EN | Sagascience - Karsenti

He heard the call of the sea as a young boy, and was taking sailing lessons even before he became a teenager. Although he was born in Paris in 1948, Eric Karsenti is a sailor at heart. When it came to choosing a discipline, he first leaned toward oceanography or marine biology. But the director of the Concarneau Marine Biology Station steered him toward genetics. After studying biochemistry and genetics in Paris, he began his professional life at the Institut Pasteur’s immunocytochemistry laboratory, working under the supervision of the French immunologist Stratis Avrameas. But his passion for the sea continued unabated: in 1979, while completing his PhD thesis in cellular biology and immunology, he gave sailing lessons in Brittany.
Karsenti was already riding high. Having joined the CNRS three years earlier, he was sent to San Francisco

in the early 1980s to work as a postdoctoral fellow at the University of California, investigating the cell cycle as part of a team led by Marc Kirschner. The young researcher conducted various experiments to determine the respective roles of centrosomes (the organelles that organize the microtubules), chromosomes (which carry genetic information) and cytoplasm in the formation of mitotic spindles of microtubules, the mechanism that enables chromosomal migration during cell division. At the time, the dynamics of the cell cycle and mitosis had already been clearly described but completely understood, especially at the molecular level. Based on his experiments, Karsenti proposed a new hypothesis according to which chromosomes play an essential role in the organization of the microtubules, forming the mitotic spindle between the two poles of the cell.

VOYAGE INSIDE THE CELL – MITOSIS

Credits

“In the 1970s, we didn’t know much about the clock-like factors that regulate the cell cycle

from one division to the next and trigger mitosis,” Karsenti recounts. “We knew that the microtubules, the tiny protein tubes that form the skeleton of the cell, are able to organize themselves just before division into a structure called the mitotic spindle. By pulling on the chromosomes, which by then have been duplicated, the microtubules in the spindle separate the two sets of homologous chromosomes and position them at the two poles of the spindle, enabling the formation of two daughter cells. Back then, the scientific community thought that the centriole,

a specific cellular structure found at the poles of the spindle, induced the assembly of microtubules and the bipolarity of the spindle in mitosis. With Marc Kirschner, we used centrioles isolated in test tubes by Tim Mitchison and discovered that even without them, the assembly process still took place. We inferred that, rather than the centrioles, it was the chromosomes themselves that triggered the process. It was so unexpected that no one believed us at first—nor for some time afterwards! As always in science, we had to work very hard to accumulate enough experimental evidence to confirm that initial observation,” Karsenti adds.

In 1985, the biologist took off for Heidelberg, where he joined the department of cell biology at the European Molecular Biology Laboratory (EMBL). There he continued his Californian research as part of a new team, with a new goal: to identify the factor(s) that cause chromosome condensation, a vital phase in mitosis. Except during the brief period of division, the cell’s DNA does not come in the form of chromosomes, usually represented as Xs and Ys. Most of the time, the DNA strands are unraveled, forming a mass at the center of the cell’s nucleus, like thousands of unwound thread reels tangled up on a huge rug. It is not possible to separate the threads into two perfectly equal bunches without rolling them up first. The same thing happens within the cell: before being divided into two batches, the chromosomes’ DNA has to be “rewound”. The question is: which protein is responsible for this tedious process?

MITOSIS AND SPINDLE

Credits

MICROTUBULES

Credits

chromosome

In parallel, a team led by the British genetician Paul Nurse had identified the protein pair responsible for the phenomenon: kinase cdc2, an enzyme that activates other proteins by adding phosphate, and cyclin, which activates the kinase. The latter, in turn, activates proteins responsible for the condensation of DNA into chromosomes and the assembly of the spindle. For this breakthrough, Nurse, along with Leland Hartwell and Timothy Hunt, was awarded a Nobel Prize in 2001. “ At the same time,” Karsenti comments, “Marcel Dorée had begun purifying the kinase from Xenopus (Xenopus is a genus of frogs whose eggs, produced in large quantities, are used as a biological model in the laboratory) eggs in my lab at the EMBL. He completed the project with a student in Montpellier. Personally, I think he should have been named co-winner of the Nobel.”

In Heidelberg, Karsenti and Dorée follow in Nurse’s footsteps, showing that the kinase remains inhibited until the cyclin concentration reaches a certain threshold, and that it deactivates spontaneously after ten minutes.

karsenti_group

“Given the autoinhibition of the kinase activity,” Karsenti continues, “I started thinking about other instances of self- organization in biology. While working with my students on the mitotic spindle during that period, I realized that a phenomenon of this type was taking place: at a certain point in the cell cycle, ‘motors’ linked to the microtubules adopt a collective behavior and organize these small building blocks into a spindle.

From microscopic chaos emerges an ordered, quasi-macroscopic structure. It’s fascinating!”

By the end of the 1980s, the mystery surrounding the universal molecular motor of the cell cycle was about to be solved.

THE CLOCK IN EVERY CELL

Each living cell is equipped with a clock-like mechanism that controls its cycle, as demonstrated by Eric Karsenti and his teams over years of experiments.

« Already at the University of San Francisco, I started ‘breaking’ Xenopus eggs in 1981-82, isolating the cytoplasm for in vitro experiments, » the researcher recounts. In Heidelberg, with the help of my first student, Marianne Felix (now a senior researcher at the CNRS), I used this method to examine how the kinase/cyclin complex was regulated over time.”

The two scientists noticed that the complex did not become active immediately upon entry into the cytoplasm. Only after a series of reactions would the cyclin reach the critical threshold, after gradually accumulating. This latency time is of great importance: it enables the complex to autoactivate at once, rather than gradually, inducing a “clean break” in the form of mitosis

Once activated, the complex remains active for 10 minutes. After that, the cyclin level drops sharply, before gradually building up again0for the next cycle. “We were able to demonstrate that the kinase/cyclin complex itself causes this drop,” Karsenti explains. “If cyclin alone is inserted into our ‘broken eggs’, its concentration remains constant. But if active kinase is inserted into in an egg containing cyclin, the latter disappears after 10 minutes. The kinase triggers a second clock that determines the moment of its own inactivation. In other words, the genetic products of the cell cycle are generated in the proportions needed for the self-organization of a biological clock, enabling an alternation between interphase and mitosis —in Xenopus eggs, every 30 minutes on the dot. It’s amazing!”

THE MOTORS
THAT DRIVE MITOSIS

In the lab, Eric Karsenti’s team has shown that without the action of certain molecular motors, proteins that use the cell’s energy to move along the microtubules, the mitotic spindle cannot form properly.

But to understand how the protein motors, microtubules and chromosomes behave collectively during the assembly of the spindle, the biologist relied on digital simulations0performed in cooperation with physicists.

Using this tool, “We understood that during mitosis, the motors bind to the microtubules,” Karsenti reports. “By forming bridges between them, and by all moving in the same direction, toward the two poles of the cell, they rearrange the microtubules in a spindle around the chromosomes.”

These same microtubules induce the separation of the chromosomes, and grow shorter with the help of the motors, exerting a pull on the chromosomes. Biologists call this depolymerization, and liken the process to a necklace losing its beads. Thus pulled apart, the X chromosomes are cut in half lengthwise, with each half going to one of the daughter cells.

“This is another case of self-organization,” Karsenti concludes. “The collective behavior of the chromosomes, microtubules and motors induces the formation of a ‘machine’, the spindle that separates the chromosomes. We are beginning to understand how complex cellular functions emerge from what seems to be chaos.”

Meanwhile, in 1988, Eric Karsenti embarked on a new adventure. In Roscoff (Brittany), he initiated the first Jacques Monod conference on the cell cycle, with a view to fostering interdisciplinarity — a principle that would guide his entire career. Hosted by the CNRS and INSERM through the Institute of Biological Sciences (INSB) and the Institute of Ecology and Environment (INEE), the conference series continues to this day, with four to six events scheduled each year, most often in Roscoff. Discussions focus on new topics in the life sciences, especially those that involve0several disciplines.

In 1996 Karsenti was named director of the EMBL department of cell biology, taking over from the Finnish biologist Kai Simons. His main goal was to promote interdisciplinarity within a structure that he found “ a bit too 20th-century”. He took on the challenge with enthusiasm, encouraging cooperation among researchers from different fields and renaming his unit the « department of cell biology and biophysics » . Karsenti insits: “If we really want to understand a phenomenon like self- organization in the cell,we need to work with physicists and statisticians to quantify it —and imaging specialists to observe it better.

We must pool our know-how!”

While he had planned to remain in Germany for three years, he finally stayed two and a half decades. “My wife moved back to France after ten years,” he says. “Having a long-distance family life wasn’t easy, but we managed. I came home regularly, and she was very understanding!”

Between 2001 and 2003, Karsenti seemed to be on all fronts. He moved back to Paris to head the Institut Jacques Monod but pursued his EMBL projects. In parallel, he became an advisor to Elisabeth Giacobino, senior researcher at the French Research Ministry. Yet he still had a dream: it was during that period that he hatched the idea of a marine expedition around the world, after reading Charles Darwin’s account of his voyage on board the Beagle. Karsenti’s dream came true with the Tara, a polar schooner that was given a second life as a floating laboratory. In 2009 the Tara Oceans expedition, with Karsenti as scientific director, set sail after a full year of preparation. The ship covered 140,000 kilometers, sailing the Mediterranean, the Red Sea, the Indian, Pacific and Atlantic Oceans, all the way to Antarctica.

“Tara Oceans was a new adventure for me,” the researcher relates.“At 60, I felt the need to change perspective, to address fascinating but troublesome questions about the past evolution of our planet and its imminent future, in which the oceans are crucial. Not only did life on Earth originate in the oceans, but it still largely depends on them. To improve our understanding of the key role of microscopic marine life, I built up a multidisciplinary team of world-class researchers.

I also wanted to reach out to others, to share the adventure of advancing the scientific understanding of our universe.”

On board the Tara, biologists worked alongside engineers, computer scientists, oceanographers, quantitative imaging experts… and even artists! According to Karsenti, “that was the strength of the expedition,” as it enabled its participants to make invaluable discoveries concerning plankton, which—albeit invisible—store 50% of the planet’s CO2. The Tara had a dual goal: to understand the functioning and diversity of marine life, but also to forecast marine ecosystems’ response to climate change. Managing logistics was a daunting task, with 126 researchers, 70 crewmembers plus artists and journalists from 35 countries taking turns on board, nearly 200 sampling stations, 53 oceanographic missions across the globe and the necessary requests for permission to drop anchor and take samples in territorial waters.

Everything was skillfully handled by Tara Expeditions, with the help of one of Karsenti’s collaborators, S. Kandels-Lewis. Based on the samples collected, the researchers have already characterized all types of viruses, bacteria and small eukaryotes from the first stations visited. In the latter category, they have identified some 150,000 genetic types, of which only 10,000 were previously known. Contrary to prior belief, the eukaryote population is much more diverse than that of bacteria. So far, the Tara project has characterized nearly 40 million bacterial genes, most of them unknown to this day —the most extensive such biological census ever undertaken. Even in the 21st century, scientists don’t always know what they are going to find.

ITINERARY OF THE TARA OCEANS EXPEDITION

© Tara Oceans

LEGEND

THE TARA OCEANS EXPEDITION (2009-2013)

© Tara Oceans

“We discovered that the ‘Agulhas rings,’giant whirlpools 400 kilometers in diameter and 4,000 meters deep that cross the Atlantic from east to west, trapping organisms along the way, eventually create a new ecosystem that evolves independently from the external environment,” Karsenti reports. “This is yet another self-organization phenomenon in which small entities adopt a collective behavior to form a larger-scale structure”—and yet another experience in which the seafaring biologist combined his dedication to the life sciences with his passion for the ocean. In late March 2012, the Tara returned to its home port, in Lorient

on the Brittany coast. It was followed by the Tara Oceans Polar Circle expedition, which mapped the biodiversity of Arctic plankton between May and December 2013 and closed the final chapter in this maritime and scientific Odyssey. But the adventure of Tara Oceans was not over yet: all of the samples gathered around the world still needed to be analyzed. After 938 days at sea, it took until May 2015 for Tara Oceans to release its first results. From the ship to the lab, the mammoth task of interpreting the data has only just started.

IMAGES OF PLANKTON

Epicancella nervosa, a unicellular eukaryote found among the zooplankton collected in the southern Pacific during the Tara Oceans expedition (station 98).

Cyttarocylis brandti morph. of Cyttarocylis ampulla, a unicellular eukaryote found among the zooplankton collected in the southern Pacific during the Tara Oceans expedition (station 111).

This specimen has the structure of a C. brandti morph. shell as well as the rounded shape of C. eucecryphalus morph.

Phronima sp, an amphipod crustacean (a form of zooplankton) whose fierce appearance inspired the monster in the film “Alien”. Its enormous eyes and four bright red retinas

give the phronima panoramic vision. This one comes from the Observatoire Océanologique in Villefranche-sur-Mer.

A crustacean of the order Euphausiacea (superorder Eucarida) collected by the Tara in the Indian Ocean.

The order includes krill, a key link in the marine food chain.

Platynereis dumereii, marine flat worm, collected by Tara in 2011, in the Indian Ocean. © Eric Roettinger, KahiKai/Tara Oceans

An elephant-shaped pteropod about 5 mm long, two copepods (left) and an ostracod (the orange creature on the right) netted by the Tara in the Indian Ocean

off the coast of the Maldive Islands. The orange fragment in the far lower left is a chip of paint from the Tara’s hull.

whose prominent heads have arms, or tentacles. The best known are octopuses and squid.

A cephalopod larva collected by the Tara in the Indian Ocean. Cephalopods are mollusks

A crustacean larva collected by the Tara in the Indian Ocean.

A copepod of the zooplankton genus Sapphirina. Planktonic copepods play an important role in both the marine food chain and the carbon cycle.

Specimen collected by the Tara in the Indian Ocean.

Ctenophores are a phylum of carnivorous plankton, which swim using eight comb-like rows of cilia.

This juvenile specimen was collected in the Mediterranean

Common examples of zooplankton: on the left, three crustaceans (a copepod, a spider crab larva and an amphipod); in the middle, a baby octopus;

on the right, an amphipod of the Phronima genus and an Atlanta pteropod mollusk.

Plankton collected in the Pacific using a 0.1 mm mesh net. Shown here, a mixture of multicellular organisms

—small animals and zooplankton larvae—and unicellular protists, including diatoms, dinoflagellates and radiolarians.

Plankton collected in the Mediterranean during the winter using a 0.2 mm mesh net, composed of a mixture of protists and zooplankton (animals and larvae).

Marine diatoms collected during the Tara Oceans expedition. Unicellular algae with an external silica structure, diatoms make up the largest group of phytoplankton algae.

A protist: the dinoflagellate g.Pyrocystis. Dinoflagellates are aquatic microorganisms with flagella. Some are photosynthetic.

Specimen collected by the Tara in the Indian Ocean.

A diatom (protist). Unicellular algae with an external silica structure, diatoms make up the largest group of phytoplankton algae

Specimen collected by the Tara in the Indian Ocean.

Planktonic protists, collected by the Tara.

An Acantharea (protist). Specimen collected by the Tara in the Indian Ocean.

PLANKTON CHRONICLES

Credits

Throughout his career, Eric Karsenti has been a pioneer in interdisciplinary approaches to cellular biology. Today, along with Sébastien Colin, he chairs the Morphological Data work group of the Oceanomics project. Providing the scientific follow-up to the 2009 first Tara expedition coordinated by CNRS senior researcher Colomban de Vargas, the group is in charge of collecting, analyzing and archiving the morphological descriptions of the plankton communities sampled during the Tara Oceans expedition. For this researcher who has devoted his professional life to unraveling the secrets of the infinitely small, the adventure continues.

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EN

VOYAGE INSIDE THE CELL – MITOSIS

MITOSIS AND SPINDLE

MICROTUBULES

THE CLOCK IN EVERY CELL

Each living cell is equipped with a clock-like mechanism that controls its cycle, as demonstrated by Eric Karsenti and his teams over years of experiments.

THE MOTORS THAT DRIVE MITOSIS

In the lab, Eric Karsenti’s team has shown that without the action of certain molecular motors, proteins that use the cell’s energy to move along the microtubules, the mitotic spindle cannot form properly.

But to understand how the protein motors, microtubules and chromosomes behave collectively during the assembly of the spindle, the biologist relied on digital simulations0performed in cooperation with physicists.

Using this tool, “We understood that during mitosis, the motors bind to the microtubules,” Karsenti reports. “By forming bridges between them, and by all moving in the same direction, toward the two poles of the cell, they rearrange the microtubules in a spindle around the chromosomes.”

I also wanted to reach out to others, to share the adventure of advancing the scientific understanding of our universe.”

ITINERARY OF THE TARA OCEANS EXPEDITION

THE TARA OCEANS EXPEDITION (2009-2013)

IMAGES OF PLANKTON

PLANKTON CHRONICLES

THE MOTORS
THAT DRIVE MITOSIS