Download induction of instability at selected loci in maize

Biol 4241 ‐ Advanced Topics in Genetics
Paper Review
Tricia O'Brien, Juliet Le, Dave Watton, Heidi Matchem
"INDUCTION OF INSTABILITY AT SELECTED LOCI IN MAIZE"
Barbara McClintock
What are “Jumping Genes”?
Jumping genes, or transposable genetic elements:
Chromosomal segments capable of moving from one position on the chromosome to another within the genome.
The genome is not static; genes have the ability to move.
The normal genetic role of this is unknown, but is a universal phenomenon and can be found in many different organisms,
including:
plants (eg. maize, parsley, and snapdragons)
humans (Alu elements)
insects (eg. Drosophila melanogaster P‐elements)
bacteria
..and many others!
How were these first discovered?
Transposable elements disrupt genetic function and, therefore, phenotype.
Fall into a broad definition of mutation.
First identified due to abnormalities observed in the maize (Zea mays) by Barbara McClintock.
Create phenotypic ratios that do not follow Mendelian genetics, therefore creating curiosity.
Spotted or "variegated kernels".
McClintock suggested a system of control tht she referred to as "extragenic units".
The Dissociation‐Activation (Ds‐Ac)two‐unit system controls instability genetic expression:
These two "extragenic units" she decribed are:
Ds (Dissociator)– disrupts gene expression
Causes chromosomal breaks or inserts itself into the gene
Affects genes in the cis configuration (i.e. adjacent to the break or insertion)
BUT
Only when Ac (Activator) is present in the genome in the trans configuration (i.e. on the homologous chromosome)
Capable of autonomous movement.
Cytological and Genetic Consequences of Ds insertion:
Ds‐induced chromosome breakage and loss of acentric fragment with downstream loci in 2 synapsed chromosomes.
Loss of the dominant alleles "uncovers" expression of recessive phenotypes.
Ds‐insertion is reversible; it may subsequently "jump out" in the presence of Ac, creating the original phenotype.
Therefore, genetic expression is unstable in those cells with Ac and Ds in the genome.
A simplified illustration & animation!
Ac Dosage
Transposition of the Ac element only occurs during chromosome replication.
0‐3 "doses" of Ac can be found in maize plants, depending on when transposition occurs.
Background
Genetic instability (mutation) was discovered at several previously known loci.
This paper looked to see if Ac controlled mutability can appear appear at other loci of known genetic action (C, Bz, or Wx
demonstrated previously)
A1 and A2 were genes selected (responsible for pigment formation) ‐ Did not affect plant vitality and easily observable
Recessives were discovered to be stable in the presence of Ac
Predictions:
1) Mutations only occur when Ac is in the nucleus
2) In absence of Ac modifications are stable 3) If Ac returns mutations no longer stable
4) Increased dosage of Ac results in later development of variegation
McClintock's Experiments:
McClintock set up a series of ingenious crosses to prove her hypothesis. In order to determine the genetic constitution of progeny kernels, further crosses with these kernels completed in successive
years.
Frequecy of phenotypes were recorded.
Results of Her Experiments:
Table 2: Ac Constitution of gametes of plants of culture 6424 determined by crosses to plants of Ac tester stock: I Ds/I Ds;
A1/A1; no Ac.
Ac Constitution of Plant
Ratios
Kernel Types on Resulting Ear (%)
Colorless (no Ac)
Variegated (Ac)
Group 1 ‐ No Ac
100
0
1:0
Group 2 ‐ 1 Ac
50
50
1:1
Group 3 ‐ 2 Ac
~6
~94
~0:1
Ac Tester Stocks:
McClintock created "tester" maize plant strains with which she could cross other plants to determine their Ac "constitution". The presence of Ac can be detected because it produces breaks at Ds leading to mutations at a loci of genic action.
These plants have NO AC FACTOR!
Many of these tester stocks utilized genetic markers on the short arm of chromosome 9 with Ds at it's standard location.
Table 3: Types of kernels appearing on ears resulting from crosses of plants of culture 6424 with tester plants of the following
constitutions. Part 1: tester plants have no Ac. Part: tester plants have 1 or 2 Ac factors.
Constitution of Tester Plant
a1 sh2/a1 sh2 (No Ac)
a1 Sh2/ a1 sh2 (No Ac)
Constitution of Plant Kernel Types (%)
Ratios Colored
Variegated
Colorless
a1m‐4 Sh2/a1m‐4 Sh2 (No Ac)
0
0
100
0:0:1
a1m‐4 Sh2/a1m‐4 Sh2 (1 Ac)
1
48
51
0:1:1
a1m‐4 Sh2/ a1m‐4 Sh2 (2 Ac)
4
83
13
0:6:1
a1m‐4 Sh2/ a1 sh2 (No Ac)
0
0
100
0:0:1
a1m‐4 Sh2 a1sh2 (1 Ac)
1
25
74
0:1:3
a1m‐4 Sh2/ a1 sh2 (2 Ac)
<1
45
55
0:1:1
a1m‐4 Sh2/ a1m‐4 Sh2 (No Ac)
0
0
100
0:0:1
a1m‐4 Sh2/ a1m‐4 Sh2 (2 Ac)
4
85
11
0:6:1
a1m‐4 Sh2/ a1 sh2 (1 Ac)
<1
24
76
0:1:3
a1m‐4 Sh2/ a1 sh2 (2 Ac)
<1
49
50
0:1:1
a1 Sh2/ a1 Sh2 (No Ac)
a1m‐4 Sh2/ a1 sh2 (No Ac)
0 0
100
0:0:1
a1m‐4 Sh2/ a1 sh2 (1 Ac)
2
24
74
0:1:3
a1 sh2/ a1 sh2 (1 Ac)
a1m‐4 Sh2/ a1m‐4 Sh2 (No Ac)
0
50
50
0:1:1
a1m‐4 Sh2/ a1 sh2 (No Ac)
0
25
75
0:1:3
a1m‐4 Sh2/ a1m‐4 Sh2 (No Ac)
0
51
49
0:1:1
a1m‐4 Sh2/ a1 sh2 (No Ac)
0
27
73
0:1:3
a1 Sh2/ a1 Sh2 (1 Ac)
a1m‐4 Sh2/ a1 sh2 (No Ac)
0
27
73
0:1:3
a1 Sh2/ a1 sh2 (2 Ac, Non‐allelic, Non‐linked)
a1m‐4 Sh2/ a1 sh2 (No Ac)
0
40
60
0:1:1.5
a1 Sh2/ a1 sh2 (1 Ac)
Trends observed from Table above:
No Ac present in either plant ‐ ALL kernels colorless (no exceptions)
No subsequent mutations due to the movement of Ds in the absence of Ac.
Recessive trait continues to be expressed in progeny.
1 Ac present ‐ resulted in either 1 variegated : 1 colorless (OR) 1 variegated: 3 colorless
1:1 ratio will occur if the plant is homozygous for the mutation
1/2 of the gametes possess Ac and 1/2 will possess the mutation.
The Ds in the cells possessing Ac can subsequently "jump out" creating variegation in the plants arising from this cross.
The other half will posses the mutated a1m‐4 locus or recessive a1 allele expressing the colorless phenotype.
1:3 ratio will occur if the plants are not homozygous for the mutation (i.e. there is only one a1m‐4).
1/2 of gametes wil posses Ac, while only 1/4 of the gametes will possess the mutation.
Therefore, less kernels will the mutation in which Ds can "jump out" of, producing less variegated kernels.
2 Ac present (allelic & linked: Ac inherited together)‐ resulted in either 1 variegated : 1 colorless (OR) 6 variegated : 1 colorless
1:1 will occur when the plant is not homozygous for the mutation.
1/2 of progeny will possess 2 Ac factors, while the other will possess NO Ac.
1/4 of the progeny receive the mutation, but the higher dosage of Ac.
More mutations at the A1 locus create a greater a proportion of variegated kernels.
6:1 will occur when the plant is homozygous for the mutation.
1/2 of the progeny will possess 2 Ac factors.
1/2 of the progeny possess the mutation.
High dosage of Ac increases the number of subsequent mutations and variegated kernels. 2 Ac present (non‐allelic & non‐linked: Ac inherited independtly; chance of cross‐over) ‐ resulted in 1 variegated : 1.5 colorless
when crossed with a plant not homozygous for the mutation.
In theory, NO completely colored kernels should have been observed
All tester plants were homozygous recessive for a1, while all plants crossed were either recessive for a1 or possessed the
mutated a1 locus).
However, may be due to changes in an Ac‐controlled mutable locus which usually take place in later developmental stages.
Could others see what she saw?
Conveying her ideas...
Jumping Genes Today!
The genome was originally thought to be a static master blueprint. A "safe deposit box"
Rearrangements only happened during meiosis with the occasional (often deletarious) mutation
It it now known that mobile DNA is not an exception but is ubiquitous
As much as 50% (!!!) of the human genome Greater than 60% of the maize genome
Originally thought to be useless. "Junk DNA"
But has some known functions:
1) Affecting Gene Expression
2) Generating Mutations
Ex: 50% of new mutations in Drosophila, 10% in Mice
3) Reorganizing Genome Structure
After McClintock's discovery in corn transposons were discovered in Drosophila
but still largely ignored or written off as uninportant
1960s E. coli
Transposons could then be studied with the aid of the new techniques of molecular biology
With the discovery of genome sequencing the sheer magnitude of the number of transposons was revealed.
3 known Categories:
1) DNA Transposons (Like McClintock's Ds‐Ac system)
2) LTR (viral‐like) Retrotransposons
3) Poly‐A (non‐LTR) Retrotransposons
Retrotransposons have an RNA phase.
There are autonomous (systhesize transposase) and non‐autonomous transposons
In the human genome most mobile DNA sequences are inactive but still 1/600 human disease causing mutations are caused by
retrotransposon jumping Some sample functions of mobile DNA:
1) Maintain the ends of Drosophila chromosomes
2) Important component of centromeres (not fully understood)
3) Arrangement of immunoglobulin genes
Also an important tool for developing mutants
Ex: P elements in flies
Sleeping Beauty transposon system
“A maverick. A visionary”
Genes are now thought of as a dynamic structure, rather than as a static linear sequence (as was up until the 1960s)
How does scientific knowledge arise? Who’s goals does it reflect?
Do we all seek the same type of explanations?
Marcus Rhoades, lifelong friend and companion:
“One of the remarkable things about Barbara McClintock’s surpassingly beautiful investigations is
that they came soley from her own labors. Without any technical help of any kind she has by virtue of
her boundless energy, her complete devotion to science, her originality and ingenuity, and her quick and
high intelligence made a series of significant discoveries unparalleled in the history of cytogenetics. A
skilled experimentalist, a master at interpreting cytological detail, a brilliant theoretician, she has had
an illuminating and pervasive role in the development of cytology and genetics.” –M. Rhoades