimulated
  end
• A vertically oriented cell would experience
  different pressures on each end

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Problems Ambiguous Evidence Universality

• Some results could lend support to either theory
  depending on how you look at it
• Starch mutants still have some ability to respond
  to gravity (especially under high g forces)
• Laser ablation of columella cells doesnt
  completely remove gravity response
• Actual movement of a statolith may not be
  required
• Germinating rice placed in high density media
  still show some root curvature
• Results from one system may not be applicable to
  others
• Not all gravity responding cells contain
  statoliths
• Chara cells are MUCH larger than most cells (on
  the order of several centimeters long)

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Common Ground

• The force of gravity acts upon some element of
  the cell (statoliths or protoplast)
• The receptor element then acts upon something
  else within the cell to initiate a signal for
  asymmetric signaling
• In both theories this initial signaling event is
  likely associated with membranes, either directly
  or though cytoskeletal connections

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Necessary Components

• Cytoskeleton
• Membranes
• Proteins
• Calcium

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Cytoskeleton and Membranes

• Amyloplasts and other organelles are enmeshed
  within an actin network
• Amyloplasts may be restrained by ER elements or
  in contact with specialized ER
• Tension/Compression forces would act directly
  upon plasma membranes
• The Altered Response to Gravity (ARG1) gene
  encodes a putative Dna-J like protein

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Chara Extracellular Matrix Connection

• Degradation of certain cell wall elements
  abolishes gravity response
• RGDS peptides applied to the top end of cells
  inhibit gravity response (tension)
• RGDS implies integrins
• Integrins would involve membranes, ECM, and
  cytoskeleton

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Integrin signaling regulates proliferation differ
entiation Motility Survival
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Protein Requirement

• The Euglena photoreceptor is inhibited by UV-B
  light which in turn inhibits phototaxis. UV-B
  light can also impair gravitaxis
• Treating the ends of Chara cells with proteases
  or UV irradiation abolishes response
• A mutation in the Arabidopsis ARG1 protein alters
  gravity response

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Calcium

• Implicated in all model systems
• Euglena - Gd, vanadate, and a calcium ionophore
  all inhibit gravitaxis
• Chara external Ca, nifedipine, La effect
  streaming
• Arabidopsis Gd/La and calmodulin Ca-ATPase
  blockers inhibit gravity reponse, Ca gradient
  seen across gravistimulated root
• IP3 signaling as well?

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Hypothetical Signaling Story

• Gravity acts upon protoplasm
• Force acts through integrins
• Ion channels open in a polar fashion
• Subsequent responses

• Gravity acts on statolith
• Statolith in contact with ER activates ion
  channels, or 2 above
• Responses

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Ceratopteris Spores as a Model System

• Single cell both senses and responds to gravity
• Only slightly larger than a typical multicellular
  plant cell
• Earliest detectable response is a gravity
  directed calcium flux
• Known time frame for gravity directed polarity
  fixation
• Subsequent steps are visually apparent

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Spore Germination Is Induced by Light and
Polarity Development Is Directed by Gravity
Period of gravity fixation ( 4-18 h)
Rhizoid emergence (72h)
Downward nuclear migration (24 h)
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Calcium Movement
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ATP
ADP Pi
Use calcium as the starting point Calcium
channels Calcium pumps Calcium transmitters
Late gravity responses
Ca2 ions
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Summary

• Gravitational forces lead to a variety of polar
  responses
• Movement, growth, differentiation
• Mediated by cytoskeletal, membrane, and protein
  elements
• Actin, microtubules, PM ER, intergrins, ion
  channels and pumps
• Actual receptor(s) and subsequent signaling
  pathway(s) still being investigated

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Articles

• Chen R., Rosen, E., Masson P. Gravitropism in
  Higher Plants. Plant Physiology. (1999) 120
  343-350
• Staves, M. Cytoplasmic streaming and gravity
  sensing in Chara internodal cells. Planta (1997)
  203 S79-S84
• Chatterjee, A., Porterfield D., Smith, P., Roux,
  S. Gravity-directed clacium current in
  germinating spores of Ceratopteris richardii.
  Planta (2000) 210 607-610