biology 624 - developmental genetics
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Biology 624 - Developmental Genetics. Lecture #8 - Tube Formation I. Tubular Organs. Tube Formation is critical to forming: Lung* Kidney* Mammary gland Blood vessels* Fly trachea C. elegans excretory system (1 cell!). From Nelson, 2003. There are three types of tubes: Multicellular - PowerPoint PPT PresentationTRANSCRIPT
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Biology 624 - Developmental Genetics
Lecture #8 - Tube Formation I
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Tubular Organs
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Tube Formation is critical to forming:
1.Lung*2.Kidney*3.Mammary gland4.Blood vessels*5.Fly trachea6.C. elegans excretory system (1 cell!)
From Nelson, 2003
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There are three types of tubes:
1.Multicellular2.Unicellular, with autocellular
junction3.Unicellular, seamless
From Lubarsky and Krasnow, 2003
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Processes of Tubulogenesis
From Lubarsky and Krasnow, 2003
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Processes of Tubulogenesis
From Lubarsky and Krasnow, 2003
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Genetic Programs involved in Tube Formation
From Nelson, 2003
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MDCK Cells in Collagen Gels
- HGF +HGF
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MDCK in collagen starts with a polarized cyst
Gp135 = apical, redb-catenin = basolateral, green
From Pollack et al, 98; Zegers et al, 03
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Upon addition of HGF, some cellsform an extension outward on their
basolateral side
Gp135, apicalE-cadherin, basolateral
From Pollack et al, 98
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The extended cell divides, and cells in the extension express E-cadherin
but not gp135
E-cadherin, basolateral
Gp135, apical
From Pollack et al, 98
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From Pollack et al, 98From Pollack et al, 98
As the lumen begins to reform in theextension, gp135 is re-expressed
on the new apical surface
E-cadherin, basolateral
Gp135, apical
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From Pollack et al, 98
As lumen formation continues, cells re-establish basolateral expression of
E-cadherin
E-cadherin, basolateral
Gp135, apical
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Model derived from this study:
1.Stimulation of migration is the first step in tubulogenesis
2.Apical/basal polarity is transiently lost and then restored
3.Discontinuous lumens form in tubules
4.Cell-cell contacts are retained throughout the process
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Apical membrane biogenesis is importantFor lumen formation in MDCK cells
From Lubarsky and Krasnow, 03
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Tube formation requires and stabilizes apical-basolateral polarity
From Bryant and Mostov, 08
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From Bryant and Mostov, 08
Tube formation requires and stabilizes apical-basolateral polarity
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From Uv et al, 2003
Drosophila Tracheal System
•Simple structure
•Powerful genetics
•Easy observation
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Embryonic Tracheal Development
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From Uv et al, 2003
I
II
III
IV
Drosophila Trachea Has Four Types of Tubes
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Drosophila Trachea Has Four Types of Tubes
From Uv et al, 2003
4-5 µM 1 µM 0.5 µM
2-5 cells compose the lumen circumference
single tube-shaped cellswith AJ encircle the lumen
doughnut shaped cells with no AJ
protrusions from single cells without AJ
1 µM or 4-5 µM
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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Specification of Tracheal Cells
From Affolter and Shilo, 2000
Trh = trachealess (bHLH PAS transcription factor)Tgo=Tango (bHLH Pas transcription factor, binding partner for Trh)Vvl/Dfr = Ventral veinless-Drifter (POU domain transcription regulator)
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Invagination of Tracheal Placode
EGFR = epidermal growth factor receptorSpitz=EGFR ligandRho = Rhomboid (EGF pathway activator)
From Affolter and Shilo, 2000
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Model of Tracheal Cell Invagination
Trh, Vvl
Rho (EGF pathway)
Apical Actin Enrichment
Localized Apical Cell Constriction
Ordered Cell invagination
Brodu V and Casanova J 2006
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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DB = dorsal branch
DTa/p = dorsal trunk (anterior/posterior)
VB = visceral branch
SB = spiracular branch
LTa/p = lateral trunk (anterior/posterior)LTp is also called GB or ganglionic branch
Branches of the Drosophila Trachea
From Cabernard et al, 2004
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From Affolter M 2002
Determination of branch identity
EGF=Epidermal growth factor : GB and DTWg= wigless: DTDpp = Decapentaplegic (Transforming growth factor β-like): DB, LT
EGF αPS1 integrin
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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FGF Signaling Directs Primary Branch Outgrowth
From Cabernard et al, 2004
bnl/FGF = blueBtl/FGFR in Trachea cells = brown
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Primary Branching Requires Bnl/Btl Signaling
From Cabernard et al, 2004green = actin-GFP (WT)Red = cells lacking Btl/FGFr
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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Dpp Signaling is Required for Dorsal Branch Migration in Addition to FGF Signaling
Affolter M 2002
Dpp (ligand)Pnt (receptor)
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From Englund et al, 2002
Branch Migration Requires Slit/Robo Signaling
blue= tracheal lumenBrown =DSRF (GB marker)
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From Cabernard et al, 2004
Branch Elongation via Cell Rearrangements
Type I Type II Type IIType I
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AJ Remodeling during Intercalation Process
From Ribeiro C et al 2003
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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From Uv et al, 2003
Formation of Specialized Tracheal Cells
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Fusion Process
E-Cadherin Membrane
Cytoskeleton Lumen
Migration Contact Adhesion Invagination Lumen Formation
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Dysfusion is Expressed in Tracheal Fusion Cells
DT
DB
LT
btl-lacZ Dys
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dys Misexpression Causes Inhibition of Migration
DysMab 2A12
Wild type
btl-gal4; UAS-dys
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• Branch identity determination• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals in addtition to
FGF signaling DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching
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From Uv et al, 2003
I
II
III
IV
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VB that has ramified to form dozens of fine terminal branches on the gut
Larval Tracheal System
Ghabrial and Krasnow MA 2003
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Terminal branch expansion in response to Hypoxia induced branchless
Jarecki J, Johnson E and Krasnow MA 1999 Type IV tube
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Model for Patterning of Terminal Branching by Bnl
Jarecki J, Johnson E and Krasnow MA 1999
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Steps of Tracheal Tube Formation
• Tracheal sac formation Specification of tracheal cells (tracheal placodes) Invagination of the tracheal placode
• General branch outgrowth via FGF signaling• Branch-specific outgrowth is controlled by regional signals and branch
identity genes Branch identity dentermination DB migration requires Dpp signaling GB migration requires slit-robo signaling Tube elongation through cell intercalation
• Determination and differentiation of distinct cell types (fusion cell, terminal cell) Fusion process Terminal branching