Root Hairs

506 Pages·2012·9.73 MB·English

GENETIC NETWORKS of PLANT DEVELOPMENT
and NUTRIENT-TREATED ROOT HAIRS
by
Christophe Liseron-Monfils

A Thesis
presented to
The University of Guelph

In partial fulfilment of the requirements
for the degree of
Doctor of Philosophy
in
Plant Agriculture

Guelph, Ontario, Canada

© Christophe Liseron-Monfils, January, 2012
ABSTRACT

GENETIC NETWORKS of PLANT DEVELOPMENT
and NUTRIENT-TREATED ROOT HAIRS

Christophe V. Liseron-Monfils Advisor:
University of Guelph, 2012 Associate Professor Manish N. Raizada

Nitrogen and water limit global crop production. Root hairs comprise the major surface
area of plant roots for nitrogen and water absorption. Despite this, no studies have reported how
added nutrients or water alter global gene expression in root hairs in any plant species.
I demonstrate that within 3 h of added nitrate, >876 genes in maize root hairs switch up
or down. These genes overlapped with relevant maize QTLs. I also demonstrate that within 3 h
of adding water to drought-stressed roots, 1831 genes show altered expression in root hairs.
These results demonstrate that root hairs are rapid sensors of the soil environment.
Following root uptake of nitrate, it is assimilated into amino acids for transport to the
shoot; a process that involves families of genes. Understanding where and when these genes
switch on/off may facilitate improved use of nitrogen. Here, the expression of 65 nitrogen uptake
and assimilation probes was analyzed in 50 maize tissues, from germination to seed
development. The results demonstrate that subsets of nitrogen-related genes are selectively
expressed in specific organs at specific stages of development. This result identifies a higher
network level of control over nitrogen-related gene expression in maize.
An important clue to identify the master regulators responsible for such coordinated changes in
gene expression can come from identifying their promoter targets. No software program has been
adapted to serve this purpose in maize. I have created an online software tool for the maize
community, called Promzea that is able to retrieve DNA motifs over-represented in the
promoters of co-expressed genes, will be described. Using Promzea, over-represented motifs
were identified associated with co-expressed genes in root hairs, diverse maize organs and
different developmental phases. These results should aid in the design of synthetic promoters for
targeted maize gene expression. As such genetic transformation will involve improving in vitro
regeneration in maize, in my last study, I describe the transcriptome effects of two QTLs that
improve in vitro plant regeneration, using a model system
Acknowledgements
First I would like to express gratitude to my advisor Manish Raizada for giving me this
great opportunity. I would like to thank him for his support and his kindness. Thank you for
giving me direction and to have believed in me. Thank you for your great scientific vision that
has made me raise the bar in science considerably. You have been and will remain an example of
dedication and caring about others for me.
Thank you to my Advisory Committee composed of Lewis Lukens, Joseph Colasanti and
Daniel Ashlock for their guidance and helpful advice.
I am grateful to François Fauteux, Gregory Downs, Lewis Lukens and Martina Strömvik
for guiding me in the computer programming learning process, and for helping to solve my first
computer problems. I am also grateful to them to have allowed me to use their server during the
first years. I am also grateful to Paul McNicholas for his advice and for kindly giving me access
to his server during the Promzea testing. I would like to express gratitude to the internet Perl
community for their help during the production of Promzea and during the initial difficulties
involving microarray analysis.
I am also very grateful to Steven Rothstein and Mei Bi for their help, good advice and
support during these years. You have been examples for me during these years. This experience
would not have been the same without the exceptional sharing and support of my lab mates:
Steve, Mike, Blair, Sameh, Amélie, David, Hannan and Vijay. Their presence has enriched me
exceptionally for the rest of my life. Thank you also to the summer students who helped during
this project: Adrienne, Sarah, Bridget, and all the others for their enthusiasm and their great
work.

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I would like to thank my fiancée, Eve. I would never have enough words to describe her
support and love through these years. Thank you for encouraging me to continue with graduate
school when my only will was to give up on this part of my life. I am also very grateful to my
family for their unconditional support and love, my parents, my sisters and my brother. You
have taught me and encouraged me to never give up and never allow me to compromise on what
I want to become in life.

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Abbreviations
AddN: Added Nitrate media
AMT: ammonium transporter
BioP: BioProspector
BLAST: basic local alignment search tool
bp: base pair(s)
cM: CentiMorgan
DIMBOA: 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one
Gln: glutamine synthetase 1-encoding gene
GOGAT: glutamate synthase.
GS: glutamine synthetase
Ler: Landsberg Arabidopsis ecotype
MaintN: Maintenance Nitrate media
MAL: Maize Adult Leaf motif
MAR: Maize Adult Root motif
Mbp: Mega base pairs
MEME: Multiple Em for Motif Elicitation
miRNA: micro ribonucleic acid
MJL: Maize Juvenile Leaf motif
MJR: Maize Juvenile Root motif
MNR: Maize Nodal Root motif
MNR: Maize Nodal Root motif
MORSL: Maize Old Reproductive Stage Leaf motif
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MORSR: Maize Old Reproductive Stage Root motif.
MSR: Maize Seminal Root motif
MSR: Maize Seminal Root motif
MV1R: Maize V1-stage Root motif
MV2L: Maize V2-stage Leaf motif
MV2R: Maize V2-stage Root motif
MVeL: Maize Ve-stage Leaf motif
MVeR: Maize Ve-stage Root motif
MYLL1: Maize Young Lower Leaf motif
N: nitrogen
NAR2: nitrate transporter 2 complex accessory protein
NiR: nitrite reductase
NiRT: nitrite transporter
NIL: near isogenic line
NO: Nitric oxide
Nos: Nossen Arabidopsis ecotype
NR: nitrate reductase
NRE: nitrate responsive element
NRT: nitrate transporter
PLACE: plant cis-acting regulatory DNA elements database
qPCR: quantitative polymerase chain reaction
QTL: quantitative trait loci
RH: root hair
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RHM: root hair media
ROS: reactive oxygen species
RxDAP: reproductive stage, x days after pollination
Vx: vegetative stage

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List of Tables
Chapter 2
Table 2.1: Position weight matrix example.
Chapter 3
Table 3.1: Software programs used in Promzea.
Table 3.2: Combination of motif discovery programs based on measures of True Positive
and False Positive nucleotides.
Table 3.3: Effectiveness of combining different motif discovery programs based on
nucleotide sensitivity scores.
Table 3.4: The effect of MNCP score cut-offs to the motif prediction from the Sandve’s
benchmark.
Table 3.5: List of input cDNAs from the maize anthocyanin pathway used for Promzea
searches.
Chapter 4
Table 4.1: Detailed description of developmental stages and tissues sampled.
Table 4.2: Tissue/stage-specific interesting genes.
Table 4.3: GO terms of the photosynthetic genes up-regulated in seminal roots compared
to nodal roots.
Chapter 5
Table 5.1: Over-represented motifs associated with vegetative phase change.
Table 5.2: Probe sets differentially expressed in leaf and root tissues
Chapter 6
Table 6.1: Detailed description of developmental stages and tissues sampled.
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Table 6.2: Promoter cis-acting motifs over-represented in the nitrogen-related genes
expressed during maize development at different stages/tissues.
Table 6.3: NRE/Acore motifs detected in nitrogen-related promoters from gene
expression Cluster 1.
Chapter 7
Table 7.1: Real time qRT-PCR primers used to determine the RH/root expression ratio of
N-related genes.
Table 7.2: Relative expression of nitrate and ammonium transporters in root hair cells.
Table 7.3: Ammonium transporters retrieved from the maize genome, and orthologs
from rice and Arabidopsis.
Table 7.4: Categories of differentially expressed genes in the root hair response to nitrate
addition.
Table 7.5: Hormone-related genes that are differentially expressed in root hairs at 0.3
mM versus 3 mM nitrate at 30 min and 3 h following addition of liquid root hair media
(RHM).
Chapter 8
Table 8.1: Relative expression of genes related to root hair growth and development
following rehydration in root hair.
Table 8.2: Hypergeometric test p-values corresponding to the over-represented gene
categories shown in Figure 7.5.
Table 8.3: Relative expression of hormone-related genes that were differentially
expressed in maize root hairs following rehydration.
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