A fly used as a model for fishing lures has led James
H. Marden, assistant professor of biology at Penn State, to a new theory
of how flight evolved in insects. His recent studies of the stonefly reveal
its ability to skim across the water surface on its feet-like a Florida
airboat equipped with pontoons. This behavior could be an ancient form
of locomotion that fostered the development of large muscles and other
factors necessary for full airborne flight.
Many fly-fishing lures look like the stonefly nymph-a prime food object for fish. "My interest in fishing had me thinking about the insect behaviors mimicked by fishing lures, then that interest evolved into a curiosity about the evolution of flight," Marden recalls. He says he was in a stream in winter teaching an undergraduate ecology class when suddenly he noticed stoneflies skimming along on the surface of the water. "I had been looking for something like that for a long time and I knew right away that it could be how insect flight evolved," he says.
The evolution of insect flight has long been one of the great mysteries in evolutionary biology. Scientists generally agree that wings evolved from the gills of water-dwelling species about 400 million years ago, but until now they have lacked a convincing explanation of how flying insects could have evolved from these nonflying swimmers.
"The dominant theory is that flying evolved from gliding, but that didn't make any sense to me because most insects just don't glide," Marden says. In addition, Marden says a glider's wings have to be rigid but all flying insects flap their wings. "A flying insect needs to have sophisticated joint articulation, a sophisticated neural pattern, and a large proportion of its muscle mass specialized for flapping-how could all that have evolved in an insect specialized for holding its wings horizontal and perfectly rigid?" he asks.
Instead, Marden proposes a direct evolutionary route from swimming to true flying. With the stonefly to demonstrate his theory, Marden has shown how surface skimming could be an intermediate stage in the evolution of strong flapping wings. Stoneflies, and their close cousins the mayflies, are thought to be almost like living fossils. "They are the lineages in which the ancestral traits are most similar to the first flying insects," Marden explains.
In late winter, wingless stonefly nymphs come out from under rocks on the bottom of streams throughout eastern and central North America, float up to the surface of the water, migrate toward the shore, and emerge there from the nymph stage into adults with wings, where they live for the rest of their lives without ever flying. But sometimes when a stonefly emerges on sticks or ice floating in the middle of a stream it must get to land by using its new wings to send it skimming across the surface of the water. Marden says this is the only time in its life that these stoneflies are known to flap their wings. "The behavior probably is maintained for periodic episodes of flooding when all the stoneflies would have to surface skim to the shore and survival of the entire group would depend on surface-skimming ability," he explains.
Because stoneflies emerge in the winter when trout are cold and sluggish, they can get away with surface skimming, which in warmer months would instantly attract lively and hungry fish. "People who fish are more aware of the stonefly's cousin, the mayfly, which emerges in warmer months and has to take flight right away to get away from the fish," Marden says.
Marden tested his theory by videotaping stoneflies brought to the laboratory by undergraduate student Melissa G. Kramer, a biology major and coauthor with Marden of a paper describing the research in the October 21, 1994, issue of the journal Science. "When they were emerging in the field I could collect 50 in 15 minutes because they were just crawling all over-they really stand out against the snow," Kramer says.
Marden and Kramer put individual stoneflies on a dish of water and videotaped them as they flapped their wings and skimmed across the surface. "They do it every time you put them on the water," Kramer says. They learned that skimming velocity improved with increasing wing size, flight muscle size, temperature, and body size. "The fastest ones reached speeds as high as 44 centimeters-or a little over 17 inches-per second," she says.
The biologists clipped the wings of some of the insects and found that even very short wings worked well for surface skimming. "The nymph's gill plates have the neuromotor pattern, the complex articulation, and the muscles for moving fluid," Marden explains, "so they just need bigger gill plates and bigger muscles to go from moving water to moving air-not that huge an evolutionary jump."
Marden and Kramer studied the feet and wings of the insects under a scanning electron microscope and discovered they are covered with hairs that look somewhat like miniature ice skates, giving a water-resistant coating to those body parts. "If you've ever had to clean a swimming pool you know what happens to insects whose wings get stuck to the water," Marden comments. Stoneflies are able to lift their water-resistant wings from the water, raising their body high above it on their long legs. The hairs on their feet help stoneflies to surface-skim by reducing their contact area with the water and its resulting surface tension.
Marden says he plans to analyze the water-resistant hairs on the feet and wings of both stoneflies and mayflies to see if their amino-acid or gene-sequence data demonstrate that they had a common ancestor. "Many scientists already think the common ancestor of these two groups was the first flying insect," he says. If the analysis shows this ancestor also had wet-resistant wings and feet, the scientists could infer that it was a surface skimmer and that surface skimming could be the evolutionary bridge to insect flight.
"We have tested our theory by demonstrating an intermediate stage between nonflight and flight that has an important survival function," Marden says. "The theory becomes much more interesting when you see an insect that might be a living fossil actually demonstrating it."
Here are two Stonefly QuickTime Movies (3.7 & 2.2 Mb) cited in our recent Science paper (Marden & Kramer 1994). They are platform-independent but require QuickTime movie player software on your machine for viewing. These movies originated as video shot with a Kodak Ectapro system at 500 frames per second. Elapsed time is displayed in a panel on the lower right side of the screen.
Barbara K. Kennedy"We analyzed in greater detail than before how these new HIV inhibitors work, so we now know exactly how they block the replication of the HIV-1 virus," says Kenneth A. Johnson, Paul Berg Professor of Biochemistry at Penn State and principal author of a paper describing the research to be published in a recent issue of the journal Science.
In order to survive, HIV must transform its genetic material, RNA, into the form of human genetic material, DNA. This transformation, the replication process, is carried out by an enzyme of the virus, reverse transcriptase. The resulting HIV-derived DNA can then work its way into the DNA of a human cell, where it is able to reproduce. Reverse transcriptase is the target of many drugs used to fight HIV.
The reverse transcriptase enzyme works like a factory assembly line, building the twisted zipper shape of DNA one piece, or base, at a time in a series of coordinated steps. It inserts its genetic code into the new DNA by using a strand of its own RNA as a template to which it attaches DNA's chemical building blocks, nucleoside compounds that it finds within the human cell.
After it grabs onto one of these building blocks, the enzyme clamps down around the growing DNA strand, maneuvering the nucleoside close to other compounds, causing a chemical reaction that adds a new piece to the growing strand of DNA. Drugs now used to fight HIV, such as AZT and ddC, work by locking into a site on the reverse transcriptase enzyme where the nucleoside compounds normally attach.
"The problem with drugs that mimic nucleoside compounds," says Johnson, "is they are very toxic." These drugs attack not only reverse transcriptase but also kill the cell's essential mitochondria, which the body needs to convert food to energy. "Its toxicity ultimately limits how much AZT a person can stand," Johnson says.
Johnson and his research team studied a new class of drugs that are not nucleoside mimics. "Nevirapine is currently being used in clinical trials and is the most effective of the nonnucleoside drugs discovered," Johnson says. "It has the advantage of being very potent and very specific for HIV-1 reverse transcriptase, so it can be given in much smaller doses, and it doesn't kill mitochondria."
The Penn State researchers used instruments they designed and built that allowed them to do studies that hadn't been done before to measure the very beginning of key reactions on a millisecond time scale.
Johnson's team studied three of the new nonnucleoside-type drugs and discovered that they attack HIV by binding to a site on the reverse transcriptase enzyme that is different from the nucleoside binding site. "Although the site where the new nonnucleoside drugs bind to the reverse transcriptase enzyme was previously known, their mechanism of action was not understood," Johnson says. "We found that the binding of nonnucleoside drugs at this site interferes with the enzyme's ability to properly position the chemical reactants."
Normally, reverse transcriptase would clamp down to force the reactants together into a very close space with exactly the right orientation so that a chemical reaction would occur, forming a new base. Then it would release and move on to build the next base, Johnson explains. But with the nonnucleoside drug attached, the enzyme cannot align the key catalytic compounds properly. Reverse transcriptase closes down but when the chemical reaction does not take place the enzyme does not move onto the next step of releasing its grip. The drug stalls the DNA-production process by freezing the reverse transcriptase enzyme in a stranglehold, binding the nucleoside even tighter.
Because the AZT-type and Nevirapine-type drugs attach to different parts of the reverse transcriptase enzyme, researchers might be able to fuse the two together to make more powerful and less toxic compounds. "If we could design a reverse transcriptase inhibitor so that part of it would bind in the nonnucleoside site and part of it would bind in the nucleotide site, we would possibly have a drug that binds tighter than either of them would alone," Johnson says. "We might be able to make a very specific inhibitor of reverse transcriptase to overcome some of the problems of toxicity and mutation-related drug resistance at each site."
The Johnson lab's research was sponsored by the National Institutes of Health.
Barbara K. Kennedy
According to William Dunson, Penn State professor of biology, the burning of coal and municipal waste in Pennsylvania deposits a lot of metals and toxic chemicals into the atmosphere, making the state an ideal model for the study of acid and metal deposition in the temporary forest ponds the salamanders use for breeding in early spring. "Because Jefferson salamanders are one of the most sensitive amphibians in this habitat, they are an excellent bioindicator of serious environmental damage," Dunson says.
Meager mating results have left much of Pennsylvania completely barren of the Jefferson salamander species, according to Dunson and his research team, which has just completed a 10-year study of the animal and its early-spring mating ponds. "This is one of the best long-term studies known in the entire world that shows convincing evidence of pollution affecting a population of amphibian species," Dunson says.
Dunson and his research team sampled over seventy temporary ponds in Pennsylvania, which he says receives some of the most acidic rain in the country. With a pH of 4, it is 40 times more acidic than normal rainfall. In Pennsylvania, the ponds are covered with ice and snow until early March, when melting begins and the salamanders migrate from the surrounding forest into the ponds to start breeding. "Sometimes we have to chip ice away to get into the ponds because the tops have refrozen," Dunson says. Equipped with four-wheel-drive vehicles, the biologists would drive as far as they could into the forest and then hike a couple of miles farther to reach the ponds, where they collected the salamander egg masses to take back to the laboratory.
The researchers looked at multiple variables when analyzing the breeding sites of the Jefferson salamander, including pond pH and metal concentration. They transplanted Jefferson Salamander embryos from the ponds to the laboratory where they could change the pH of the water and monitor the salamanders' progress. "Once you remove low pH-or high acidity-as a potential toxin, metals become the primary toxins in the pond," Dunson explains.
The research indicates that metals like aluminum, copper, lead, and zinc, which enter freshwater ecosystems either by rain or watershed leaching, could be responsible, in part, for the reproductive failure of the Jefferson Salamander. "Groundwater with a low pH extracts metals from the soil, allowing them to enter ponds and streams, where they bind to the gills of amphibians and fish causing respiration and other physiological problems, even death," states Dunson. "There is a direct correlation between low pH and high aluminum levels, and our field study revealed that aluminum concentrations were significantly higher in the ponds lacking successful breeding of the Jefferson Salamander," he says.
"We now need to broaden the approach and look at what's happening in other habitats in the surrounding forest as well," says Dunson, who recently was awarded a major grant by the Environmental Protection Agency to study atmospheric effects of pollution on other forest species.
Dunson's 10-year study of Pennsylvania salamanders was funded primarily by grants from the Environmental Protection Agency.
Shorna R. Broussard '94 ERM science-writing intern, fall semester 1994
The research also explores important similarities with and differences from another known motor protein, myosin, which powers movement in muscles.
Kinesin is thought to be important for the regeneration of severed nerves and could be involved in conditions such as Parkinson's and Lou Gehrig's diseases that are caused by the degeneration of nerve cells.
The Penn State research shows that kinesin transports neurotransmitters and other chemicals produced by the nerve cell along microtubule strings strung through the long arm, or axon, of nerve cells. Some of these microtubules can be up to a meter long. "There are single nerve cells that start out in the base of the neck and go all the way out to the fingertip," says Kenneth A. Johnson, Paul Berg Professor of Biochemistry at Penn State and leader of the research team. "Everything that is needed at the tip of the finger from that nerve is synthesized in the cell body and bundled into packages that kinesin carries all the way to the axon tip," he says.
According to Johnson, the research is the first to show exactly how kinesin uses chemical energy stored in the compound ATP to fuel transportation of the neurotransmitter-containing packages.
Johnson says the kinesin molecule looks like a snake with two heads. "The kinesin motor has both a microtubule binding site and an ATP binding site on each of its heads, and these sites interact in a way that leads to the production of force and movement," he explains.
Johnson's lab developed new high-speed tests to reveal how the molecule's two heads take turns attaching to and detaching from the microtubule to move neurotransmitters from the nerve-cell center to the tip of the axon. "Our methods allowed us to make measurements over time scales of milliseconds with very small amounts of material so that we could examine the reactions occurring during each step in kinesin's force-production cycle," he says.
Johnson explains that when ATP attaches to a kinesin head it undergoes a chemical transformation to the chemical compound ADP plus phosphate. The resulting energy, and the structural changes this chemical reaction produces, together provide the force that powers kinesin's race along the microtubule with its neurotransmitter cargo attached to its tail. "If you drew it to scale, it would look like an ant carrying a basketball along a tightrope," Johnson says.
The research solved a related puzzle that had until now baffled researchers. They wondered how a single kinesin molecule could move its cargo so smoothly and efficiently while muscle movements require the coordinated activity of many myosin molecules, the biological motors in muscle cells.
Johnson's sensitive experimental measurements answered both questions by revealing that "kinesin is 100 times slower to come off and 2,000 times faster to rebind" to its microtubule than is the muscle motor, myosin, which pulls against muscle filaments. In other words, after kinesin produces a prolonged force against the microtubule, it lets go-but then rapidly reattaches and exerts a prolonged force again. Myosin, on the other hand, pulls for a shorter time then lets go right away, spending a lot of time detached.
Johnson says the difference makes sense in terms of the biology of the two systems. "Kinesin is an isolated worker rapidly carrying a single large package, while many myosin molecules are pulling together but at random rates against the same filament to make a muscle contract." Johnson explains the myosin molecule has to let go quickly so it doesn't impede the uncoordinated efforts of its coworkers.
According to Johnson, the research shows that although kinesin's two heads are identical to each other when they are in solution, there is some interaction between the two when they are next to each other on the microtubule. "Because the heads take turns detaching, kinesin is always in contact with the microtubule, which explains its remarkable persistence in running along the entire length to the very end," he says.
"The underlying principles revealed in this study apply to a lot of systems," Johnson explains, noting that there are about 20 different kinesin-like molecules involved with a variety of movements within and between cells. He says one of them, a protein called NCD, "has mutations that seem to be related to changes in the rate of cell division, which has implications for diseases like cancer."
Johnson's research was supported by the National Institutes of Health.
Barbara K. Kennedy
"Mixing individual samples together collected from the same polluted site and then analyzing these composited samples will lower regulation costs, because fewer overall samples will be examined in costly laboratory procedures," says Glen Johnson, a doctoral researcher in Penn State's Center for Statistical Ecology and Environmental Statistics. He is a former scientist in Pennsylvania's Department of Environmental Resources.
"This composite sampling will still provide an accurate assessment of environmental or public health risk from pollutants," Johnson says.
Chemical analysis of individual samples is the costliest aspect of pollution monitoring programs.
"Because monitoring for chemical pollutants is so expensive, a lot of tax and private dollars are spent without much actual site cleanup," Johnson says. He worked on applying composite sampling along with colleagues Ganipati P. Patil, distinguished professor of statistics and director of the Penn State Center for Statistical Ecology, Charles Taillie, research associate, and Sharad Gore, professor of statistics at the University of Poona in Pune, India. Patil, Tallie, and Gore conducted several studies on combining individual samples before laboratory analyses. "The key is to know the right time to use composite sampling," he says. "Although it cuts costs, you don't use it if it's going to produce biased estimates or ruin the integrity of the samples."
For example, the technique won't work if mixing samples changes their chemical composition or dilutes contaminated samples with clean ones to the point where pollutants can't be detected.
The researchers call for continued testing of composite sampling and other innovative sampling methods to achieve what they call "observational economy," or the most cost-effective way to determine the risk from harmful chemicals or other substances.
One of their papers on sampling, prepared by Johnson and Patil, will be published in the proceedings of the Air and Waste Management Association's International Specialty Conference, "Cost Efficient Acquisition and Utilization of Data in the Management of Hazardous Waste Sites."
Much of the center's research also appears in a series of papers and reports on composite sampling prepared for the Environmental Protection Agency.
Scott Turner
"We are struggling to understand how our images fit into the general picture of quasar creation and evolution," says Donald P. Schneider, associate professor of astronomy and astrophysics. Schneider made the observations along with his colleagues John Bahcall and Sofia Kirhakos of the Institute for Advanced Study at Princeton University.
Since their discovery in 1963, quasars, or quasi-stellar objects, have fascinated astronomers because they emit prodigious amounts of energy and luminosity from a very compact source. "They are 100 to 1000 times more luminous than our entire galaxy but no bigger than our solar system," Schneider says.
Before Schneider's observations, the most widely accepted model had been that quasars consist of a supermassive black hole-as massive as 100 million to 1 billion Suns-devouring an exceptionally luminous and massive but otherwise normal galaxy. "We think it takes about one solar mass per year to fuel a quasar," Schneider says. He explains that violent shocks from collisions between different streams of star material as they careen into the black hole are thought to emit the intense light characteristic of quasars. However, confirming this model had been difficult because Earth's atmosphere blurs a quasar's intense light over a broad area, obscuring astronomers' view of the stars in the suspected host galaxy. With the Hubble Space Telescope, Schneider's research group thought they could get a better idea of what the environments of quasars are like. "Using the Hubble Space Telescope can improve angular resolution by a factor of 10 over what we can do from the ground," Schneider explains. He says his group's study is the largest and most systematic quasar survey done with the Hubble Space Telescope. While others have studied individual quasars, Schneider's team selected a sample of about 20 of the brightest and nearest quasars-all about 50 times as bright as our Milky Way galaxy and several billion light years from Earth-to study with the Hubble's Wide Field Planetary Camera 2. "We wanted to understand quasars in general, not just specific cases, so we could make statistically valid statements about quasar environments," he explains.
Instead of finding the bright host galaxies most astronomers expected, Schneider says, "We were astonished to see that our first eight quasars seemed to have almost nothing surrounding them." Subsequent observations of six quasars revealed an amazing diversity. "Some environments look like ordinary spiral galaxies except they are much brighter, some look very bizarre and distorted-as if they are being ripped apart by tremendous gravitational forces, and some quasars appear to have very little at all around them.
The astronomers found additional companion galaxies associated with many of the quasars they surveyed-many more than would be expected in a random sample, Schneider says. "It appears from the Hubble images that quasars tend to have galaxies nearby. These companions typically are separated from the quasar by approximately 30,000 light years, which is the distance from the Sun to the center of the Milky Way. These observations provide a new challenge for theorists because current models do not predict the variety of quasar-galaxy interactions unveiled by the new images. "This is the most enigmatic data I ever have analyzed, and it is much too early to know what the final conclusions will be," Schneider says. In addition, astronomers are wondering where the apparently naked quasars are getting the fuel to power their luminous glow. "We can't yet say for sure there is 'nothing' there," Schneider says. "We have to ask whether something really is there but it is just too faint for the Hubble to see." Schneider's team plans to observe some of their apparently "naked" quasars for longer periods in an attempt to get a better look.
The team described their research at the 185th meeting of the American Astronomical Society in Tucson, Arizona, earlier this year.
The Hubble Space Telescope is a project of international cooperation between the National Aeronautics and Space Administration (NASA) and the European Space Agency (ESA). In addition to Donald Schneider, other Penn State astronomy faculty who have had observing time on the Hubble Space Telescope include Matthew Bershady, Robin Ciardullo, France Cordova, George Pavlov, and Lawrence W. Ramsey.
Barbara K. Kennedy