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Lecture 7 OutlineChimeras II
1. History of “Sports”2. The Chimera Concept3. Sectorial Chimeras4. Definitions5. Green-white-green (GWG)
periclinal chimeras
History and Application of Grafting
1. Plant Collectors 2. Darwin’s travels3. Winkler’s Chimera Concept4. Types of Grafts5. Modern Applications
Grafting occurs naturally
Figure 1A from Stegemann et al., 2012, PNAS 109:2434
Chloroplast genomes can be transferred from one tree to the other across the graft junction
History1. Explorers, like Charles Darwin,
traveled to new and exotic places and collected ‘treasures’in the form of novel species of plants and animals
2. New varieties of plants were highly prized and breeders were therefore motivated to generate new and improved stocks
a) by making interspecific crossesb) by vegetative propagation (or the
generation of a “sport”)
History1. A Sport was defined as, “A
spontaneous change or mutation in a part of a plant that created a feature not previously known within the species, their sexual hybrids and descendants”.
2. Sports were notoriously difficult to maintain and would often be unstable
3. The first reproducible and stable variant was described in 1674 and was called the ‘Bizzarria orange’
http://www.homecitrusgrowers.co.uk/citrusvarieties/bizzarria.html
Pietro Nati (1625-1685), who was the director of the Pisa Botanical Gardens, published one of the first
descriptions of a graft chimera known as “La Bizzarria”
Resulted from grafting a sour orange scion to a citron stock. A bud arose at the graft junction with a unique mix of parental characteristics.
Historical paradox: What was the nature of graft hybrids?
Were the resulting hybrid branches the result of sexual hybridization or a union of vegetative tissue (somatic segregation)?
The argument against sexual hybridization was the you could not produce such plants by conventional breeding methods.
Winkler 19071. Did experiments to test the nature of graft
hybrids and, using tomato and nightshade plants, reproduced results akin to those seen previously
2. Got shoots that were all tomato, all nightshade or 1/2 nightshade and 1/2 tomato
3. Winkler was responsible for coining the term “chimera” (a fire breathing monster from Greek mythology that had the foreparts of a lion, torso of a goat and the hindquarters of a dragon)
ConventionPericlinal chimeras are described
according to the relative contribution of graft partners, starting with the L1
1. If plant A is grafted to plant B and a periclinal chimera is produced, the layer contributions are listed beginning with the L1, then the L2 and finally the L3
2. ABA: L1 is A, L2 is B, L3 is A3. BBA: L1 is B, L2 is B, L3 is A
L1L2
L3
SAM
Vegetative Propagation1. Plants can be maintained by taking
cuttings and rooting them2. By grafting a cutting onto a stock3. Maintaining them in tissue culture4. Grafting is very important to
a) The horticultural industry (roses)b) For maintaining grape varieties for
wine production (root stocks resistant to pathogens)
c) The fruit industry (apples)
Two types of chimerasPericlinal and Sectorial
Within Organs
At the SAM
Sectorial Chimeras1. Plants with sectorial meristems show
sectors that run vertically down the shoot
2. Meristem is divisible into circumferential lineages and therefore there must be a small number of initial cells at the top of the meristem (subset of central zone)
3. Sectors are not usually stable (do not persist for the lifetime of the plant)
Sectorial Chimeras1. Position of an initial
does not restrict the fate of the initial’s descendants
2. The position of a marked band is independent of leaf initiation and may or may not encompass a whole leaf or even part of a leaf
Sectorial ChimerasWithin organs reveals division patterns
and clonal relationships
Daughter cells arising from one marked cell
VariegationSpontaneous sectors can arise from the disruption of pigment biosynthetic genes
caused by transposon insertion and excision.
DefinitionsGenotype: The DNA constitution
a) the genetic constitution of a cell, an organism, or an individual (i.e. the specific allele makeup of the individual) (from Wikipedia)
b) the actual alleles present in an individual
Phenotype: Visible consequencesa) an observable characteristicb) any observable characteristic or trait
of an organism (from Wikipedia)
Gene expression patternsIn theory, every nucleus in a multicellular organism
has the same genetic information but expression patterns are different. Genes are turned on and
off by multiple factors (temporal/spatial).
A direct relationship: gene is on, function is on or gene is off and function is off
Factors affecting gene expression
Location, location, location Developmental Regulation
A gene may be off if it resides in a cell in the L1 but not in the L2 (where you live matters!)
Activation due to a change in fate
Through cell division, an L1 daughter cell may end up in an L2 neighbourhood and hence different genes become activated and inactivated (turned on/off)
Green-white-green chimeras
L1=green, L2=albino, L3=greenConventionally written as GWG
What does that mean?1. The L1 cells are genetically green but
visibly (phenotypically) white because chloroplasts do not mature (suppressed)
2. The L2 cells are genetically albino or white (genes do not function: they have mutations) and phenotypically white
3. The L3 cells are genetically and phenotypically green (green genes are functional)
Green-white-green chimeras
The L2 meristematic layer is albino
From Leyser and Day, Mechanisms in Plant Development. 2003 Blackwell Publishing,Chapter 3.
Genotype of L1, L2 and L3L1=green, L2=albino (white), L3=greenPigments are cell autonomous markers
because they are contained within a plastid
Leaf in cross-section
Phenotype of L1, L2 and L3
L1=white (gene expression suppressed)L2=albino (genes have mutations)L3=green (genes are expressed)
Layer displacementsL1 cells that divide periclinally (rare) will give rise to L2
daughter cells that can make chloroplast pigment while L2 cells cannot, even if displaced into the L3. L2
cells invading the L3 occurs more commonly.
L1
L2 L3 layer
GWG Chimeras Demonstrate that….
1. L1 layer is very stable and rarely invades the subjacent L2 layer
2. L2 cells typically remain in the subepidermal layer in most of the leaf but near leaf margins are more likely to become displaced into the L3
3. If L2 cells invade, there is a compensatory reduction in cell divisions in the L3 so that the number of cell layers in the leaf remains constant
4. Hence one sees this fairly common pattern of variegation in garden and house plants (white margins and green centre)
Sectorial Chimeras are Rare
1. GWG to WWG, GWW or GGG sectors can occur but they tend not to be stable
2. GWW sector would give rise to a white sector on the stem and would run down the vertical axis of the shoot (leaf and stem tissues would be white)
3. GGG sector would produce a solid green band with no variegation
4. Seeds produced in any of these sectors would have the genotype of the L2 only
What have we learned so far?
1. Meristem layers are relatively stable (L1>L2>L3).
2. Proteins and RNAs can move between layers and there seems to be directionality.
3. Directionality is determined on a case-by-case basis.
4. There are tissue-specific constraints.
L1L2
L3
SAM
Some additional questions
1. Who is in charge? For example, is the coordination of meristem activity due to the response of L2/L3 tissues to signals from the L1?
2. Is there lateral movement of signals within a layer?
3. Are there communication rules and if so, what are the rules?
L1L2
L3
SAM