14.3: The 'Jumping Genes' of Maize

Barbara McClintock’s report that bits of DNA could jump around and integrate into new loci in DNA was so dramatic and arcane that many thought the phenomenon was either a one-off or not real! Only with the subsequent discovery of transposons in bacteria (and in other eukaryotes) were McClintock’s jumping genes finally recognized for what they were.

To begin the tale, let’s look at the maize reproduction in Figure 14.7.

Figure 14.7: Life cycle of maize: sperm from male plant pollen land on stigma atop corn silk to tunnel through a pollen tube of their own making to the base of the silk. There, one sperm fertilizes the egg; another fertilizes poplar bodies left over from oogenesis, which become triploid endosperm cells.

The different colors of corn seeds (kernels) result from anthocyanin pigments, which are expressed differentially by cells of the aleurone tissue, derived from the triploid endosperm. McClintock was studying the inheritance of color variation, which ranged from colorless (white or yellow, due to an absence of anthocyanins) to brown, purple, spotted, or streaked. The mosaic of kernel colors is vivid in the corncobs in the photo in Figure 14.8.

Figure 14.8: Examples of mosaic corn cobs with different color kernels on the same cob

Clearly, kernel color is inherited. The inheritance of colorless and purple seed color does indeed follow Mendelian rules, but the genetics of mosaicism does not. Mosaic color patterns after genetic crosses were not consistent, implying that the mutations responsible for kernel colors were not due to mutations in germ cells. Rather, genes controlling anthocyanin synthesis must have been undergoing mutations in the somatic cells that would become (or already were) the ones in which the pigments were produced.

242 What Interested McClintock about Maize

14.3.1 Discovering the Genes of Mosaicism: The Unstable Ds Gene

As we describe McClintock’s experiments, keep in mind that her research and intuitions about gene regulation and epigenetic inheritance came long before molecular technologies made it possible to prove or to name these phenomena. McClintock was looking for a genetic explanation for seed-color variation in the 1940s and early 1950s. DNA structure had only recently been published, and gene cloning and DNA sequencing were decades into the future! Her only available technologies were based on an understanding of Mendelian allelic assortment in traditional breeding studies. Since seed color is expressed in cells derived from endosperm, McClintock knew that the inheritance of the kernel-color phenotype must be studied against a triploid genetic background. She was also aware of speculations that the variegated-color phenotype might result when a specific unstable mutation, which usually produced colorless kernels, “reverted” in some cells but not in others, thereby creating a spotted or streaked phenotype. Just what made for an “unstable mutation” was, of course unknown. McClintock identified three genes involved in seed-kernel coloration and ultimately solved this puzzle.

CHALLENGE

Later in your study, you should be able to design one or more experiments based on molecular technologies that might have quickly revealed how transposons influence maize mosaicism.

Two of the genes studied by McClintock controlled the presence vs absence of kernel color. These are the C and Bz genes:

1. C′ is the dominant inhibitor allele, so-called because if even one copy were present, the kernels would be colorless (yellow), regardless of the rest of the genetic background.
2. Bz and bz are dominant and recessive alleles of the Bz gene, respectively. In the absence of a dominant C′ allele, the presence of a Bz allele would lead to purple kernels. If the bz allele were present without both C′ and Bz alleles, the kernels would be dark brown.

The third gene—the one required to get the variegated kernel color—was the Ds (Dissociator) gene. McClintock knew that without a viable Ds gene, kernels were either colored or colorless, depending on the possible genotypes dictated by the C and Bz alleles. In other words, the Ds gene must suffer the “unstable mutations” that led to variegated kernel color.

The mutations occurred at random among aleurone layer cells, so the mutations must have been occurring in a region of chromosomal instability (prone to damage or breakage) in some cells but not in others. Let’s look at what McClintock did to figure out what was going on in corn-kernel color genetics. Having already demonstrated crossing-over in maize (another remarkable achievement!), McClintock mapped the C’, Bz, and Ds genes to chromosome 9. She then selectively mated corn with genotypes shown in the protocol in Figure 14.9.

Figure 14.9: An experimental cross between CCbzbz triploid (triple recessive) female with no Ds genes and C′C′BzBzDsDs (triple dominant) male with Ds genes, showing predicted genotypes and kernel color phenotypes, based on Mendelian assumptions.

Remember that triploid cell genotypes are being considered in this illustration! You can refer to the phenotypic effects inherent in three-gene allelic backgrounds as we follow McClintock’s cross. Her cross of a homozygous recessive with a homozygous dominant plant should ring a bell! Let’s look more closely at this cross. Figure 14.10 shows the expected triploid genotypes from the cross. Aleurone cells resulting from this cross should all be colorless (yellow) because of the presence of the dominant C’ allele.

Figure 14.10: Triploid genotypes expected for the cross in Figure 14.9.

But while there were indeed many colorless kernels on the hybrid cob, there were also many mosaic kernels with dark spots or streaks against a colorless background. McClintock’s interpretation of events is illustrated in Figure 14.11.

Figure 14.11: McClintock’s interpretation of results of the triploid cross in Figure 14.9

According to McClintock, if some aleurone-layer cells in some kernels were to suffer chromosomal breakage at the Ds (Dissociator) locus (indicated by the double slash, //), the C’ allele would be inactivated. Without a functional C’ allele, the operative genotype in the affected cells is CCbzbz. These cells will revert to making the brown pigment as directed by the bz allele. When these cells divide, they form clusters of brown cells that are surrounded by cells with an unbroken chromosome (and thus an active C’ allele), creating a mosaic—that is, the appearance of pigment spots or streaks in some kernels, against the otherwise-colorless background of the surrounding cells.

243 Variegated Maize Kernels Result from "Loss" of the DS Gene

14.3.2 The Discovery of Mobile Genes: The Ac/Ds System

The experiments just described were reproducible using her original, single breeding stock of maize. But when McClintock tried to repeat the experiments by crossing the homozygous dominant males with homozygous recessive females from a different breeding stock, all the kernels of the progeny cobs were colorless, as if the Ds gene had not caused any chromosomal damage.

CHALLENGE

Outline the cross escribed above to show (i.e., explain) the described genetic outcomes.

It seemed that the Ds genes contributed by the males were unable to function mutate (i.e., “break”) chromosomes in females of this new breeding stock. McClintock hypothesized that the female in the original cross must have contributed a factor that could somehow activate the Ds gene to break, and that this factor, yet another gene, was absent or inactive in the females of the new breeding stock. McClintock called the new factor the activator, or Ac gene. Based on the dependence of Ds on the Ac locus, McClintock recognized that mosaicism in maize kernels was controlled by these two “genes” as part of a two-element, Ac/Ds system.

She then demonstrated that Ac-dependent Ds “breakage” was in some cases also associated with inactivation of a normal Bz gene, leading to a loss of purple kernels. At this point McClintock concluded that, far from simply “breaking” the chromosome at a fragile Ds locus, the Ds gene had moved to (or into) the Bz gene, disrupting its function. Again, this could not happen in the absence of an active Ac gene.

McClintock had discovered the first transposon, earning her the 1983 Nobel Prize in Physiology or Medicine, albeit an honor belated by decades! View the homage to, and a brief history and summary of, McClintock’s work and impact at A short history of McClintock Science. With the advent of recombinant DNA technologies, we now know the following:

1. The Ds transposon lacks a gene for a transposase enzyme required for transposition.
2. The Ac element has this gene and is capable of independent transposition.
3. Ac provides the transposase needed to mobilize itself and the Ds element.
4. The sequence similarity of Ds and Ac elements supports their common ancestry.

The basic structural features of the maize Ac/Ds system are listed here:

1. Ac is 4,563 bp long.
2. Ds is a truncated version of Ac.
3. There are eleven bp inverted repeats at either end of the Ac and Ds elements.
4. There are eight bp direct repeats of insertion site DNA created at the time of transposition flanking each of the elements.

Look for these features as we describe prokaryotic and eukaryotic transposons.

244 Discovery of Mobile Elements and the Ac-Ds System

245 The Ac-Ds System Today