Oligogalacturonide (OG) Signalling in Plant Innate

Immunity

23 March 2010

Background

• Activation of plant innate immunity– Specific detection of pathogen-associated

molecular patterns (PAMPs) – Host encoded pattern-recognition receptors

(PRRs) • Perception of PAMPs

– signaling cascades• activation of plant defenses and enhancement of

basal immunity towards potential pathogens

Background• Recognition of modified-self including damage-associated

molecular patterns (DAMPs)

• Plant cell-wall fragments– Degrading enzymes secreted by

the pathogen during the infection process

– Recognised as DAMPs

• None of the hypothetical DAMP receptors have yet been identified– Signaling processes not properly

characterized.

Background• Pectin

– Central component in plant cell walls

• Release of oligogalacturonide (OG) fragments from plant cell walls

• Many plant pathogens produce pectin degrading enzymes as crucial virulence factors

• Recognition as DAMP• Activation of immune responses

Previous work in the group

• Activation tagging - Karen

• 35,000 BASTA resistant plants selected from 1 million seeds• 1500 pools of 24 plants collected• Germinated on Whatman paper with water• Infiltrated with OGs• Selection for oligogalacturonide-tolerant seedlings

• OGs of DP 2-6(8) : 300 putants

• Reselection of putants (2x) : 30 ”mutants”

BAR4 x 35S enh

Repeating Karen´s results

• On 6 and 12 well plates with MQ water• Problems:

– Seedling germination and growth rate• Several lines very slow• Start to die before they are “ready”

7 days old 9 days old

• Solutions– New “fresher” seeds– Grow in MS media from the start?– One line per plate?

• Infiltrate when “standard” size?

New screen• Previous test to germinate on plates then move to liquid

medium containing OGs.

• Try to grow in 96-well plates.• Germinate in liquid ½ MS medium

– Low amount of liquid so seed can grow ”out” of liquid– After germination: add OGs– Selection for tolerant seedlings

• Problems– Drying out at the

edges of the plates

– Time consuming to transfer seeds one by one

• Seed loader?– Insufficient

oxygen?

New screen

8 days old – Grown in different volumes of media (μl indicated on the right side)