From EMBL: “Life and death of proteins”

European Molecular Biology Laboratory

15 February 2018
Berta Carreño

EMBL scientists create a turnover catalogue of almost 10.000 proteins from primary cells

Architecture dependent turnover of the nuclear pore subunits. Top row shows the nuclear pore subunits seen from top, bottom row shows subunits of the nuclear pore cut in half. IMAGE: Jan Kosinski/EMBL.

Proteins perform countless functions in the cell, including transporting molecules, speeding up metabolic reactions and forming structural parts of the cell such as the nuclear pore complex. Protein turnover is a measure of the difference between protein synthesis and protein degradation and it is an important indicator of a cell’s activity in health and disease.

EMBL group leaders Mikhail Savitski and Martin Beck, in close collaboration with Cellzome scientists Marcus Bantscheff and Toby Mathieson, have improved the accuracy of the detection of small changes in protein turnover by developing a better algorithmic treatment of raw mass spectrometry data. As a result, the researchers have published a turnover catalogue of 9699 unique proteins in Nature Communications. The paper focuses on protein complexes and demonstrates that subunits of protein complexes have consistent turnover rates.

What did you do?

We wanted to study protein homeostasis, or the balanced process behind protein synthesis and degradation in primary cells extracted from blood or living tissue. Primary cells provide a better understanding of the in vivo situation than cultured cells but, unfortunately, they have a short lifespan when compared to the protein complexes we wanted to study. To overcome this problem, we developed a better algorithmic treatment of raw mass spectrometry data. The improved algorithm accurately determines very small changes in proteins, allowing us to measure the turnover of 9699 unique proteins, including very long-lived proteins, such as the Histone H1.2 protein which has a half-life of 2242 hours. For the first time, we have a view of protein turnover at a cellular scale in several primary cell types, which will be a valuable resource for the scientific community.

We focused our analysis on protein complexes, particularly on the nuclear pore complex, which is very big and is composed of several sub-complexes. We discovered that there are protein turnover levels that are specific to a given sub-complex. Proteins which are peripheral to the complex, that joined later in evolution, turn out to have much faster turnover than the ones that form the core structure and have been there for a longer time. Contrary to previous understanding, our data clearly suggests that there is a turnover mechanism for the nuclear pore in non-dividing cells. This is exciting because it opens new research in this direction.

Why is understanding protein turnover important?

Protein turnover is important for understanding cellular homeostasis. Our work delineates the tools to study the mechanisms controlling it and will help researchers study a wide range of things, such as ageing, brain function, cancer and neurodegeneration.

Science paper:
Systematic analysis of protein turnover in primary cells, Nature Communications.

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “Multipurpose enhancers and promoters in embryonic development”

European Molecular Biology Laboratory

30 January 2018
Iris Kruijen

Enhancer activity (green) and promoter activity (purple) in the same regulatory element. IMAGE: EMBL / Eileen Furlong.

EMBL scientists show that some promoters can act as enhancers and vice versa.

During gene expression, the information stored in our DNA is transcribed: turned into instructions to produce RNA and proteins that perform specific functions within each cell. DNA regions called promoters are located at the beginning of genes, and determine the starting point where transcription is initiated. Other snippets of DNA called enhancers control when and where specific genes are expressed. Enhancers are often located far away from genes and must relay their regulatory information to a gene’s promoter.

Now, Olga Mikhaylichenko and colleagues in Eileen Furlong’s group at EMBL have gained new insights into the role of enhancers and promoters during embryonic development, a life stage where very tight regulation of gene expression is essential. Furlong explains the main findings of the paper, that explores the balance between enhancer and promoter activity within individual regulatory elements in vivo, and that was published in Genes & Development on January 29, 2018.

What is the key finding in this paper?

“It used to be thought that there was a black-and-white distinction between enhancers and promoters: they can only act as one or the other. Our paper shows that there is actually a large grey area in-between, with elements that can perform both functions to varying degrees. The level of enhancer or promoter activity is reflected by both the amount and the direction of transcription from the regulatory element, so whether the element can be read in one or two directions, unidirectional or bidirectional. We also developed a new framework to measure enhancer and promoter activity for the same element, in the same embryo (see figure), which we suggest should become the standard for future studies.”

Why is this important?

“First of all, we were able to show that things are not as black and white as they seemed. Enhancers and promoters are in various states of evolution with some having exclusive promoter function, others having predominantly enhancer function, and yet other elements, distal enhancers, having weak promoter activity.

One of the findings that I am most excited about is when we looked at activity in the other direction, asking if gene promoters can act as developmental enhancers. Here, we found that promoters that are bidirectionally transcribed can function as both strong enhancers and promoters, for the same gene. This suggests that they regulate both the levels (promoters) and spatial expression (enhancer) of the gene. Interestingly, promoters that are unidirectionally transcribed cannot perform this function.

Hints from other studies suggest that these general features are conserved from fruit flies to humans. Our findings uncover a new aspect of promoter and enhancer function during embryogenesis, and provide interesting insights into how these elements might have evolved to regulate robust embryonic development.”

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “Cell division offers hope to fight antibiotic resistance”

European Molecular Biology Laboratory

9 March 2017 [Not good to hide great science from social media.]
Sonia Furtado Neves

Preventing bacteria like Helicobacter pylori from untangling chromosomes could be a new way to treat infections. IMAGE: AJC1 (CC BY 2.0)

Keeping bacterial chromosomes tangled could offer hope in developing novel approaches to treating bacterial infections.

Growing levels of antibiotic resistance pose a serious threat to global public health and scientists are racing to find novel ways to tackle bacterial infections. EMBL’s Orsolya Barabas explains a recent study by her lab on how bacteria untangle their chromosomes during division, and the hope that this opens for developing new antibacterial treatments.

What did you find?

A bacterium’s DNA is in a ring-shaped chromosome. When the bacterium divides to produce two daughter cells, the DNA has to be repackaged into two rings, one for each new cell. The DNA in the two daughter rings often gets tangled. If those chromosome rings cannot be untangled, the two daughter cells cannot separate from each other and the bacteria will ultimately die.

When a bacterium divides to produce two daughter cells, the DNA in the two daughter rings often gets tangled. IMAGE: Orsolya Barabas/EMBL

Most bacteria have a protein that cuts any DNA tangled during cell division and sticks it back together as two distinct daughter chromosomes. We discovered that this protein doesn’t start cutting as soon as it binds to DNA. First, another protein has to activate it by changing its shape. This means one could look at designing drugs to interfere with that activation process. And that’s really good news because the alternative – preventing proteins from binding to DNA – is very difficult.

Why is it important?

At the moment, it seems like for every antibiotic we have, there’s at least one bacterium that’s resistant to it. This means common infectious diseases are getting harder to treat and procedures that we routinely use today such as organ transplants, chemotherapy, surgery and diabetes management could become very risky. If we’re going to overcome that, we need to look at new ways of fighting bacterial infections. Preventing the untangling of chromosomes during bacterial cell division could be one option. The untangling seems to work in a similar way in most types of bacteria, although the proteins are slightly different from one species to another. So with the knowledge that we’ve acquired in this study, we could either look at developing generic drugs that would target all bacteria equally, or we could also envision going for a specific therapy for specific bacteria.

SOURCE ARTICLE
Bebel A et al. eLife, 23 December 2016. DOI: 10.7554/eLife.19706

RELATED LINKS

Barabas lab | EMBL
WHO factsheet: antimicrobial resistance
ARTICLE TAGS
Antibiotic resistance
Health
Heidelberg
Structural Biology

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “Metabolism matters”

European Molecular Biology Laboratory

2 March 2017
Sonia Furtado Neves

Scientists can now study glycolysis in embryos, thanks to the EMBL scientists’ pyruvate sensor. IMAGE: Vinay Bulusu/EMBL

Cells at different stages of differentiation get energy in different ways, a new approach developed at EMBL shows.

Life requires energy. The strategy a cell uses to obtain that energy can influence not only how fast it multiplies but also a variety of other processes, like which of its genes are turned on. This process – called metabolism – is challenging to track in time and space, so it has not been studied much in developing embryos. Alexander Aulehla shares how, in work published this week in Developmental Cell, his lab is starting to fill that gap.

What did you find?

To study energy metabolism during development, we looked at how somites – the parts of the embryo that will eventually give rise to the vertebral column and striated muscles – are formed in the mouse embryo. Somites develop from presomitic mesoderm (PSM). But cells in the PSM don’t all become somite cells at once. The tail bud, at one end of the PSM, contains cells that are still in a stem-cell-like state, while at the other end, cells are differentiating into somites.

We found that cells at different points of the PSM generate energy in different ways. Undifferentiated cells in the tail bud showed a higher rate of glycolysis – they get their energy mainly by breaking down glucose. In contrast, cells that are about to differentiate had a higher rate of respiration. And there’s a gradient between these two extremes, with more glycolysis the closer you get to the tail bud.

The sensor developed by the EMBL scientists changes colour when cells have higher pyruvate levels (right). IMAGE: Vinay Bulusu/EMBL

How did you do it?

This is one of those projects that was really only possible at EMBL. Vinay Bulusu had an EMBL Interdisciplinary Postdoc fellowship (EIPOD) to work in my lab and Carsten Schultz’s lab. That allowed us to say “let’s try something quite daring: can we visualise pyruvate (an important product of glycolysis) in a mouse embryo?” Working with the Schultz lab, Vinay was able to design a sensor (called a FRET sensor) that measures the amount of pyruvate, not just in cells in a dish, but in an embryo. Then we created a transgenic mouse line that expresses that FRET sensor in all cells and used those mice to analyse pyruvate levels during PSM development using real-time imaging. Thanks to a collaboration with Uwe Sauer’s lab at ETH Zurich, we were also able to use mass spectrometry to directly measure other products of glycolysis, to confirm that there’s a higher rate of glycolysis in the tail bud. It will be exciting to see how others can now use the FRET sensor mouse line in different contexts.

What questions does this raise?

The main question this raises is ‘why?’ Why do PSM cells, which all seem to proliferate at a similar rate, get energy in different ways? We have evidence this could be linked to – and even influence – the signaling machineries that control differentiation. A companion study published alongside ours confirms this link and we are now investigating exactly how this happens at the molecular level.

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From ScienceNews via EMBL: Women in STEM -“…These STEM women are all about biology”

European Molecular Biology Laboratory

ScienceNewsforStudents

[These are just a few of the Women in STEM presented in this article.]

Sep 16, 2016
Bethany Brookshire

From birds to salamanders, these STEM women are all about biology

Lisa Chella studies Zulu sheep — a type of sheep that is native to South Africa. Namhla Mdutshane

Lisa Chella

“My research and life’s work so far is conservation of indigenous species,” says Chella. She’s a biologist at the University of Zululand in Mangeze, South Africa. She studies a type of sheep — the Zulu sheep — and has mapped out its genome.

Chella is fearless around some animals. She’s handled crocodiles, pythons and chameleons, for instance. But her weakness? “I am utterly and completely terrified of moths and butterflies,” she says.

When Science News for Students asked for women in science, technology, engineering and math to send us pictures of themselves, we got answers from all aspects of STEM. We received more than 150 responses! They include 18 countries and all seven continents.

We’ve put some photos in our feature story on women in science. But we wanted to make sure they all got their chance to shine. So this week, we are sharing their stories. Here are women who are taking on the study of life. Meet our biologists!

Joanna Bagniewska has lived in seven countries! Now, she helps immigrants in England adjust to their new lives. Andrew Steele

Joanna Bagniewska

“I’m a zoologist, and I study invasive species,” says Bagniewska. “These are species that have been brought by humans, accidentally or intentionally, to new places and have become a threat to native biodiversity there.” She works at the University of Reading in England.

Bagniewska’s life has taken her around the globe. Originally from Poland, she’s lived in Italy, Thailand, China, Germany, the United States and England. “In my spare time,” she says, “I now work with migrant communities in Britain.”

When Lorelei Eschbach isn’t handling giant millipedes, she’s tearing it up on the roller derby rink. Sean Hale

Lorelei Eschbach

Ever wondered where the animals in your classroom come from? Eschbach knows. She works at Ward’s Science, located in Rochester, N.Y. “Basically, whatever science needs your classroom has, we provide it,” she says. “I more specifically work in the algae and protozoa lab. If a class needs amoeba or tardigrades, I culture them and prepare them for shipping.”

As part of her job, Eschbach gets to take her work on the road. She heads to schools to show off what is housed in the entomology lab, she says. “We are the only company that sells giant tropical millipedes in the [United States], so it is an awesome experience for the students to get to hold one.”

Charlotte Milling holds one of her study subjects, a pygmy rabbit. Laura McMahon

Charlotte Milling

Milling is a graduate student in ecology and conservation at the University of Idaho in Moscow. She is studying why animals choose to live where they do and the influence of physiology on their behavior and personality. “Knowing how and why an animal uses the habitat and resources it does allows us to make informed and effective wildlife management decisions,” she notes.

Scientist is definitely Milling’s first choice of career. But if she could be anything other than a scientist, she says, “I would be a baker and a track coach, because I love pastries, running and teaching, and the schedules are compatible.”

K. Nicholson went on a bear hunt for science, and she got a big one. Don Young

Kerry “K” Nicholson

Many people might see a bear and run. Not Nicholson! She’s a carnivore biologist who’s studied everything from bears to lizards in places as varied as Texas, Antarctica, Sweden and South Africa. But in the end, it was Alaska that drew her. “Although Sweden was filled with meatballs and Prinsesstårta, it was just not the same midnight sun,” she says. “The moose were too small, reindeer too nice and too few wolves or bears — this Goldilocks needed to go home.”

Now she’s a biologist with the Alaska Department of Fish and Game in Fairbanks. But that’s not all. Nicholson throws pots, kayaks, does traditional Native Alaskan bead work — and has even learned how to breathe fire.

[There are many more Women in STEM presented in this article.]

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “How new HIV drugs lock virus in immaturity”

European Molecular Biology Laboratory

14 July 2016
Sonia Furtado Neves

For HIV to mature, a crucial cutting point has to be exposed. IMAGE: Florian Schur/EMBL

Study provides insights into workings of new HIV drugs and how virus becomes resistant

A new type of HIV drug currently being tested works in an unusual way, scientists in the Molecular Medicine Partnership Unit, a collaboration between EMBL and Heidelberg University Hospital, have found. They also discovered that when the virus became resistant to early versions of these drugs, it did not do so by blocking or preventing their effects, but rather by circumventing them. The study, published online today in Science, presents the most detailed view yet of part of the immature form of HIV.

HIV, the virus that causes AIDS, comes in two forms: immature and mature. The immature form is assembled inside an infected person’s cells. After an immature virus particle has left the cell, it has to change into the mature form before it can infect other human cells. A new group of drugs that inhibit this maturation is currently undergoing clinical trials, but so far it was unclear how exactly these drugs act.

To go from immature to mature, HIV has to cut the connections between its main building blocks, and rearrange those pieces. John Briggs’ lab at EMBL and Hans-Georg Kräusslich’s lab at Heidelberg University Hospital looked at a particularly important cutting point. It connects building blocks known as the capsid protein and the spacer peptide 1, and if it is not cut, the virus cannot mature. The scientists used a combination of cryo-electron tomography and subtomogram averaging to reveal exactly what this part of the immature form of HIV looks like in 3D. They found that the cutting site is hidden in a position where the virus’ cutting machinery can’t sever it. So for the virus to mature, the structure first has to change, to expose that cutting point.

“When we looked at the virus with one of these inhibitor drugs on it, we found that the inhibitor doesn’t prevent the cutting machinery from getting in, as you might expect,” says Florian Schur, who carried out the work in Briggs’ lab. “Rather, the drug locks the immature virus structure in place, so that it can’t be cut.”

The scientists determined the cutting site’s 3D structure in whole HIV particles. IMAGE: Florian Schur/EMBL

When the new inhibitor drugs were first developed, scientists found that HIV viruses with certain mutations in their genetic sequence were unaffected by the drugs – they were resistant. Having determined what the cutting point looks like and how the drugs act, Briggs and colleagues are now able to understand the effects of those mutations.

“Rather than stopping the drug from binding, the virus becomes resistant through mutations that destabilise the immature structure,” says Kräusslich. “This allows it to rearrange and be cut even when the drug is in place.”

The researchers would now like to probe the virus and the inhibitor drugs in even greater detail, to understand exactly how the drugs attach themselves to the viral proteins, and potentially gather data that could help to search for better drugs – or to design them.

The method used in this study – combined cryo-electron tomography and subtomogram averaging – enables scientists to study structures inside irregular viruses like HIV, or within cells. In essence, the scientists use an electron microscope to obtain a 3D image of the sample – in this case, whole HIV-1 particles. They then identify all the copies of the object they want to study – all the instances of the capsid protein-spacer peptide 1 cutting point – and use software to rotate the 3D image of each copy so that they are all facing the same way. By repeating this procedure with thousands of images, the scientists can obtain an accurate picture. With this approach, researchers can study such samples without having to purify them in a test-tube, which means that they see them in their real state. The EMBL scientists’ work now proves that the method can provide the level of detail that is crucial to understanding how molecular machines work and to informing drug design.

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “Revealing structure of nuclear pore’s inner ring”

European Molecular Biology Laboratory

14 April 2016
Sonia Furtado Neves, Science

The architecture of the nuclear pore. The outer ring is colored in orange and blue, whereas the newly characterized inner ring is seen in green and lemon. Credit: Jan Kosinski/EMBL

Study published today in Science sheds light on structure of nuclear pore complex, which plays a crucial role in controlling molecular traffic to a cell’s nucleus

It was a 3D puzzle with over 1000 pieces, with only a rather fuzzy outline as a guide. But scientists at EMBL have now put enough pieces in place to see the big picture. In a study* published today in Science, they present their latest findings, bringing the nuclear pore complex into focus.

The nuclear pore is a passage into the cell’s nucleus. A typical cell has hundreds of these pores, playing a crucial role in controlling the hundred of thousands of molecules that enter and exit this compartment every minute. Nuclear pores are used by many viruses to inject their genetic material into a host and they are known to change when cells become cancerous, so knowing how they work is important. Scientists understood many of the components of the nuclear pore, but exactly how those building blocks fitted together was unclear.

“The nuclear pore is the biggest, most complicated protein complex in a human cell. We now understand how it is structured,” says Martin Beck, who led the work at EMBL. “This is a very important first step towards understanding what actually happens to nuclear pores in cancer, during ageing, and in other conditions.”

The nuclear pore is composed of three layered rings: a nuclear ring facing the nucleus; a cytoplasmic ring facing the rest of the cell; and an inner ring in between those two. Having already pieced together how the building blocks of the nuclear and cytoplasmic rings are arranged, Martin Beck’s group at EMBL have now worked out the arrangement of the pieces that form the inner ring.

“Surprisingly, we found that although it is made of different building blocks, the inner ring has the same basic architecture as the other two rings,” says Shyamal Mosalaganti from EMBL, who studied the ring using cryo-electron microscopy. “This very complicated structure is built using simple principles . We were able to uncover that because we interweaved a lot of different techniques here.”

Access mp4 video here .

*SOURCE ARTICLE

Molecular architecture of the inner ring scaffold of the human nuclear pore complex

Science team:

Jan Kosinski1,*, Shyamal Mosalaganti1,*, Alexander von Appen1,*, Roman Teimer2, Amanda L. DiGuilio3, William Wan1, Khanh Huy Bui4, Wim J.H. Hagen1, John A. G. Briggs1,5, Joseph S. Glavy3, Ed Hurt2, Martin Beck1,5,†

Affiliations:

1Structural and Computational Biology Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
2Biochemistry Center of Heidelberg University, Im Neuenheimer Feld 328, D-69120 Heidelberg, Germany.
3Department of Chemistry, Chemical Biology and Biomedical Engineering, Stevens Institute of Technology, 507 River Street, Hoboken, NJ 07030, USA.
4Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec, Canada.
5Cell Biology and Biophysics Unit, EMBL, Heidelberg, Germany.

↵†Corresponding author. E-mail: martin.beck@embl.de

↵* These authors contributed equally to this work.

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “Foetus, or placenta?”

European Molecular Biology Laboratory

24 March 2016
Mary Todd Bergman

New study on embryonic development by EMBL-EBI’s Marioni group sheds light on early cell-fate decisions. IMAGE: Zernicka-Goetz lab, University of Cambridge

New research in Cell shows subtle differences between seemingly identical cells at a very early stage of development

When exactly does an embryonic cell decide whether it will become part of the foetus, or part of the placenta? Scientists at the University of Cambridge and EMBL-EBI shed light on this important question by studying the development of mice embryos only four cells in size. The findings, published in Cell, have implications for the understanding of mammalian development.

When an embryo first starts to develop, genetic factors influence whether its cells will become part of the supporting structure around a new organism, or part of the organism itself. Today’s study shows that seemingly identical cells in a two-day-old mouse embryo already begin to display subtle differences.

Once an egg has been fertilised by a sperm, it divides several times to become a free-floating ball of stem cells. At first, these cells are ‘totipotent’, able to divide and give rise into any type of cell, placental or organismal. Some of those cells then switch to a ‘pluripotent’ state, in which their development is restricted to generating the cells of the whole body, rather than the placenta. Understanding that switching mechanism, and when it occurs, is the subject of intense research.

“We know that life starts when a sperm fertilises an egg, but we’re interested in when the important decisions that determine our future development occur,” says Magdalena Zernicka-Goetz from the Department of Physiology, Development and Neuroscience at the University of Cambridge. “We now know that even as early as the four-cell stage – just two days after fertilisation – the mouse embryo is being guided in a particular direction and its cells are no longer identical.”

The team used the latest sequencing technologies to understand embryo development in mice, looking at the activity of individual genes at a single-cell level. They showed that some genes in each of the four cells behaved differently. The activity of several genes in particular differed the most between cells. These genes form part of the ‘pluripotency network’: they are targeted by key pluripotency factors, including Sox2 and Oct4. The team showed that when activity of such genes is reduced, the activity of a master regulator, which directs cells to develop into the placenta, was increased.

John Marioni of EMBL-EBI, the Wellcome Trust Sanger Institute and CRUK-CI, adds: “We can make use of powerful sequencing tools to deepen our understanding of the molecular mechanisms that drive development in individual cells. Because of these high-resolution techniques, we are now able to see the genetic and epigenetic signatures that indicate the direction in which early embryonic cells will tend to travel.”

The research was funded by the Wellcome Trust, the European Molecular Biology Laboratory and Cancer Research UK.

The science team:
Mubeen Goolam
, Antonio Scialdone
, Sarah J.L. Graham
, Iain C. Macaulay
, Agnieszka Jedrusik
, Anna Hupalowska
, Thierry Voet
, John C. Marionicorrespondenceemail
, Magdalena Zernicka-Goetzcorrespondenceemail

See the full article here .

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Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.

From EMBL: “The cellular crystal factory”

European Molecular Biology Laboratory

Free-electron laser beam hits a peroxisome containing protein crystals. IMAGE: EMBL/CFEL, Thomas Seine

Scientists from the Wilmanns group have teamed up with experts across the Deutsches Elektronen-Synchrotron (DESY) research campus in Hamburg and at the SLAC National Accelerator Laboratory in California to show that naturally formed crystals can diffract X-rays. The first crystals successfully analysed with a free-electron laser inside the cells that produced them are unlikely to be the last.

While structural biologists are familiar with the concept of growing protein crystals in the lab for X-ray crystallography experiments, many may not know that some organisms produce crystals naturally within their cells. “When we heard about these naturally forming crystals, we wondered whether we could use them for crystallography experiments,” says Daniel Passon, a postdoc in the Wilmanns group at EMBL Hamburg. “Producing protein crystals in the lab for crystallography experiments is not always easy – imagine if we could get cells to do this for us: a tiny crystal factory in a cell!”

Crystallography uses X-rays to probe the 3D atomic structure of proteins that have been captured in their crystalline form, but the technique has its limitations. In a study published recently in the International Union of Crystallography Journal (IUCrJ), the team of Hamburg scientists instead used crystals grown inside yeast cells for crystallography experiments at the Linac Coherent Light Source [LCLS], an X-ray free-electron laser facility.

Inside the LCLS

“This study would not have been possible without access to one of only two X-ray Free-Electron Lasers currently operational in the world,” says Matthias Wilmanns, Head of EMBL Hamburg, who oversaw the research, “It is a great example of the importance and potential of emerging infrastructures for the field of structural biology.”

Size matters

The group studied crystals that occur naturally in parts of the cell called peroxisomes. These organelles break down large molecules such as fatty acids, keeping toxic processes safely within their bounds and away from the rest of the cell. In Hansenula polymorpha yeast cells, a protein called alcohol oxidase breaks down methanol molecules into useful byproducts.

To make effective use of the restricted space within the peroxisome, alcohol oxidase molecules are packed tightly into crystals; despite being so densely packed, the enzyme molecules inside this crystal are still active. “We think of crystals as rigid entities, but in fact they are not entirely solid,” explains Arjen Jakobi, a postdoc in the Wilmanns group and the Sachse group at EMBL Heidelberg, who carried out the work together with Passon and Wilmanns. “The methanol molecules pass through the crystals, reacting with the oxidase to become detoxified.”

Daniel Passon prepares nozzles for sample delivery via liquid jet. PHOTO: EMBL/Daniel Passon

While all of these yeast cells have peroxisomes, some cells have more peroxisomes than others, and peroxisome size varies from cell to cell, too. This natural variation posed a problem, as the researchers needed the crystals to be as large as possible, and as similar to each other as possible.

The Wilmanns group worked closely with colleagues at the University of Groningen who identified a mutant strain of yeast that only produces one large peroxisome per cell, each containing one large crystal. “Large is of course relative,” says Passon. At 0.001mm, the crystals were still too small for observations at even the most advanced synchrotrons, where they were likely to be combusted before data could be collected. “Free-Electron Lasers produce a large amount of photons in small bursts and have a very small parallel beam,” explains Wilmanns, “This makes them ideal for looking at such small crystals.”

A novel experience

The experiment was novel not only for EMBL’s scientists. “This field is its infancy and there are few leading experts worldwide,” says Wilmanns. “We teamed up with the Coherent Imaging Division at the neighbouring Center for Free Electron Laser Science (CFEL) at DESY and University of Hamburg, and benefited from the considerable experience and expertise of division Director, Henry Chapman and his group” he adds.

Having done some initial validation experiments on the beamlines in Hamburg, Wilmanns, Chapman and their teams set off to the Linac Coherent Light Source at SLAC with their precious peroxisomes. “For such a novel and exciting experiment, I was really keen to be there in person!” says Wilmanns. “It reminded me of being at the synchrotron 20 years ago – it is a very experimental set-up, but the SLAC staff are skilled and efficient.”

Wilmanns checks that nozzles for sample delivery are working during a test run. PHOTO: EMBL/Daniel Passon

A complementary method

The group prepared two types of samples for the experiment: one with the peroxisomes inside their cells and the other with just the peroxisomes, removed from cells. “Surprisingly, we got better data when we measured the peroxisome inside the cell,” says Jakobi. “There was a lot less interference from the surrounding cell material than we expected.”

Having shown that it is possible to get data from crystals within a cell, the group now hopes to harness the natural ability of the peroxisome to produce crystals of other proteins, thereby side-stepping the need for laborious crystallisation experiments. “This could become a complementary method for structural biologists studying challenging proteins,” Wilmanns concludes.

SOURCE ARTICLE

Jakobi A J et al. IUCrJ (published online 12 January 2016). DOI: 10.1107/S2052252515022927

See the full article here .

Please help promote STEM in your local schools.

Stem Education Coalition

EMBL is Europe’s flagship laboratory for the life sciences, with more than 80 independent groups covering the spectrum of molecular biology. EMBL is international, innovative and interdisciplinary – its 1800 employees, from many nations, operate across five sites: the main laboratory in Heidelberg, and outstations in Grenoble; Hamburg; Hinxton, near Cambridge (the European Bioinformatics Institute), and Monterotondo, near Rome. Founded in 1974, EMBL is an inter-governmental organisation funded by public research monies from its member states. The cornerstones of EMBL’s mission are: to perform basic research in molecular biology; to train scientists, students and visitors at all levels; to offer vital services to scientists in the member states; to develop new instruments and methods in the life sciences and actively engage in technology transfer activities, and to integrate European life science research. Around 200 students are enrolled in EMBL’s International PhD programme. Additionally, the Laboratory offers a platform for dialogue with the general public through various science communication activities such as lecture series, visitor programmes and the dissemination of scientific achievements.