adventures in...
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Adventures in Multicellularity
The social amoeba (a.k.a. slime molds)
Dictyostelium discoideum
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Dictyostelium discoideum
ÿ the most studied of the social amoebae / cellular slime molds
ÿ predatory soil amoeba that feeds on a variety of microorganisms anddecaying matter
(in the lab exist primarily on a diet of E. coli although strains have been selected that grow on complex media alone).
ÿ asexually reproducing amoebae (unicellular) under conditions when food isabundant
ÿ under starvation conditions a complex developmental cycle is initiated – anadventure in multicellularity.
ÿ they have features of both plants and animals• cellulose and development of spores (plant-like)• cell movements involved in morphogenesis (animal-like)
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Embryogenesis Dictyostelium Development
Aggregation of manyidentical cells
Successive divisionsof large precursorcell (zygote)
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The Natural History of Dictyostelium discoideum
Vegetative Cycle:
Amoeba lives solitary existencefeeding on bacteria
Reproduce by binary fission (i.e.divide in half)
Continues as long as food isavailable
Feeding is by phagocytosis – essentiallyengulfing their prey with membrane.
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
Chemotaxis and Aggregation:
When the cells begin to starve, theybegin a developmental cycle
They produce and release cAMPthat acts as a chemoattractant andalso as a signaling molecule
The cells comes together (streaming)to form a multicellular tippedaggregate .
This elaborates into a finger orstanding slug
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
The standing slug has two fates – itcan topple over and become amigrating slug or it can proceed todevelop into a fruiting body. Theelaboration of the fruiting bodybegins with culmination
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
Scanning electron micrographs ofthe different stages of development
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
1.
2.
3.
4.
1. Chemotaxis to signals producedcoordinately by the cells(primitive morphogens)
2. Aggregation and cell adhesion(primitive patterning andmulticellularity)
3. Differentiation and patterning inthe migrating slug andfruiting body (primitivepolarity)
4. Competition to be spores(cheating)
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
Enclosed in a sheath (muco-polysaccharide and cellulose)
The tip is a specialized group ofcells that control development(remove it and development stops until anew tip is formed)
Under the control of the tip theaggregate elongates and forms thestanding slug.
RH Kessin (2001)
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The Natural History of Dictyostelium discoideum
The slug typically contains about100,000 cells and behaves as asingle entity. It is capable of bothphototaxis and thermotaxis.
RH Kessin (2001)
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Dictyostelium aggregates have many properties of an embryo :
• They are essentially a multicellular entity enclosed within a sheath
• They have polarity
• They have exquisite proportioning (1/5 prespore ; 4/5 prestalk)
• They have an organizing center (tip)
• morphogenesis involves cell movements
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The essential steps in cell motility
RH Kessin (2001)
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Three dimensional reconstruction of a D. discoideumamoebae moving towards a cAMP gradient.
cAMP source
cAMP
Wessel and Soll, Univ. of Iowa
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Chemotaxis of single cell towards a point source of cAMP.
The tip of the capillary tube is the source of cAMP. See howquickly the cell responds to changes in concentration. Note thenumber of pseudopodia protrusions in the amoeba.
Time hr:min:sec:
G. Gerisch, Max Planck Institute, GDR
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D. discoideum amoebae chemotaxing toward cAMP
Cells are expressing a GFP-coronin fusion protein: coronin is acytoskeletal protein that is localized in the extended pseudopods.
G. Gerisch, Max Planck Institute, GDR
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Starvation and cAMP signaling
During starvation, cells develop the ability tosynthesize, detect and degrade cAMP.
When one cell releases a pulse of cAMP (1),neighboring cells detect it and move up thegradient towards the source (2) .
After a delay of about one minute these cellsrelease cAMP of there own (2) and processgets repeated about every six minutes (3).
This process results in waves of migration (4).
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2
3
4
RH Kessin (2001)
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Star logo simulation
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Aggregation patterns
The patterns of aggregation can beseen by dark-field microscopybecause moving and stationary cellsreflect light differently – movingcells appear lighter.
Note the waves of cAMP tend to propagatefrom a central point (‘organizing center’)
RH Kessin (2001)
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Dark waves of D. discoideum cells on caffeine agar. Movieconstructed from 36 seconds intervals.(F. Siegert and C.J. Weijer J.Cell Sci. 93:325-335 (1989))
Aggregation Patterns
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Core of a Spiral Wave in Aggregating D. discoideum cells
10 sec between images
Siegert and Weijer, J. Cell Sci. 93:325-335.
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Siegert and Weijer, Curr Biol 1995
Rotation of cells within the mound.
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The classical studies of Raper (1940’s)
RH Kessin (2001)
Prestalk and prespore cells do not mix throughout development.
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The classical studies of Raper (1940’s)
RH Kessin (2001)
The tip controls events in the slug.
Apical fractions from foreign slugs (B-D) were grafted onto anormal slug. Four independent slugs emerge after ~45 min.
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JT Bonner (2000)
Positional Sorting of Prestalk and Prespore cells within the slug
Prestalk cells were grafted onto theanterior end of a slug.
These cells eventually migrate tothe proper position at the posteriorend of the slug.
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JT Bonner (2000)
There is little mixing of prestalk and prespore cells in the slug.
The prestalk cells at the tip move morerapidly in the slug than the pre-spore cells.
Prestalk Prespore
This study used an ingenious method developed by Bonner to generate “twodimensional” slugs. They could contain as few as 50 cells (typical slugs have >100,000 cells) but the correct proportioning of prestalk and prespore cells wasalways maintained.
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Wolpert, 1998
Movement of Cell Types During Culmination
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Weijer, Univ. of Dundee
Note that cells move more rapidly in the tip (prestalkcells) than in the body of the slug (prespore cells). Notealso that the tip is further partitioned into Prestalk a ndPrestalk O cells.
Movement of Prestalk Cells in the Slug
Two types of prestalk cells:1) Green (GFP labeled) Prestalk A cells2) Red (red-shifted GFP labeled) Prestalk O cells
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Weijer, Univ. of Dundee
Movement of Prestalk Cells During Culmination
Two types of prestalk cells:1) Green (GFP labeled) Prestalk A cells2) Red (red-shifted GFP labeled) Prestalk O cells
Watch closely for the movement of prestalk cellsdown the core of the culminating slug.
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Mound formation can be described by two assumptions: 1.) by considering thepopulation of cells as an excitable medium and 2.) by considering cell movementas the flow of a compressible liquid. The cell movement (fluid flow) is controlledby chemotactic forces dependent on increasing concentrations of the cAMP. Themodel allows to describe the whole early development from isolated single cellsinto bifurcating aggregation streams to the formation of a 3-dimensional,hemispherical mound, (see Vasiev B., Siegert F. & C. J.Weijer (1997), J.Theor.Biol. 184, 441-450
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A three dimensional hybrid cellular automata/partial differentialequation model capable of describing the morphogenesis ofDictyostelium discoideum “from single cells to crawling slug".Using simple local interactions we can achieve the morphogenesiswith only three processes: production of and chemotaxis to cAMPand cellular adhesion. The interplay of these processes causes theamoebae to spatially self-organize leading to the complex behaviourof stream and mound formation, cell sorting and slug migration allwithout any change of parameters. Our model describes thesubstrate for morphogenesis on which higher cell functions canoperate.
Savill and Hogeweg (1997) J. Theor. Biol. 184:0229-235.
http://www-binf.bio.uu.nl/~njs/dicy.html
P. Hogeweg et al Model of Development (Univ. of Utrecht)
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Streaming and Mound Formation
Hogeweg Model (cont’d)
• Each amoeba is represented by connected automata in CA system (notjust as a single cell)
• Cell-cell adhesion an chemotaxis are represented as simple CA rules
• Amoeba are not rigidly defined by the number of cells but can fluctuateslightly allowing flexible interactions and movements past one another
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cAMP waves
Slug development
Hogeweg Model (cont’d)
Yellow PrestalkGreen Prespore
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Siegert and Weijer, J. Cell Sci. 93:325-335.
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General References:
First Signals : Evolution of Multicellular Development.J.T. Bonner (2000)Princeton University Press
Dictyostelium: Evolution, Cell Biology and the Development of Multicellularity.Richard H. Kessin (2001)Cambridge University Press
Web sites containing numerous links to research sites, resources and databases:
http://dicty.cmb.nmu.edu/dicty/dicty.html
http://dictybase.org/dicty.html