1

UMB/UMN Workshop Report

from the

Cell

Wall Group (CWG)

Kate VandenBosch, Jerry D. Cohen, Steve Gantt, UMN

Carl Gunnar Fossdal, Norw. Inst. of

Forest & Landscape 

Odd Arne Rognli, Trine Hvoslef‐Eide, UMB

Abscission 2

Overview

of

the

CWG presentationGoals for UMB‐UMN Cell

Wall Group (CWG) 

•

Inventory

CW‐related

projects

of

group

members

1.

Kate VandenBosch

et al.2.

Simo Sarkanen

and Steve Gantt3.

Jerry Cohen4.

Trine Hvoslef‐Eide5.

Odd Arne Rognli6.

UMB’s

Cell

Wall Group•

Identify

possible

other

collaborators

•

Define

possible

shared

interests

•

Next

steps

–

Sabbatical

projects

for Trine and Odd Arne–

Define

what

could

be done

towards

a publication–

FY10 grant proposal

planning–

Develop

long

term collaboration

plan 

3

Plant Cell Walls

Primary cell wall

Secondary cell wall

Ferulate-lignincrossbinding

1.  Kate VandenBosch  Systems Biology of Cell Wall Biogenesis and Stem 

Development in Medicago

Participants:U OF MN PLANT BIOLOGYJane GlazebrookFumi

KatagiriMesfin

Tesfaye

Goals:Identify regulatory components controling

cell wall compositionunderstand co-ordinate regulation of cell wall and stem development

Publications: Tesfaye

et al. 2009. Bioenergy

Research; 1 in press + 1 in prep

Funding: NSF PGRP application planned for 1/2010; NSF BREAD and NIFA possible

USDA-ARSHans JungDebby SamacJoAnn

LambJohn GronwaldCarroll VanceSam Yang

Willow/Salix

Alder/AlnusHazelnut/CorylusQuercus/Oak

Angiosperm Phylogenyhttp://www.mobot.org/MOBOT/research/APweb/welcome.html

Rationale•Forage crop improvement•Digestibility•Biofuel

feedstock modification•Translation to

related foragecrops andwoody plants

Tools & Approaches

Elongation zone (Primary Growth)

Post-elongation(secondary growth)

Red stain = lignin

Evaluation of diverse genotypes• Association genetics• QTL analysis• SNPs

Transcriptional profiling•Expression polymorphisms •eQTLs•Genes co-regulated with markers of lignocellulose

deposition in 2ry walls

Reverse genetics

Mer

iste

mat

icro

ot

Uni

nocu

late

dro

otN

onm

eris

tem

atic

root

Nod

ule

4 dp

iR

oot

0 dp

i

Flow

er

Nod

ule

10 d

piN

odul

e 14

dpi

Nod

ule

28 d

piLe

af

Bud

Youn

g st

em in

tern

ode

Pod

Seed

10d

Seed

12d

Seed

16d

Seed

20d

Seed

24d

Seed

36d

Pet

iole

Ste

m

Old

ste

m in

tern

ode

Evaluated public Medicago

Affymetrix

data for genes co-expressed with:

•CESA•COBL4

Co-expressed genes included :

•Ferrulate

hydroxylase•Cytochrome

P450•β-galactosidase•Arabinogalactan

Proteins•Fasciclin-like AGPs•Receptor-like kinases•Transcriptional regulators in AP2, MYB, NAC & GRAS families

Genes co‐expressed with markers of  secondary cell wall synthesis

2:  Simo

Sarkanen

&  Steve Gantt Reducing Recalcitrance of Next Generation 

Lignocellulosic

FeedstocksParticipants: 

(SS, SG, and Mark Distefanon

(UMN)

Funding: Pending application with DOE Advanced  Projects Agency –

Energy (DOE ARPA‐E)

–

Transformational R&D emphasizes high‐risk concepts with 

potentially high‐payoff. •

Transformational energy technologies are those that have the 

potential to create new paradigms in how energy is produced, 

transmitted, used, and/or stored. 

–

Preproposal

was chosen as one of about 100 (out 3,500 

applications) to submit full proposal

Recent work has shown that not all  monolignols

are randomly polymerized

•

β‐O‐4 ether  linkages are most 

easily digested

•

High proportion of  these linkages are 

found in spruce

•

We propose that  proline‐rich proteins 

are involved in  directing their  formation

G. Gellerstedt, 2007, http://rfparois.free.fr/LIG2G/Seminaire%20LIG2G‐WEB‐vs‐tout‐public.htm;

Hypothesis and Experimental  Approaches 

•

Structure of lignins are encoded by cell wall proline‐rich proteins 

(PRP)

•

Supporting evidence:–

low concentrations of polymeric lignin in open solution promote selective 

formation of high molecular weight species during the dehydrogenative 

polymerization of coniferyl alcohol to lignin‐like macromolecules

–

PRP epitopes have been correlated with lignin both spatially and

temporally in developing cell walls of the maize coleoptile and in 

secondary walls of differentiating protoxylem elements in the soybean 

hypocotyl

•

Experimental approach:–

Knock down loblolly pine PRP gene expression by RNA interference

–

Examine the resulting lignin structure by 2‐D HSQC NMR spectroscopy

4.  Jerry Cohen

Major Interests:

Auxin

biosynthesis

and metabolism;  protein turnover

Potential

Cell

Wall‐Related

Interests:

•

Developmental

transition

from juvenile to mature tissues

•

Stem development

in shade

avoidance

•

Roles

of

plant hormones

in cell

wall

development

•

Analytical

tools

for evaluating

cell

walls

Participants:

UMB (THE), Bioforsk

(Jihong

Clarke), UMN  (Jerry D. Cohen)

Rationale: Norwegian

poinsettia

growers

defined abscission

as one

of

3 priority

problems

Goals: prevent

precocious

abscission

Tools: RNA in situ

hybridization, RNAi, IAA transport  studies, gene

characterization

Publications: 

Lee et al., The Plant Journal, June 2008;   Ayeh

et al, BMC Plant Biology, June 2009

Funding: Pending

in NFR

13

Induction

of

abscission in poinsettia

•

Decapitating

the

buds to intiate

abscission

•

Initiated

in 7 days

•

98‐100 % induction

if

done

accurately

Abscission 14

Immunolabelling

analysis in abscission  zones in poinsettia using anti pectic

antibodies

Lee et al, The Plant Journal, 2008

Abscission 15

Fourier‐Transform

InfraRed microspectroscopy

(FT‐IR)

D0 D2 D4 D6 D7

Lee et al, The Plant Journal, 2008

Lee et al, in prep

Abscission 16

Abscission‐less

mutants in pea

Ayeh 2008

17

Description of six riboprobes

from differentially  displayed clones from poinsettia flower abscission 

(Munster 2006; Ayeh

2008)Riboprobe BLAST search Accession

numberSpecies Orientation

in the pCR4TOPO

6a PSBX(photosys temIISU X)

NP 565335.1

A. thaliana Forward

25a No significant matches

- - Reverse

38b No significant matches

- - Forward

84 Putative pol protein

AAL66759 Zea mays Forward

90a No significant matches

- - Forward

136b Immunophilin NP196845 A. thaliana Forward

5.  Odd Arne Rognli

Participants:

UMB (OAR), Bioforsk

(Liv Ostrem, Jihong Clarke), Graminor

(A. Larsen), UMN (Kate 

VandenBosch)

Rationale: Improved

nutritive quality

of

forages is a key factor

in sustainable

livestock

farming. The same 

traits

also

determine

conversion

of

lignocellulose

to  biofuel

Goals: Genetic regulation

of

cell

walls 

Tools: sequencing, comparative

genetics, SNPs, field trials, phenotyping, feeding

experiments

Funding: NFR, industry

(Graminor)

19

Composition Stem/leaves

Forage with highnutritive value

Harvesting

Products

Grazing

Conservation

Ongoing project: Festulolium with improved forage quality and winter survival for Norwegian farming

20

Forage quality studies 

•

QTL mapping of NIRS trait predictions  (15 traits) in F. pratensis

–

MSc

thesis

•

Available NIRS predictions for ADL  (Acid detergent lignin) not good 

enough•

Calibrated FT‐IR models for prediction 

of ADL and KLASON

lignin –

The accuracies of the ADL and KLASON were 92% and 89% –

Small data set, model calibration are being expanded in collaboration with 

IBERS, Wales (Gordon Allison)

•

Good forages have:–

low lignin content–

low S/G ratio (syringyl/guaiacyl)–

little cross‐linking by ferulates

We have generated sequences and SNP information of genes involved in lignin biosynthesis from Festuca and Lolium:

4-Coumarate CoA ligase (4CL)Cinnamate-4 hydroxylase (C4H)Cinnamyl alcohol dehydrogenase

(CAD)Caffeoyl-CoA O-methyl transferase

(CCoAOMT)Cinnamoyl CoA-reductase (CCR)Caffeic acid O-methyltransferase

(COMT)

PhD student (China/Norway) starting Jan 2010

Transformation of Brachypodium to manipulate lignin biosynthesis

6.  Norwegian Cell Wall Group Network organizers

•

Vincent Eijsink, UMB

•

Øystein Johnsen, Skog og Landskap

•

Sigbjørn Lien, UMB

•

Svein Knutsen, Nofima

Mat

•

Ellen Merethe Magnus, Bioforsk

•

Odd‐Arne Olsen, UMB

•

Margareth Øverland, UMB

•

Birger Svihus, UMB22

Festulolium

prosjektmøte

21.04.0823

Festulolium

prosjektmøte

21.04.0824

Festulolium

prosjektmøte

21.04.0825

Festulolium

prosjektmøte

21.04.0826

Goals

for UMB‐UMN CWG  Short term

•

Define

collaborative

sabbatical

projects

for THE and OAR.   Possible

approaches

with

Medicago

materials:

–

Genetic studies•

Bioinformatics?•

QTL mapping

of

morphological

traits?•

RNAi

knockdown using

hairy

root

cultures?•

RNA in situ

hybridization

of

selected

gene

sequences?–

Cell

biology

approaches

to selected

phenotypes

•

Antibodies

to cw

components?•

Studies on

secondary

cell

wall

deposition

(lignin)?•

Hormone

studies?

•

1 publication

on

cell

wall

studies resulting

from  sabbatical

project(s)

•

Consult

on

grant proposal(s) under preparation

Goals for UMB‐UMN CWG  Long term

•

Continue

to identify

other

potential collaborators, their

interests

and expertise

•

Development

of

model

systems to study

CW

•

Development

of

analytical

methods

to  elucidate

on

CW assembly, composition

and 

degradation