Transposable elements: Barbara McClintock and early experiments in maizeIn 1938 Marcus Rhodes provided the first genetic description of an unstablemutation, an allele of a gene required for the production of pigment in maize.This instability resulted in a variegated phenotype. Variegation describesirregular patterns of pigmentation and reflects changes in the expression ofgenes that control pigment production. Rhodes defined two genes required forthis instability: the unstable allele defined one gene, and a dominant mutationcalled Dotted that controlled this instability mapped to a separate locus, defininga second gene. In the 1940s and 1950s Barbara McClintock showed that this typeof instability was caused by genetic elements that moved or transposed fromone locus to another. It was not until the description of transposable elements inbacteria two decades later that McClintock's work was truly appreciated.McClintock won the Nobel Prize for her work on transposable elements in 1983.The maize life cycle and kernel developmentA single corn plant contains both male and female germlines and is capable ofself or cross fertilization. Fertilization begins when a pollen grain lands andgerminates on a silk, a long structure connected to the ovule that entraps pollen.Each pollen grain contains several nuclei. The diploid male pollenprecursor cell undergoes meiosis, giving rise to four haploid microspores. Eachmicrospore divides mitotically several times to generate one pollen grain. Two ofthe products of these mitotic divisions are the two genetically identical spermnuclei. The diploid female macrospore mother cell undergoes meiosis togenerate four haploid macrospores, three of which degenerate. The remainingmacrospore divides mitotically three times to generate the embryo sac, whichincludes one egg nucleus and two haploid nuclei in the central cell.When a pollen grain lands on a silk, it produces a pollen tube that runsdown the silk to the ovule. The two pollen sperm cells migrate down the tubeand are deposited in an embryo sac. Fertilization is actually a double fertilizationevent. Upon reaching the embryo sac, one sperm cell fuses with the egg cell toproduce the diploid zygote, and the second sperm cell fuses with the twoprimary endosperm nuclei in the central cell to produce the triploid endosperm(and aleurone). Thus the embryo that develops from the fertilized egg cell andthe endosperm that develops from the fertilized central cell are geneticallysimilar except that the endosperm is triploid, containing two copies of thematernal genome. The kernel consists of an internal diploid embryo, a triploidstarchy endosperm, a triploid pigmented aleurone layer that surrounds theendosperm, and a hard pericarp that is of maternal origin. Each kernelrepresents the product of a single cross and many phenotypic traits can bescored in the triploid endosperm and aleurone layers of the kernel before it isgrown into a plant. A single ear of corn contains the products of multiplematings.The genetic and cytological markers used by McClintockIn the crosses we will describe here, McClintock was working on the short armof chromosome 9, where the order of genes and chromosomal structuralelements that we will consider are: telomere - Colorless (C+) - Bronze (Bz+) -Waxy (Wx+) - centromere. The Colorless gene is required for pigmentproduction in the aleurone layer and is assayed visually: C+ results in a purplecolored aleurone layer; the C-I mutation is dominant to C+ and results in acolorless phenotype; c mutations are recessive and also result in a colorlessphenotype. Bronze is also required for pigment production in the aleurone layerand assayed visually. Bronze encodes an enzyme that modifies the brown ofbronze colored product of the C+ gene, changing it to a purple pigment. Thus,Bz+ results in a purple colored aleurone layer, and bz mutations are recessive andresult in a brownish aleurone layer. If both the Colorless and Bronze genes aremutant, for example, in a C-I bx/C+ bz or a c bz/c bz kernel, the kernel iscolorless because the C+ pigment is not produced and the genotype at theBronze locus is irrelevant.The Waxy gene is required for the production of a specific starch in theendosperm that is assayed by a staining reaction: Wx+ confers a starchyphenotype in which the endosperm stains dark blue with an I2-KI solution;recessive wx mutations confer a waxy phenotype in which the endosperm stainsreddish brown with I2-KI.In addition to the genetic markers, McClintock used a cytological marker,a chromosome with a knob at the telomere. The normal chromosome lacks thisknob.McClintock's analysis of chromosomal breakage in maize led to thediscovery that two elements are involved, which she termed Dissociation (Ds)and Activator (Ac). Ds is located at the site of chromosome breakage and can bedefined both genetically and cytologically. Ac is dominant and is required forchromosome breakage at Ds.With an understanding of these genetic and cytological tools, there arefour experiments that we will discuss. These experiments showed that: (1) Ds is agenetic element that results in chromosome breakage; (2) Ds can move; (3) Accontrols Ds breakage and can also move; and (4) Ds can move into a gene toproduce an unstable allele.Dissociation (Ds) is a genetic element that results in chromosomal breakageKernels from a cross between C-I Bz+ Wx+ / C-I Bz+ Wx+ pollen and C+ bz wx /C+ bz wx ovules were of the genotype C-I Bz+ Wx+ / C+ bz wx. Most werecolorless and starchy (the Bz+ phenotype is not expressed in a C-I or c/cbackground). McClintock noted some kernels had colored (bronze), waxysectors; i.e., all three dominant traits derived from the male parent were lost.McClintock proposed that chromosomal breakage occurred at a hotspotbetween Wx+ and the centromere, and she called this hotspot Dissociation (Ds).She predicted that breakage at Ds resulted in an acentric fragment containing theC-I, Bz+ and Wx+ genes and that this fragment was lost during development ofthe kernel.The variegation was heritable, and McClintock mapped Ds 1 cM from Wx.She also provided cytological evidence for breakage at Ds to confirm herhypothesis. The telomere of the C-I Bz+ Wx+ chromosome was marked with theknob discussed above, while the C+ bz wx telomere was cytologically normal.Loss of the dominant markers coincided with the loss of the telomeric knob.Ds can moveMcClintock also discovered that the position of Ds is unstable
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