Chromosome Connections: Compelling Clues to Common Ancestry

INQUI RY & INVESTIGATI ON Chromosome Connections: Compelling Clues to Common Ancestry RECOMMENDATION LARRY FL AMMER ABSTRACT Students compare banding patterns on hominid chromosomes and see striking evidence of their common ancestry. To test this, human chromosome no. 2 is matched with two shorter chimpanzee chromosomes, leading to the hypothesis that human chromosome 2 resulted from the fusion of the two shorter chromosomes. Students test that hypothesis by looking for (and finding) DNA evidence of telomere segments at the fusion site, thus reinforcing the likelihood of our common ancestry with chimps and showing that we all carry the molecular fossils of telomere fusion! Students see how multiple lines of evidence make a compelling case for common ancestry. Key Words: Hominid chromosomes; common ancestry; DNA fusion; hypotheses tested. BANG! What was that? If you actually heard this sound with the unmistakable qualities of a gunshot you certainly would notice it and wonder what was happening. Depending on where you are, this sound could signal a real crime, with real bullets. And sooner or later, crime-scene investigators may gather, looking for clues to what happened. Crime-scene investigations are among the closest applications of the process of science to real-world questions especially to explain events of the past that were not witnessed by anyone living today. Analyzing clues produced by an unwitnessed earlier event is fascinating science in its own right and characterizes the historical sciences (e.g., astronomy and paleontology). As you may know, a bullet used in a crime can be linked to the gun that fired it by analyzing the scratch marks on the bullet (Figure 1). When a second bullet (safely fired from the suspect gun) has a pattern of marks identical to those on a bullet found at the crime scene, we can be very confident that those two bullets came from the same gun. In other words, they had a common origin: the gun that fired them (Wallace, 1966). Each pair of chromosomes in an organism s full set of chromosomes has its own unique pattern of bands, characteristic for each species. The Chromosome Connection You may ask, what does all this have to do with chromosomes? When the chromosomes from a certain stage of the cell cycle are stained in a special way, they will appear under a microscope to have a series of dark and light bands (Figure 2). When many photos of each chromosome are carefully analyzed, diagrams of each can be developed for easier comparison of details. Each pair of chromosomes in an organism s full set of chromosomes has its own unique pattern of bands, characteristic for each species. When the chromosomes from two different species are compared and are found to have very similar banding patterns, what do you suppose this suggests? These matched patterns, as we find with bullet marks, indicate a common source a shared biological origin of those chromosomes. The chances that two sets of identical complex banding patterns had independent unrelated origins are vanishingly small. We conclude that those two species must have had a common origin a common ancestry. In Figure 3 (adapted from Yunis & Prakash, 1982), we compare, side by side, four of the chromosomes (nos. 3, 4, 5, and 6) from two different species we ll call C and G. Do you see the identical or nearly identical patterns on the corresponding chromosomes? These line patterns are too complex to exist in two species by chance. What does that suggest? If you said that they must have a shared source or a common ancestor you would be right. Furthermore, when you learn that nearly all the chromosomes from one species appear to be identical in whole or in part to those of a second species, this strengthens the likelihood that those two species have a common ancestor. Ask your students to count the total number of clear differences between the C and G chromosomes shown here and record that number privately as count 1. Announce that the two species being compared here are chimpanzees (C) and gorillas (G), and that their chromosome similarities clearly point to them having a common (shared) ancestor at The American Biology Teacher, Vol. 75, No. 2, pages 108 113. ISSN 0002-7685, electronic ISSN 1938-4211. 2013 by National Association of Biology Teachers. All rights reserved. Request permission to photocopy or reproduce article content at the University of California Press s Rights and Permissions Web site at www.ucpressjournals.com/reprintinfo.asp. DOI: 10.1525/abt.2013.75.2.7 108 THE AMERICAN BIOLOGY TEACHER VOLUME 75, NO. 2, FEBRUARY 2013

Figure 1. Matched bullets from a common source. ask them to raise their hands for the opposite result: their count 1 was less than their count 2. Because counting differences involves some judgment, they will vary, but one count will usually be higher. Also, tell your students that they need to realize that these are only four chromosomes out of the 23 or 24 pairs of chromosomes that these three species have, but even those other chromosomes are remarkably similar. At this point, project a copy of Figure 5 on your screen. This shows the diagrams of each chromosome found in four hominid species, with closely matched chromosomes side by side. This figure is available directly from http://www.indiana.edu/~ensiweb/ lessons/chr.4.all.pdf (courtesy of Jorge Yunis, 1982). From all of this, you can probably agree that both species P and species G are certainly very similar to C (and therefore to each other), which means that they all probably shared a common ancestor at some time in the distant past. Of course, your students will be asking What is species P? Delay answering! The suspense is most engaging. Ask them to guess. Write down their guesses on the board, without revealing whether any guess is right or wrong. Eventually, you must give in and tell them that P is us people Homo sapiens. Furthermore, in careful studies of the differences in these chromosomes, it turns out that chimps are more like us than they are like gorillas (Figure 6)! Figure 2. Stained chromosome: photo and diagram. some time in the past that was neither a chimp nor a gorilla. That ancestor produced two offspring in the distant past that were slightly different (genetically) and probably became part of two different populations that became reproductively separated, and their offspring inherited those chromosome differences. Over many more generations, additional changes accumulated in each population, eventually resulting in two new species: C (chimps, Pan troglodytes) and G (gorillas, Gorilla gorilla). We find many similar but different species today that show various stages in their growing genetic dissimilarity. In Figure 4 (adapted from Yunis & Prakash, 1982), we compare this same selection of chimpanzee chromosomes (C) with another species (P). With so much of their banding patterns being identical, you would have to conclude that these two species must also have had a common origin a shared ancestor. Ask your students to study these chromosomes, count the obvious differences between C and P for all four chromosomes combined, and privately record the result as count 2. When all have finished their counts, ask your students to raise their hands if their count 1 was greater than their count 2. Then Figure 3. Compare 4 chromosomes (3, 4, 5, 6) between 2 species (C and G). THE AMERICAN BIOLOGY TEACHER CHROMOSOME CONNECTIONS 109

Figure 4. Compare 4 chromosomes (3, 4, 5, 6) between 2 species (C and P). In fact, those studies tell us that the chromosomes of both chimpanzees and gorillas have changed more than our chromosomes have since the split, which suggests that we may be more like our common ancestor than chimps and gorillas are (Yunis & Prakash, 1982). Again, remind them that these are only 4 of the 23 pairs of chromosomes in us and of the 24 pairs of chromosomes in chimps, gorillas, and orangutans. A Molecular Fossil: Chromosome 2 If your students pick up on this difference (23 vs. 24), perhaps, with a few hints, you can then shift to our chromosome 2, and its comparison with a chimp s two shorter chromosomes, 2a and 2b. In Figure 7 (adapted from Yunis & Prakash, 1982), these two chimp (C) chromosomes (2a and 2b) are positioned next to our no. 2 (P), and they match perfectly! How could this happen? You might consider two possible explanations (hypotheses): A. The common ancestor had one long chromosome that split in half sometime in the chimp population, to make the two short ones that we see in chimps today, while we kept our one long chromosome 2 to the present, or B. The common ancestor had two short chromosomes that fused end-to-end early in the unique human branch, to make the long chromosome 2 we see in us today, while the chimp branch kept its two short chromosomes. For reasons discussed in the Chromosome Fusion lesson (below), we choose explanation B to test. We should then predict what we will find only if the hypothesis is true. So, if fusion of the two chromosomes is what happened and you closely examine our chromosome 2, what might you expect to find in the supposed fusion area that could be considered evidence of this fusion? Is there, perhaps, something special about the DNA in that region? Something that is not obvious here, and was unknown to Yunis and Prakash (1982), is that we now know the DNA sequence in the ends of chromosomes (called telomeres ). They typically consist of a long series of tandem repeats, like this (showing only one DNA strand here, assuming the usual complement on the other strand): ttagggttagggttaggg etc., with ttaggg repeated many hundreds of times. So, if two short chromosomes (like those found in chimps today) came together end-to-end to make our chromosome 2, we can predict that we should find the remains of those telomeres near the middle of our chromosome 2. On the other hand, finding telomere sequences in the middle of a chromosome that was not the result of fusion would be very unlikely. By 1990, we knew the DNA sequence near the middle of our chromosome 2, so that hypo thesis (choice B above) could be tested by looking for those telomere remnants in the supposed fusion zone of our chromosome 2 (Ijdo et al., 1991). When we go into the online database of DNA for our chromosome 2 and search for the telomere sequences in the exact region where fusion must have happened, we do, indeed, find what is clearly the tandem repeats of ttagggttaggttaggg, then suddenly aatcccaatcccaatccc, the complementary sequence in the telomere DNA of the other short chromosome. Actually, the sequence isn t exactly like this, because of mutations since the fusion time several million years ago, but it s very close! In a very real sense, these damaged telomere sequences in our chromosome 2 are actually molecular fossils the remains of the telomeres of two separate chromosomes in our ancestor and every one of us has those molecular fossils in our cells! Our prediction is confirmed, reinforcing our confidence in the way we got our chromosome 2 (Ijdo et al., 1991; Fan et al., 2002). Your students can actually do this search, following the directions in the Chromosome Fusion lesson on the ENSI website (see list below). Or, if you lack the time or computer access to do this, you can use a copy of the DNA strands in the fusion zone, taken from the data bank and provided in that lesson for your students to search directly for their molecular fossils. At this point, perhaps you can see another prediction about our chromosome fusion hypothesis that we could test. Look at the diagram of our chromosome 2 next to the two matching chimp chromosomes (Figure 7). What do you see that we could look for from the fusion of those two short chromosomes? If you need a clue, look at the number of centromeres (constricted parts) in each chromosome. Do you see it? Obviously, the centromere in one of the short chromosomes continues to function as the centromere of our chromosome 2 (chromosomes can have only one centromere). But what happened to the centromere in the other short chromosome? What would you expect to find in the region of our chromosome 2 that coincides with that centromere? Perhaps some kind of DNA evidence for the remains of that centromere? In 1992, Avarello and colleagues reported their discovery of the centromere evidence from that other fused chromosome. Hypothesis strengthened: fusion confirmed again. Once scientists figured out the chimp genome (Chimpanzee Sequencing and Analysis Consortium, 2005), we could compare human DNA with chimp DNA. When we compare all the 110 THE AMERICAN BIOLOGY TEACHER VOLUME 75, NO. 2, FEBRUARY 2013

Figure 5. All chromosomes from four hominid species: human, chimp, gorilla, and orangutan. With kind permission of Jorge Yunis (Yunis & Prakash, 1982). THE AMERICAN BIOLOGY TEACHER CHROMOSOME CONNECTIONS 111

Figure 6. Chromosome differences (Yunis & Prakash, 1982). Figure 7. Chromosome 2 from species P and chromosomes 2a and 2b from species C (Yunis & Prakash, 1982). corresponding genes among apes and humans, we generally find identical or nearly identical sequences. As anyone who knows how DNA transcription and translation work, nucleotide replacement mutations in DNA can still produce functionally equal proteins even if you get a few different amino acids here and there in noncritical parts of the protein. In addition, most of the differences occur in noncoding (non-gene) segments and reflect the randomness of a regular rate of change that can be used to approximate how much time has passed since they branched from their respective shared ancestor. Examination also reveals that the chimpanzee branch had more DNA changes than the human branch since they separated from their common ancestor (Caswell et al., 2008). So now we have another DNA confirmation of the common ancestry of humans and chimps, plus an indication of how long ago this branching happened (about 3 6 million years ago). Multiple Lines of Evidence & Fair Test When several different lines of evidence, and their confirmed predictions, all consistently point to the same event, we call this consilience, one of the strongest kinds of evidence for what really happened. This is not a consensus, with scientists agreeing about an interpretation of data (or evidence). Rather, these are different kinds of independent data that all fit a particular hypothesis and are not all consistent with any other hypothesis. The chance that there is another explanation has become less and less. Unfortunately, many students in school don t get to experience consilience, so they miss seeing one of the most compelling features of science, especially for the science that points consistently to evolution the origin of species from previous species. These lessons using hominid chromosomes also serve as an excellent example of the Fair Test gauntlet that further strengthens scientific conclusions (Nelson, 2000). Fair Tests are challenges to two or more alternative possible solutions (hypotheses) to a problem. When the test of these hypotheses does not have the same basis used in forming those solutions, and could potentially support any of those alternatives, we have a Fair Test. When the Fair Test results are analyzed according to several criteria, we should find that one solution is better than the others meaning that it explains all the observations of the problem better than the alternatives. We call this the Best Explanation. This is what science is usually looking for. Why Hominid Chromosomes? Comparing molecules like DNA and proteins (e.g., Molecular Sequences and Primate Evolution at http://www.indiana.edu/~ ensiweb/lessons/mol.prim.html) reveals clear signs of evolution, but we can t directly see those molecular details, so these rather abstract kinds of evidence are probably not as convincing to students as clues that are more visual. Fossils can also be used and are certainly visual (see the Skulls Lab at http://www.indiana.edu/~ensiweb/lessons/hom. cran.html). However, their features showing gradual change are often quite subtle and not always obvious to the novice. But when students can actually see the striking similarities of chromosome banding patterns side-by-side from living species, it becomes very hard to ignore the obvious relationships that they imply. A good place to insert one or more of these chromosome lessons would be near the end of your chromosome genetics unit. In doing this, you also show how evolution fits into other topics of your course (Flammer, 2006). Furthermore, focusing on human evolution (rather than the evolution of moths or camels) is particularly engaging to students and provides many examples of how multiple lines of evidence ( consilience) can be so compelling. ENSI Lead-Teacher Beth Kramer has aptly pointed out that if students can be convinced that evolution happens in people, then extending that process to all other organisms is a snap! Most importantly, the experiences presented here have the greatest impact when students are actually engaged in making those observations. Access to Internet databanks of DNA and protein sequences enables students, with very little guidance, to make these discoveries themselves. The four interactive lessons below have been developed and classroom-tested to take students into different aspects of hominid chromosome comparisons. They are found on ENSIweb, the website for the Evolution & Nature of Science Institutes. All these lessons provide freely downloadable handout materials and suggested strategies for their presentation and assessment. Go to 112 THE AMERICAN BIOLOGY TEACHER VOLUME 75, NO. 2, FEBRUARY 2013

the Evolution Lessons Index at http://www.indiana.edu/~ensiweb/ evol.fs.html, click on List of Titles, then, under Human Evolution Patterns, find these lessons: Comparison of Human & Chimpanzee Chromosomes: Intro to hominid chromosomes. Chromosome Connection 2: Comparisons with more details of differences. Chromosome Fusion: Using DNA database to search for telomere vestiges. Mystery of the Matching Marks: Forensic approach, using interactive PowerPoint. Acknowledgments My thanks to Beth Kramer, ENSI Lead Teacher and colleague who created the excellent Comparison of Human & Chimpanzee Chromosomes lesson. She kindly reviewed my draft of this article and offered several changes that improved it. Also, my thanks to the person who reviewed my MS, responding with positive comments, and all the many teachers who used these lessons in their classes and sent me helpful feedback. Special thanks to the teachers who let me teach these lessons to their classes. Caswell, J.L., Mallick, S., Richter, D.J., Neubauer, J., Schirmer, C., Gnerre, S. & Reich, D. (2008). Analysis of chimpanzee history based on genome sequence alignments. PLoS Genetics, 4(4), e1000057. Chimpanzee Sequencing and Analysis Consortium. (2005). Initial sequence of the chimpanzee genome and comparison with the human genome. Nature, 437, 69 87. Fan, Y., Newman, T., Linardopoulou, E. & Trask, B.J. (2002). Gene content and function of the ancestral chromosome fusion site in human chromosome 2q13 2q14.1 and paralogous regions. Genome Research, 12, 1663 1672. Flammer, L. (2006). The evolution solution: teaching evolution without conflict. American Biology Teacher, 68. [Online article.] Available at http://www.nabt.org/websites/institution/file/pdfs/publications/abt/ archived-table/2006/068-03-0001.pdf. Ijdo, J.W., Baldini, A., Ward, D.C., Reeders, S.T. & Wells, R.A. (1991). Origin of human chromosome 2: an ancestral telomere telomere fusion. Proceedings of the National Academy of Sciences USA, 88, 9051 9055. Nelson, C.E. (2000). Effective strategies for teaching evolution and other controversial topics. In The Creation Controversy and the Science Classroom. Washington, D.C.: NSTA. Wallace, B. (1966). Chromosomes, Giant Molecules, and Evolution, pp. 7 and 81. New York, NY: Norton. Yunis, J.J. & Prakash, O. (1982). The origin of man: a chromosomal pictorial legacy. Science, 215, 1525 1529. References Avarello, R., Pedicini, A., Caiulo, A., Zuffardi, O. & Fraccaro, M. (1992). Evidence for an ancestral alphoid domain on the long arm of human chromosome 2. Human Genetics, 89, 247 249. LARRY FLAMMER is a retired high school biology teacher and currently Webmaster for ENSIweb (Evolution and Nature of Science Institutes). He lives in San Jose, CA. E-mail: flammer4@gmail.com. THE AMERICAN BIOLOGY TEACHER CHROMOSOME CONNECTIONS 113