SignalTransduction

Pathways

Signaling Pathways ControlMany Cellular Processes

Cell CycleCell PolarityCell Shape

Development ….. and DiseaseDevelopment ….. and Disease

Signals

Stop!

Go Faster?

Go.

……….Pathway OFF

….…Basal

……..Pathway ON

Stimulus

Receptor

PAK/G-protein

MAP KinaseCascade

Txn Factors

Generic MAP Kinase Pathway

Cell BiologyGFP Collection

GenomicsDNA Microarrays

Yeast GeneticsGenomic Screens

BiochemistryTAP Tag Collection

Genetics and High-throughput Technologies

Budding Yeast is a Model Organism

Filamentous Growth (FG) in Yeast

FG is required for fungal FG is required for fungal pathogenesis.pathogenesis.

Saccharomyces cerevisiae

PRP

Ste2/3/4/18

Ste20/Cdc42

Ste11Ste7Fus3

Ste12

Ste5

FGP

Sho1 + ?

Ste20/Cdc42

Ste11Ste7Kss1

Ste12/Tec1

HOGP

Sho1

Ste20/Cdc42

Ste11Pbs2Hog1

Hot1

FGP

?

Ste20/Cdc42

Ste11Ste7Kss1

Ste12/Tec1

MAPK Pathways Share Factors

Mutants defective for agar-invasion (FG)

Genome-Wide Genetic Screen

UP

DOWN

NO CHANGE

DNA Microarray Analysis

Comparison of Genomic Data

MSB2 was identified in both screens.

Scaffold at the Head of the FG Pathway

Msb2

Cdc42 Sho1

Mucin Domain

Role of Mucin Domain in Signaling

Mucin domain is inhibitory.Mucin domain is inhibitory.

MSB2 Δmucin

1220swt wt

50

10

40

30

20

FU

S1-l

acZ

N…(698)S Q V S D T SV S Y T T S S S S V S Q V S D T PV S Y T T S S S S V S Q V S D T PV S Y T T S S S S V S Q V S D T PV S Y T T S S S S V S Q V S D T PV S Y T T S S S S V S Q V S D T SV P S T S S S S S V S Q V S D T PV P S T S S S S S V S Q (818) …C

Go, Go, Go!

Hyperactivity

Questions about the FG pathway

What is Msb2 sensing?

How are general factors specified by Msb2?

In what other ways is specificity achieved?

Stop!

Negative Regulation

FGP

Sho1 + Msb2

Ste20/Cdc42

Ste11Ste7Kss1

Ste12/Tec1

HOGP

Sho1

Ste20/Cdc42

Ste11Pbs2Hog1

Hot1Rck2

Novel Negative Regulator: a Calmodulin Kinase

Questions about Rck2

Where does the kinase act?

How does P~lation by Rck2 cause pathway shut-off?

Is the response specific?

Rotate in Paul Cullen’s Lab?

Sure!

Training in Paul Cullen’s Lab

Question-based approach

Learn many different techniques (Genomics)

Working with yeast is Fun!