Did GDF6 “gene tweak” allow humans to become upright?

The short answer is, “Not really.” But as is often the case, the real story behind so many headlines last week is a bit more complicated.

smh.

smh. Links to the first, second, third, and fourth stories.

What are they talking about, Willis?

These headlines, each saying something slightly different, are referring to a study by Indjeian and colleagues published in Cell.  Researchers identified a stretch of DNA that is highly conserved across mammals, or in other words, it is very similar between very different organisms. In humans, however, this conserved region is actually missing (“hCONDEL.306”):

Fig. 4A from Indjeian et al. 2016. A stretch of DNA, "hCONDEL.306" is completely missing in humans (as is another stretch, hCONDEL.305) but otherwise very similar between chimpanzees, monkeys and mice.

Fig. 4A from Indjeian et al. 2016. A stretch of DNA on Chromosome 8, “hCONDEL.306,” is very similar between chimpanzees, macaque monkeys, and mice, but is completely missing in humans (as is another stretch, hCONDEL.305).

That a stretch of DNA should be highly conserved across diverse animal groups suggests purifying natural selection has prevented any mutations from occurring here – alterations to this stretch of DNA negatively affected fitness. But that humans should be missing such a highly conserved region suggests that this deletion came under positive natural selection at some point in human evolution. This strategy, of seeking stretches of DNA that are similar between many animals but very different in humans, has led to the identification of hundreds of genetic underpinnings of human uniqueness. Some of these, such as the case in question, involve deleted sequences and have been termed “hCONDELs,” for “regions with high sequence conservation that are surprisingly deleted in humans” (McLean et al., 2011: 216). Others involve the accumulation of mutations where other animals show few or none (e.g., HACNS1; Prabhakar et al. 2008). In many (most?) cases these are “non-coding” sequences of DNA.

How can “non-coding” DNA help make humans upright?

As was predicted 30 years ago (King and Wilson, 1975), what makes humans different from other animals isn’t so much in the protein-coding DNA (the classical understanding of the term, “genes”), but rather in the control of these protein-coding genes. “Non-coding” means that a stretch of DNA may get transcribed into RNA but is not then translated into proteins. But even though these sequences themselves don’t become anything tangible, many are nevertheless critical in regulating gene expression – when, where and how much a gene gets used. It’s wild stuff. Indeed, “Many human accelerated regions are developmental [gene] enhancers” (Capra et al., 2013).

In the present case, hCONDEL.306 refers to the human-specific deletion of a developmental enhancer located near the GDF6 gene, which is a bone morphogenetic protein. The major finding of the paper, as stated succinctly in the Highlights title page, is that “Humans have lost a conserved regulatory element [hCONDEL.306] controlling GDF6 expression…. Mouse phenotypes suggest that [this] deletion is related to digit shortening in human feet.”

How do they link this “gene tweak” to digit shortening?

Since humans have lost this gene enhancer that is highly conserved in other mammals, Indjeian and team reasoned that the chimpanzee DNA sequence associated with this deletion, retaining the enhancer sequence, is likely the ancestral condition from which the human version evolved. They inserted the chimpanzee version into mouse embryos and watched what happened as they developed. The enhancer was only active in the mice’s back legs, specifically in the cartilage that would later become the lateral toe bones and cells that would become a muscle of the big toe (abductor hallucis). These are areas where humans and chimpanzees differ: our lateral toes are shorter than chimps’, and we only have one abductor hallucis muscle whereas chimpanzees have an additional, longer abductor hallucis  (Aiello and Dean, 2002). So, we’re on our way to seeing how hCONDEL.306 might relate to our big toe or upright walking, as the headlines say.

But this still doesn’t explain how this deletion affects GDF6 gene expression, and therefore what this does for our feet. Pressing onward, the scientists compared the size of certain bones in mice with a normal Gdf6 gene, and those in which the Gdf6 gene was completely turned off (or “knocked out”).  The Gdf6 knock-out mice had shorter lateral toe bones than regular mice, but they also had shorter big toes as well – the previous experiment staining mouse embryos showed the ancestral enhancer was expressed more in the latter toes, not so much the big toe.

Figures 5-6 from Indjeian et al. (2016) sum up the findings. Figure 5 (left) shows that the ancestral version of the GDF6 enhancer (blue staining) is most strongly expressed in the lower limb, especially the fifth toe bone. Figure 6 (right) shows that a lack of GDF6 expression (black bars) results in shorter skull and toe bones. Combining these findings, humans lack a gene enhancer associated with the development of long lateral toes.

Figures 5-6 from Indjeian et al. (2016) sum up the findings. Figure 5 (left) shows that the ancestral version of the GDF6 enhancer (blue staining) is most strongly expressed in the lower half of the body, especially the fifth toe bone. Figure 6 (right) shows that a lack of Gdf6 expression (black bars) results in shorter skull and toe bones. Combining these findings, humans lack a gene enhancer associated with the development of long lateral toes.

hCONDEL.306 doesn’t completely turn off GDF6, so this second experiment doesn’t really tell us exactly what the hCONDEL does. But the results are highly suggestive. Indjeian and team showed that Gdf6 affects toe length, among other skeletal traits, in mice. The ancestral enhancer that humans are missing seems to affect GDF6 activity in the leg/foot only. This illustrates a mechanism of modularity – as the authors state, “Loss of this enhancer would thus preserve normal GDF6 functions in the skull and forelimbs, while confining any … changes to the posterior digits of the hindlimb.” In other words, developmental enhancers allow different parts of the body to evolve independently despite being made by some of the same genes (such as GDF6).

As with any good study, results are intriguing but they raise more questions for future studies. The researchers conducted two experiments to investigate the function of hCONDEL.306: first putting the chimp version in mouse embryos to see where the ancestral enhancer is expressed, and then turning off Gdf6 completely in mice to see what happens. A more direct way to see what hCONDEL.306 does might be to put a longer stretch of the human sequence surrounding GDF6 containing (or rather missing) the ancestral enhancer into mouse embryos. I’m not a molecular biologist so maybe this isn’t possible. But this is important because the ancestral (chimpanzee) enhancer appeared to be most strongly expressed in the little toe, but of course this isn’t our only toe that is short compared to chimps. Similarly, how hCONDEL.306 relates to the abductor hallucis muscle remains in question – does it reduce the size of the intrinsic muscle present in both humans and chimps, or does it prevent development of the longer muscle that chimps have but we lack? We can expect to find hCONDEL.306 in the genomes of Neandertals (and Denisovans?), since they also have short toes, but what would it mean if they retained the ancestral enhancer?

So how does this gene tweak help with upright walking?

This is a really cool paper with important implications for human evolution, but something seems to have been lost in translation between the paper and the headlines (the news pieces themselves are good, though). The upshot of the study is that humans lack a stretch of non-coding DNA, which in chimpanzees (or chimp-ified mice) promotes embryonic development of the lateral toes and a big toe muscle. This may be a genetic basis for at least some aspects of our unique feet that have evolved under natural selection for walking on two legs.

But the headlines misrepresent this result, with words like “led to,” “allowed,” and “caused,” especially when these are followed by “big toe” or “upright walking.” hCONDEL.306 doesn’t really have anything to the big toe bone itself, although it might relate to a muscle affecting our big toe. The only sense in which the “Gene tweak led to humans’ big toe” (first title above) is that hCONDEL.306 might be responsible for our short lateral toes, which make our first toe look big by comparison. The other headlines are misleading since we know from fossil evidence that hominins walked upright long before we have evidence for short toes:

These little piggies get none. Fourth toe bones of living apes and humans (left) and possible hominins from 3-5 million years ago (right).

These little piggies get none. Fourth toe bones of living apes and humans (left) and (probable) hominins from 3-5 million years ago (right). I did my best to get all images to scale.

“Epigenetic,” from the fourth article headline, is simply wrong. Modern day epigenetics is a field concerned with the chemical alterations to the structure of DNA. Even the broad concept of epigenetic as originally devised by Conrad Waddington was about how environments (cellular or outside the body) influence development.

ResearchBlogging.orgIt’s hard to fit all the important and interesting information from scientific papers into news headlines. Still, it would be good if headlines more accurately portrayed scientific findings, especially avoiding such definitive verbs as “caused.” Especially in the realm of biology, people should know that there’s a lot that we still don’t know, that there’s lots more important work left to be done.

References

Aiello and Dean, 2002. Human Evolutionary Anatomy. Academic Press.

Capra et al., 2013. Many human accelerated regions are developmental enhancers. Philosophical Transactions of the Royal Society B 368: 20130025.

Indjeian et al. 2016. Evolving new skeletal traits by cis-regulatory changes in bone morphogenetic proteins. Cell http://dx.doi.org/10.1016/j.cell.2015.12.007

King and Wilson, 1975. Evolution at two levels in humans and chimpanzees. Science 188: 107-116 DOI: 10.1126/science.1090005

McLean et al., 2011. Human-specific loss of regulatory DNA and the evolution of human-specific traits. Nature 471: 216-219.

Prabhakar et al., 2008. Human-specific gain of function in a developmental enhancer. Science 321: 1346-1350.

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Osteology Everywhere: Bacon or first rib?

I went to a cafe today to eat breakfast and get some work done. Write, write, write. It’s important to be properly nourished to ensure maximal productivity.

The Ron Swanson diet.

The Ron Swanson diet.

But I was aghast to behold the food they placed before me:

More bacon, please.

What on earth is this?

First of all, this is not a sufficient amount of bacon.

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Secondably, this bacon is a spitting image of a first rib:

First ribs, from left to right: Human, chimpanzee, bacon. First two images from eSkeletons.org.

First ribs from the right side of the body, viewed from the top. From left to right: Human, chimpanzee, bacon. First two images from eSkeletons.org.

At the top of the ribcage, just beneath the clavicle and subclavian artery and vein, the first rib is much shorter and flatter than the rest of the ribs. As Jess Beck at Bone Broke points out, “The first and second rib give something of an awkward ‘slow song at a middle-school dance’ kind of a hug, while the lower ribs provide a more comfortable and self-assured embrace.” I mean, just lookit how sheepishly the bacon dances with the eggs in the first picture, it has ‘middle-school dance’ written all over it.

But the bacon is not totally identical to the human and chimpanzee counterparts. It’s missing their anteromedially sweeping arc, and the distal portion reaching out to the egg is fairly straight. This suggests we’re probably missing much of the original distal end. Posteriorly or dorsally (toward the bottom in the pic), it also appears to be missing much of the lateral portion including the vertebral facet. In this regard, this bacon rib looks a lot like the first rib of Homo naledi:

Full stack of ribs. From left to right: Human, bacon, Homo naledi, Dmanisi Homo erectus, Australopithecus sediba (x2), Australopithecus afarensis specimen "Lucy," Ardipithecus ramidus, and chimpanzee. Images not to scale except Lucy and Ardi.

Full stack of ribs. Left to right: Human, bacon, Homo naledi, Dmanisi Homo erectus, Australopithecus sediba (x2), Australopithecus afarensis specimen “Lucy,” Ardipithecus ramidus, and chimpanzee. Images not to scale except Lucy and Ardi. Image credits given below.

It is hard to make good homologous comparisons among these fossils and bacon, since so many are so incomplete. But it looks like the hominins are relatively longer (front to back, or dorsoventrally) compared to the chimpanzee. That is, oriented along the rib “neck,” the ventral/distal end projects far more medially beyond the proximal vertebral facet in the chimp, while in the hominins the two ends are more flush.  Ardi is really incomplete and so very hard to orient, but it may be more like the chimp (I think it needs to be rotated to the right more, to make the lateral edge more vertical like all the other specimens).

It will be interesting to see what my colleagues working on the Homo naledi thorax have to say about rib shapes and their functional importance, hopefully not too long from now.

Anyway, I really wish I had more bacon.

Fossil rib sources
ResearchBlogging.orgDmanisi Homo erectus: Lordkipanidze D, Jashashvili T, Vekua A, Ponce de León MS, Zollikofer CP, Rightmire GP, Pontzer H, Ferring R, Oms O, Tappen M, Bukhsianidze M, Agusti J, Kahlke R, Kiladze G, Martinez-Navarro B, Mouskhelishvili A, Nioradze M, & Rook L (2007). Postcranial evidence from early Homo from Dmanisi, Georgia. Nature, 449 (7160), 305-10 PMID: 17882214

Australopithecus sediba: Schmid P, Churchill SE, Nalla S, Weissen E, Carlson KJ, de Ruiter DJ, & Berger LR (2013). Mosaic morphology in the thorax of Australopithecus sediba. Science, 340 (6129) PMID: 23580537

Homo naledi: Morphosource.

Australopithecus afarensis and Ardipithecus ramidus: White TD, Asfaw B, Beyene Y, Haile-Selassie Y, Lovejoy CO, Suwa G, & WoldeGabriel G (2009). Ardipithecus ramidus and the paleobiology of early hominids. Science, 326 (5949), 75-86 PMID: 19810190

Osteology everywhere: Graffiti

Astana, the wedding-cake capital of Kazakhstan, is notably bereft of graffiti and street art, at least in my somewhat limited exposure to the city. The larval metropolis is all about commercial appearance, so I’d guess that aspiring street artists likely face much more than the Marge Simpson treatment for turning around to brag about their work.

Dire consequences await those who graffito tag public property.

Dire consequences await those who graffito tag public property.

Once, I did see a pretty badass street mural,

But it was in München.

but it was in München, a mere 2,620 miles from Astana.

No, there is not much in the way of secretly donated street art here in Astana, and there’s generally little hope to see graffiti-grafted Osteology Everywhere. But this weekend, I noticed these four magical letters, quickly quietly scrawled on the side of my apartment building:

2015-01-17 14.34.37

DAKA.

Two disconcerting thoughts immediately come to mind reading this. First, why the hell is “DAKA” written in Latin instead of Cyrillic script characteristic of the FSU? Second, what does “DAKA” mean out here? Nothing in Russian so far as I know, but Google Translate claims it could mean “Dakar” in Kazakh, which if true raises even more questions.

No, the safest assumption is that this tagger, my streetwise and marker-wielding dopplegänger, was referring to the ~1 million year old Homo erectus partial skull from Ethiopia, dubbed “Daka” after the Dakanihylo site of its discovery.

The Daka calvaria (Figure 2. of Asfaw et al., 2002). Counterclockwise from the top left: view from the back, view from the top (front is to the left), view from the left, a mosquito net, view from the bottom (front is at the top), viewed from the front.

BOU-VP-2/66, the Daka calvaria* (Figure 2. of Asfaw et al., 2002). Counterclockwise from the top left: view from the back, view from the top (front is to the left), view from the left, a mosquito net, view from the bottom (front is at the top), and view from the front. *Calvaria is the fancy word for ‘bony skull without a face.’

Daka isn’t the first hominin fossil to be embraced outside of anthropology. A few years ago I noticed the 4.4 million year old Ardipithecus ramidus skeleton strutting across the label of a Dogfish Head beer bottle:

2011-11-12_14-41-32_247

GOODGRIEF, this was almost 5 years ago.

In downtown Tbilisi, Georgia I recently spotted a Dmanisi-based duo whose tech savvy belies the fact they’re based on 1.8 million year old fossils:

20141013_141725

(Let’s not forget this one, from before they got smartphones)

We’ll have to do some serious fossil-finding here in Kazakhstan before they’ll let anyone put up something this awesome on the side of anything here in Astana. (Or wait…)

Results of the toe-tally easy lab activity

Alternate title: Dorsal canting in primate PPP4s

Earlier this year I suggested a classroom activity in which students can scrutinize the evidence used to argue that the >5 million year old (mya) Ardipithecus kadabba was bipedal. To recap: Ar. kadabba is represented by some teeth, a broken lower jaw, and some fragmentary postcrania. The main piece of evidence that it is a human ancestor and not just any old ape is from a single toe bone, and the orientation of its proximal joint. In Ar. kadabba and animals that hyperdorxiflex their toes (i.e., humans and other bipeds when walking), this joint faces upward, whereas it points backward or even downward in apes. This “dorsal canting” of the proximal toe joint has also been used as evidence that the 4.4 mya Ardipithecus ramidus and 3.5 mya owner of the mystery foot from Burtele are bipedal hominins. A question remains, though – does this anatomy really distinguish locomotor groups such as bipeds from quadrupeds?

Use ImageJ to measure the canting angle between the proximal joint and plantar surface. Proximal to the right, distal to the left.

STUDENT SCIENTISTS TO THE RESCUE! Use ImageJ to measure the canting angle between the proximal joint and plantar surface, as I’ve done on this Japanese macaque monkey (they are not bipedal). Proximal to the right, distal to the left Note I changed the measured angle from the March post.

I sicked my students in Ant 364 (Human Evolutionary Developmental Biology) here at NU on this task. I had students look at only 11 modern primates from the awesome KUPRI database. Most groups are only represented by 1 (Homo sapiens, Hylobates lar and Macaca fuscata) or two (Pongo species and Gorilla gorilla) specimens, all adults. For chimpanzees (Pan troglodytes) there is one infant and four adults. The database has more individuals, and it would be better to include more specimens to get better ideas of species’ ranges of variation, but this is a good training sample for a class assignment. The fossil group includes one Ardipithecus ramidus, one Ar. kadabba, one Australopithecus afarensis, and the PPP4 of the mystery foot from Burtele. The human and all fossils except Ar. kadabba are based off of lateral photographs and not CT scans like for the living primates, meaning there may be some error in their measurements, but we’ll assume for the assignment this is not a problem. Here are their results:

Dorsal canting angle of the fourth proximal pedal phalanx in primates.

Dorsal canting angle of the fourth proximal pedal phalanx in primates. The lower the angle, the more dorsally canted the proximal joint surface. The “Fossil” group includes specimens attributed to ArdipithecusAustralopithecus and something unknown.

Great apes have fairly high angles, meaning generally not dorsally canted proximal joint surfaces. The two gorillas fall right in the adult chimpanzee (adult) range of variation, while chimp infant and orangutans have much higher angles (≥90º means they’re actually angled downward or plantarly). The gibbon (Hylobates) is slightly lower than the chimpanzee range. The macaque has an even more dorsally canted joint, and the human even more so. The fossils, except the measurement for Ar. ramidus (see note above), have lower angles than living apes, but higher than the human and the monkey. If dorsal canting really is really a bony adaptation to forces experienced during life, then the fossil angles suggest these animals’ toes were dorsiflexed more so than living great apes (but not as much as the single monkey and human).

This lab helps students become familiar with CT data, the fossil record, taking measurements (students also measure maximum length of the toe bones and look at the relationship between length and canting), analyzing data, and hypothesis testing. You can also have fun exploring inter-observer error by comparing students’ measurements.

Here’s the full lab handout if you want to use or modify it for your own class: Lab 5-Toe instructions and report

Osteology everywhere: Pelvis has left the building

The vernal awakening has brought rain to Ann Arbor, and right on here on main campus I spotted the rain-splotched silhouette of an articulated human pelvis (left).

Check out those short and flaring iliac blades, and the shortness of the ischium. These features are associated with repositioning key muscles for walking and running on two feet, and are very unlike what is seen in the four-legged, suspensory climbing apes.

But just how ‘human’ are these features? The crushed pelvis of Oreopithecus bambolii, a ~8 million year old fossil ape from Italy, has somewhat human-like short ilia (left). This pelvis also has weak anterior inferior iliac spines (Rook et al. 1999), which anchor the hip/trunk flexor muscle rectus femoris, and are allegedly a developmental novelty seen only in hominids (Lovejoy et al. 2009). These traits have led some to claim that Oreopithecus was a hominid, or at least bipedal. Without getting into that debate, I’ll just say that seeing these ‘bipedal’ features in this late Miocene ape’s pelvis weakens the case that their presence in Ardipithecus ramidus indicates a unique connection between Ardi and later, true hominids like australopiths.

UPDATE: Check the comments for notes on the Ardi and Oreo fossils from someone who’s actually studied them (I myself have only seen pictures and read about them).

ResearchBlogging.orgReferences
Lovejoy, C., Suwa, G., Spurlock, L., Asfaw, B., & White, T. (2009). The Pelvis and Femur of Ardipithecus ramidus: The Emergence of Upright Walking Science, 326 (5949), 71-71 DOI: 10.1126/science.1175831

Rook, L. (1999). Oreopithecus was a bipedal ape after all: Evidence from the iliac cancellous architecture Proceedings of the National Academy of Sciences, 96 (15), 8795-8799 DOI: 10.1073/pnas.96.15.8795

ARDIPITHECUS BEER!!!

I just made what what may be the most amazing discovery of the century at a local booze emporium. Dogfish Head brewing company makes a beer whose label is adorned with Jay Matternes’s reconstruction of an upright Ardipithecus ramidus. Note that the left foot grasps the earth with it’s ape-like big toe.

In a whimsical use of artistic license, whoever adopted this image added a curlicue pig’s tail. In animals with a tail, a number of caudal vertebrae continue off the set of fused vertebrae called the sacrum. Humans and other apes don’t have true tails but a coccyx, a small clump of tiny, fused vertebral segments. Our tail vestige may not help us hang onto trees like in Ateline monkeys, or sting our enemies like a scorpion, but the coccyx is still pretty important. In people this evolutionary memory of a tail anchors some muscles of the pelvic floor (including sphincter ani externus and levator ani), which are critical for the to control of our bowels.

Below is a close up of the Ardipithecus ramidus pelvis fossils (from White et al. 2009, fig. 3). No coccyx was discovered for Ardi, and little is said about the sacrum, other than that it’s merely broken piece of the end of the bone (Lovejoy et al. 2009). Nevertheless, I’m sure this end of sacrum would lead one to reject this artist’s hypothesis that Ardipithecus had a tail.

Had I been in charge of labeling at Dogfish Head, the beer would’ve been called “Party-pithecus” instead of “namaste,” and this label would’ve been slapped on some exotic IPA or porter instead of a wheat beer. Still pretty awesome, though.

Learn about Ardi and its pelvis
Lovejoy, C., Suwa, G., Spurlock, L., Asfaw, B., & White, T. (2009). The Pelvis and Femur of Ardipithecus ramidus: The Emergence of Upright Walking Science, 326 (5949), 71-71 DOI: 10.1126/science.1175831

White, T., Asfaw, B., Beyene, Y., Haile-Selassie, Y., Lovejoy, C., Suwa, G., & WoldeGabriel, G. (2009). Ardipithecus ramidus and the Paleobiology of Early Hominids Science, 326 (5949), 64-64 DOI: 10.1126/science.1175802

ResearchBlogging.org

Tess Tossed Tyrone

What’s the secret to becoming a good father? What would William Cosby do?

I for one have no idea BUT! a study published today in PNAS early edition finds an association between studly vs. paternal behavior, and levels of everyone’s favorite hormone, testosterone (T).

Using longitudinal data, researchers (Gettler et al. in press) found that, in general, a young guy with higher levels of circulating T is more likely than a guy with low T to become a father w/in a few years. MOREOVER! this erstwhile-high-T-and-now-father then experiences a relatively sharper decrease in T than would be expected simply because of aging. PLUS! fathers who interacted with their kids on a daily basis had lower T than fathers who didn’t hang around their kids too often.

One thing neat about this study is that it uses longitudinal instead of cross-sectional data.  A cross-sectional version of this study would’ve sampled a bunch of dudes (hopefully somewhat randomly) only once. This can be problematic because it’s then hard to interpret the results in light of the many sources of variation between people. This study, on the other hand, sampled a tonne (n = 694) of guys at more than one occasion, so they can tell how individuals’ testosterone levels tend to change in paternal vs. free-spirited circumstances.

The last line of the paper is pretty intriguing: “[these results] add to the evidence that human males have an evolved neuroendocrine architecture shaped to facilitate their role as fathers and caregivers as a key component of reproductive success.” (Gettler et al. in press: p. 5/6) This is especially interesting in light of the Ardipithecus ramidus-related evidence for a great antiquity of humans’ paternal proclivity (Lovejoy 1981, Lovejoy et al. 2009). Just how and why testosterone responds to/mediates this fatherly ‘reproductive strategy’ is mysterious to me. And of course, linking this hormonal phenomenon with anything as old as Ardi is a challenge I’m certainly not up to. Still neat, though.

ResearchBlogging.org
My personal circulating T levels are consistently through the roof. So in the event that I become a father, it will be interesting to see if the subsequent, astronomical hormone drop, predicted by this study, won’t cause my entire body to collapse in on itself.

Reference
Gettler LT et al. in press. Longitudinal evidence that fatherhood decreases testosterone in human males. Proceedings of the National Academy of Sciences… doi: 10.1073/pnas.1105403108

Lovejoy, C. (1981). The Origin of Man Science, 211 (4480), 341-350 DOI: 10.1126/science.211.4480.341

Lovejoy CO (2009). Reexamining human origins in light of Ardipithecus ramidus. Science (New York, N.Y.), 326 (5949), 740-8 PMID: 19810200

Photo credit: google (image) “Bill Cosby Fatherhood”