Next lecture InductionSignaling Requirements of inducer and responder

Next lecture: Induction/Signaling • Requirements of inducer and responder cells • Cascades of inductive events are involved in forming organs • Examples of the kinds of cell communication • Important signaling pathways in development – Notch, TGFb, SHH, Wnt, FGF

“Official glossary” from Wolpert • Induction: The process whereby one group of cells signals to another group of cells in the embryo and so affects their development • Competence: The ability of a tissue to respond to an inducing signal. Embryonic tissues remain competent for a limited time (Can depend on prior inductive events)

Competence factors • Not all tissues are able to respond to the same signals. Competence is actively acquired – Receptors, signaling molecule – Transcription factors (chromatin state) • Competence factors are specifically required in the responding tissue, not the inducer – Demonstrated in recombination experiments

Recombination experiments: First name the players…(fig 6. 1 from Gilbert)

Then play… Fig 6. 3 Gilbert

Cascades of inductive events form the organ: Reciprocal induction Of Gilbert

Two kinds of induction (plus one) • Instructive interaction: A signal or factor which tells the developing cell what it is • Permissive interaction: A signal or factor that allows the cell to become what it is • Selective interaction. A factor allows the cell to be “selected” into one lineage or another after a stochastic (random) change

For example, CD 4 vs. CD 8 T cells Selection vs. Instruction TCR-I CD 8 TCR-II CD 4

Juxtacrine interactions • Involve cell surface receptors on inducer and responding cells (no soluble factors) • Cell death/apoptosis pathways – Fas (CD 95)/Fas. L • Notch/delta pathway – Involved in many binary cell fate decisions – Examples in flies, worms and mice

Cell death pathways Figure 6. 27 of Gilbert

Notch is involved in a wide array of binary cell fate decisions • C. elegans: Ventral uterine vs. Anchor cell and vulval development • Drosophila: Neural vs. Epidermal cells • Mouse: Embryonic lethal, Demonstrated affects in the immune system and others • Human: Notch deficiencies cause birth defects

Notch/delta pathway(fig 6. 29 of Gilbert) This model is simplistic little evidence for nuclear localization of Notch in developing organisms

Players in the Notch pathway • Receptor family: Notch/lin-12, glp-1 • Ligands: (DSL) Delta, Serrate, Lag 2, Jagged • Processing: ADAMs (Kuz, TACE) and Secretases (Sel-12/presenilins) • Downstream effectors: CBF 1, Su(Hairless), Lag 1 (Collectively called CSL)

Proteolytic processing of Notch is complicated Figure 1 from Weinmaster (2000)Curr. Opin. Genet. Dev. 10: 363 -369

Presenilins • Multipass transmembrane proteins • Mutated in inherited, early onset Alzheimer's disease • Involved in cleavage of amyloid precursor protein (APP) • Evidence that they are proteases or cofactors of a protease

Evidence linking presenilins with Notch signaling • Homology between PS 1 and Sel-12 of C. elegans (sel-12 facilitates Notch signaling) • PS and Notch deficient animals have similar phenotypes (Drosophila and mice) • PS is required for access of Notch to the nucleus and, thus, Notch signaling • Defects can be rescued by providing exogenous PS

Current view of Notch signaling Figure 2 from Weinmaster (2000) Curr Opin Genet. Dev. 10: 363 -369

Notch involvement in cell fate • Examples of Notch involvement in inductive interactions (signaling between non-equivalent cells) • Lateral specification (occurs in a population of equivalent cells) – Involves the amplification of a stochastic small difference between equivalent cells

Lateral specification in C. elegans From Greenwald (1998) Genes. Dev. 12: 1751 -62

Evidence for feedback mechanism in lateral specification • Constitutive active mutants of lin 12 have no AC • Mutants eliminating lin 12 activity have 2 AC • Mixing experiments between two types above – Lin 12 - cells always became AC – Lin 12 WT cells always became VU • Behavior different from WT in non-mosaic situation where these cells have equal chance to become AC or VU

One way to generate a bias in Notch activity-C. elegans vulval development From Greenwald (1998) Genes. Dev. 12: 1751 -62 fate adoption is 2 o-1 o-2 o

Gilbert Chapter 6 errors • Lag 2 is not secreted but transmembrane. This is the ligand for lin 12/Notch (p. 169) • Figure 6. 40 on T-lymphocyte signaling: The pathways leading from the receptors are not correct. (Notice, there is no reference) • Figure 6. 19: Details appear to be in dispute

Paracrine factors and interactions • • • Transforming Growth Factor (TGF)-b Sonic Hedgehog Wnt Fibroblast Growth Factor (FGF) Retinoic Acid (RA)

TGF-b • Over 30 members of the TGF-b family • Subfamilies, TGFb, Activin, BMP, Vg 1 • Processed proteolytically with the C-terminal region conatining the mature peptide • TGFb peptides can homo- or hetero-dimerize • TGFb signal transduction involves multiple receptors which activate cytoplasmic “Smads”

Smad pathway (fig 6. 20) p. 159

Division of labor among Smads Courtesy of J. F. Doody (J. Massague lab @MSKCC) Smad 6 and 7 are inhibitory proteins induced by antagonists of the TGFb signaling pathway, such as g-interferon (Smad 7)

The Smad family From Piek, et. al. (1999) FASE�B J. 13: 2105 -2124

A more complete TGFb pathway From Piek, et. al. (1999) FASE�B J. 13: 2105 -2124

Evidence for non-redundant TGFb family involvement in development (due to expression patterns) • TGF-b 1 knockout mice have defects in blood and vasculature (many die prenatally) • TGF-b 2 knockout mice have multiple organ malformation (perinatal death) • TGF-b 3 knockout mice die shortly after birth due to defects in pulmonary development • Information from Piek, et. al. (1999) FASEB J.

Next lecture: Induction/Signaling • Requirements of inducer and responder cells • Cascades of inductive events are involved in forming organs • Examples of the kinds of cell communication • Important signaling pathways in development – Notch, TGFb, SHH, Wnt, FGF