Tag Archives: #lipids

Synthesis Of One Of The Most Abundant Organic Lipids Elucidates Its Structure (Chemistry)

Crenarchaeol is a large, closed-loop lipid that is present in the membranes of ammonium-oxidizing archaea, a unicellular life form that exists ubiquitously in the oceans. In comparison to other archaeal membrane lipids, crenarchaeol is very complex and, so far, attempts to confirm its structure by synthesizing the entire molecule have been unsuccessful. Organic chemists from the University of Groningen have taken up this challenge and discovered that the proposed structure for the molecule was largely, but not entirely, correct.

Crenarchaeol contains 86 carbon atoms and is a ‘macrocycle’, a large closed loop. No fewer than 22 positions in the molecule are chiral. The molecule can be present in two forms that are each other’s mirror image, like a left and a right hand. In the crenarchaeol molecule, all 22 chiral centres have their own specific ‘handedness’. Furthermore, crenarchaeol contains a very uncommon cyclohexane group.

Adri Minnaard | Photo University of Groningen
Adri Minnaard | Photo University of Groningen


This complex molecule was first isolated in 2002 by Jaap Sinninghe Damsté and colleagues at the Royal Netherlands Institute for Sea Research, NIOZ. They identified its structure using spectroscopic techniques but their result has never been confirmed. This is surprising because ammonium-oxidizing archaea play a key role in the oceanic nitrogen cycle and fossil crenarchaeol and its companion lipids are widely used by molecular paleontologists to reconstruct past sea temperatures. ‘The structure of crenarchaeol is a formidable challenge for synthetic organic chemistry,’ says Adri Minnaard, Professor of Organic Chemistry at the University of Groningen . ‘And we decided to take it up.’

First, 3 milligrams of the natural compound were isolated and purified at NIOZ, which took about three months. There are only tiny amounts of crenarchaeol in each cell, just 0.000000001 gram, but because there are so many of these cells in the world’s oceans and since the molecule is very stable and has accumulated in the sediment for millions of years, it is believed to be one of the most abundant organic molecules in marine sediments.

Mount Everest

There are several problems with synthesizing crenarchaeol; getting all the chiral centres in the correct orientation is one of them. ‘And the molecule contains a lot of carbon-carbon bonds, which are difficult to construct.’ But it is a challenge that a synthetic organic chemist cannot resist – just like an ambitious mountaineer cannot resist Mount Everest. Minnaard showed the structure to his PhD student Mira Holzheimer, who worked on palladium-catalyzed synthetic reactions. ‘Her literal response was: “I want to climb that mountain”.’ They made a plan of attack on paper, which involved taking the molecule apart into the building blocks that could be synthesized. This produced a tentative route towards the synthesis of complete crenarchaeol molecules, which Holzheimer explored.

Similar to climbing a mountain for the first time, the synthetic route that they designed at the start sometimes led to a dead end. This meant retracing these steps and trying a new approach. ‘You start with multiple grams of the basic compounds. But in each of the more than 65 intermediate steps, you lose material, sometimes up to 50 percent. And if you run out of intermediates, you have to go all the way back again,’ Minnaard explains.

The structure of crenarchaeol | Illustration A. Minnaard / University of Groningen
The structure of crenarchaeol | Illustration A. Minnaard / University of Groningen


After three years of hard work, Holzheimer had produced a large chunk of the molecule, roughly half of the macrocycle. Minnaard: ‘At that stage, we decided to check it against the corresponding part of natural crenarchaeol.’ This was done using coupled gas chromatography-mass spectrometry. The comparison was made at NIOZ and the results came as a shock: ‘We synthesized the correct carbon skeleton, but the chromatographic behaviour was not the same as that for natural crenarchaeol. Something was wrong’, Minnaard recalls.

After two days of checking, Minnaard and Holzheimer concluded that they really had made the proposed structure. And as it did not match completely with natural crenarchaeol, this could only mean one thing: the proposed structure was not entirely correct. The results pointed towards one of the chiral centres of the unusual cyclohexane group. ‘Our collaborators at NIOZ mis-assigned just one chiral centre out of 22.’ The structure correction was further supported by calculations on the spectra of both natural and prepared crenarchaeol, which were performed by Prof. Remco Havenith and Dr Ana Da Cunha. Minnaard: ‘This shows the value of synthetic chemistry: building a proposed structure is the gold standard for validation.’


Such a complex build as with crenarchaeol does bring some additional pay-offs: ‘We had to develop new synthetic tools, that are now added to the organic synthesis toolbox.’ Furthermore, having the correct structure is relevant for scientists who are studying archaeal membranes. This is usually done through molecular dynamics simulation, explains Minnaard, and the entirely correct structure is now available. Yet this pay-off is not the biggest motivation for Minnaard to take on these projects: ‘It doesn’t always have to have purpose. To me, building molecules can be art.’

Reference: Mira Holzheimer, Jaap S. Sinninghe Damsté, Stefan Schouten, Remco W. A. Havenith, Ana V. Cunha, Adriaan J. Minnaard: Total Synthesis of the Alleged Structure of Crenarchaeol Enables Structure Revision, Angewandte Chemie, online 11 juni 2021

Provided by University of Groningen

Researchers Discover New Mechanism for Lipid Storage Inside Cell (Biology)

In a study published in Journal of Cell Biology, a research group led by Prof. HU Junjie from the Institute of Biophysics of the Chinese Academy of Sciences reported that endoplasmic reticulum (ER) tubule-forming proteins and septin cytoskeletons are involved in the regulation of lipid droplet (LD) biogenesis via their interactions with ER membrane protein FIT2.

LD is the major place for lipid storage, and it plays crucial roles in lipid metabolism and cellular stress responses. In eukaryocytes, ER accomadates neutral lipid synthesis and nascent LD biogenesis. The accumulation of neutral lipids between the leaflets of ER membranes leads to a growing lens-like structure, which eventually buds into the cytosol as nascent LD. However, the details mechanism and regulation of LD biogenesis is largely unclear.

The ER consists of two distinct morphological domains: tubules and sheets. The tubular ER network is generated by reticulons(Rtns)/REEPs, which stabilize membrane curvature, and subsequently connected by dynamin-like GTPase atlastin (ATL). Converging evidence suggests that ER tubules are mechanistically linked to LD formation.

Prof. HU’s team used C. elegans to screen new regulators of ER morphology, and found the deletion of FIT2 caused ER sheets expansion.

Biochemical analysis revealed that FIT2 physically interacted with ER tubule-forming proteins, including Rtn4 and REEP5, and septin cytoskeletons. Deletion or depletion of these proteins caused defective LD formation in mammalian cells and in worms. FIT2-interacting proteins were up regulated during adipocyte differentiation. Importantly, FIT2, ER tubule-forming proteins and septins were transiently enriched at nascent LD formation sites, as shown by super-resolution live cell imaging.

In summary, neutral lipids in the ER accumulate with FIT2, which in turn recruits ER tubule-forming proteins to stabilize the curvature at the growing oil lens and promote lipid preparation. The outward bulging of the oil lens is reinforced by FIT2-interacting septins, serving as a handrail.

This study provides important insight into the early steps of LD formation.

Featured image: Fang Chen, Bing Yan, Jie Ren, Rui Lyu, Yanfang Wu, Yuting Guo, Dong Li, Hong Zhang, Junjie Hu; FIT2 organizes lipid droplet biogenesis with ER tubule-forming proteins and septins. J Cell Biol 3 May 2021; 220 (5): e201907183. doi: https://doi.org/10.1083/jcb.201907183

Provided by Chinese Academy of Sciences

Atomic-level Insights Gained For A Key Lipid-binding Protein Implicated in Cancer (Biology)

Scientists at Sanford Burnham Prebys have identified, at an atomic level, how a part of a protein called PLEKHA7 interacts with a cell’s membrane to regulate important intercellular communications.

The research, published in the journal Structure, points to hotspots within PLEKHA7 as targets for drugs. These targets could be key in designing treatments for advanced colon, breast and ovarian cancers.  

The region, or domain, in PLEKHA7 that the researchers examined, pleckstrin homology (PH), is commonly found in proteins that regulate the movement of cells as well as other important cellular activities. If the interaction between the PH domain and the lipids that comprise cell membranes is disrupted, diseases such as cancer can occur. 

“PH domains have been studied for some time but examining their interactions with membrane-associated lipids has been challenging,” says Francesca M. Marassi, Ph.D., professor and director of the Cell and Molecular Biology of Cancer Program at Sanford Burnham Prebys and corresponding author of the study. “We optimized our structural investigations by using an artificial membrane disc several nanometers in size. This nanodisc is a ‘substitute’ cell membrane and has been an important tool in overcoming obstacles in studying structural aspects of fatty interfaces, such as those involving lipids.” 

Nanodisc ‘substitute’ cell membrane: Protein is blue, lipids are wheat and the two helical lipoproteins that corral the lipid nanodisc and hold it together are pink. © SBP Discovery

The investigators used several techniques to detail the precise areas where the PH domain interacted with lipids in a cell’s membrane. Each of the three techniques was equally important: X-ray crystallography provided a snapshot of the structure; magnetic resonance showed the precise association of the PH domain with the lipid membrane surface; and computer simulations put all of this information together to generate a dynamic movie of the interaction. 

“We were able to determine that the PH domain interacts with the cellular membrane at several locations simultaneously, thereby demonstrating the key role of the membrane as a platform for directing cell signaling and adhesion,” says Marassi. “We found at least three sites along the PH domain that are engaged in binding lipids. Having multiple sites is crucial, because if you think of the PH binding sites as a zipper between proteins and lipids, each additional notch engaged by a zipper makes the binding stronger and more resistant to being pulled apart.” 

One of the reasons the scientists were interested in exploring PLEKHA7 was that the protein has recently been identified as a potential anti-cancer target. However, its molecular mechanism of action has been uncertain. Additionally, the researchers were interested in the PH domain because it has been implicated in advanced breast, renal and ovarian cancers and, most prominently, plays an important role in colorectal cancer.  

“Clinicians have shown that PLEKHA7 is elevated in patients with colorectal cancer, and its levels increase as the disease worsens. It has also been shown that inhibiting PLEKHA7 decreases cellular proliferation and migration,” says Marassi. “Therefore, the PLEKHA7 PH domain might be a useful drug target, but more work is needed to develop agents with greater potency and optimized pharmacological properties.” 

As a next step, the scientists hope to share their structural images with colleagues in other labs at Sanford Burnham Prebys, and elsewhere, to screen for drugs that target the PH domain and lipid membrane binding sites. 

Additional study authors include Alexander E. Aleshin, Yong Yao, Andrey A. Bobkov, Jinghua Yu, Gregory Cadwell, Michael G. Klein, Laurie A. Bankston, Robert C. Liddington and Garth Powis of Sanford Burnham Prebys as well as Amer Iftikhar, Chuqiao Dong and Wonpil Im from the Departments of Biological Sciences, Chemistry and Bioengineering at Lehigh University, PA.  

The study’s DOI is: 10.1016/j.str.2021.03.018. 

Research reported in this press release was supported by grants from the National Institutes of Health (GM118186, CA179087, CA160398, CA030199) and the National Science Foundation (MCB1810695). 

Featured image : Francesca M. Marassi, Ph.D. © SBP

Reference: Alexander E. Aleshin et al., “Structural basis for the association of PLEKHA7 with membrane-embedded phosphatidylinositol lipids”, Cell, 2021. DOI: https://doi.org/10.1016/j.str.2021.03.018

Provided by SBP Discovery

Researchers Reveal How Lipids and Water Molecules Regulate 5-HT Receptors (Biology)

Serotonin, or 5-hydroxytryptamine (5-HT), is a kind of neurotransmitter. 5-HT can regulate multifaceted physiological functions such as mood, cognition, learning, memory, and emotions through 5-HT receptors. 5-HT receptors are a type of G protein-coupled receptor and can be divided into 12 subtypes in humans. As drug targets, they play a vital role in the treatment of schizophrenia, depression, and migraine. 

However, the structural and functional mechanisms of 5-HT receptors have been largely unknown. 

In a study published in Nature on March 24, Prof. H. Eric XU and Prof. JIANG Yi from the Shanghai Institute of Materia Medica (SIMM) of the Chinese Academy of Sciences, together with Prof. ZHANG Yan from Zhejiang University, and their collaborators, have clarified the critical role of PtdIns4P and cholesterol in G-protein coupling and ligand recognition as well as the molecular basis of basal activity and the drug recognition mode of 5-HT receptors, by resolving the cryo-electron microscopy (cryo-EM) structures of five 5-HT receptor–Gi complexes. 

These five 5-HT receptor–Gi complexes include three with 5-HT1A structures (one in the apo state, one bound to 5-HT, and one bound to aripiprazole, an antipsychotic drug), one with 5-HT1D bound to 5-HT, and one with 5-HT1E bound to the 5-HT1E- and 5-HT1F-selective agonist BRL-54443. 

PtdIns4P is one of the major classes of phosphoinositides. In this study, the researchers first identified PtdIns4P as a major phospholipid at the 5-HT1A–G protein interface, which stabilizes the 5-HT1A-G protein complex. 

They found that PtdIns4P is sandwiched between two cholesterol molecules surrounding the 5-HT1A receptor, therefore providing a structural basis for the modulation of 5-HT1A signaling by cholesterol and phospholipids. 

Researchers also found several structured water molecules that form hydrogen bonds with the apo receptor within the orthosteric binding pocket. Water molecules mimic the polar functionalities of 5-HT in the active apo-5-HT1A–Gi complex, thus revealing the key role of water molecules in sustaining the basal activity of 5-HT receptors. 

In addition, the researchers revealed the basis of ligand selectivity and drug recognition in 5-HT receptors. They identified residue at position 6×55 as a key determinant for the BRL-54443 and 5-CT selectivity of 5-HT receptors. 

An outward shift of the extracellular end of TM7 in 5-HT1A stabilizes the quinolinone group of aripiprazole, resulting in 5-HT1A’s high selectivity for aripiprazole.  

A cholesterol molecule was further found to be involved in the stabilization of the aripiprazole pocket and causes aripiprazole to have a higher binding affinity for 5-HT1A. 

The observations in this study have wide implications for a mechanistic understanding of 5-HT signaling and for drug discovery targeting the 5-HT receptor family. 

Featured image: Cryo-EM structures of the 5-HT1A-Gi, 5-HT1D-Gi, and 5-HT1E-Gi complexes (Image by H. Eric Xu’s group) 

Reference: Xu, P., Huang, S., Zhang, H. et al. Structural insights into the lipid and ligand regulation of serotonin receptors. Nature (2021). https://doi.org/10.1038/s41586-021-03376-8

Provided by Chinese Academy of Sciences

Ultrasound Has Potential to Damage Coronaviruses (Medicine)

Simulations show ultrasound waves at medical imaging frequencies can cause the virus’ shell and spikes to collapse and rupture.

The coronavirus’ structure is an all-too-familiar image, with its densely packed surface receptors resembling a thorny crown. These spike-like proteins latch onto healthy cells and trigger the invasion of viral RNA. While the virus’ geometry and infection strategy is generally understood, little is known about its physical integrity.

A new study by researchers in MIT’s Department of Mechanical Engineering suggests that coronaviruses may be vulnerable to ultrasound vibrations, within the frequencies used in medical diagnostic imaging.

Through computer simulations, the team has modeled the virus’ mechanical response to vibrations across a range of ultrasound frequencies. They found that vibrations between 25 and 100 megahertz triggered the virus’ shell and spikes to collapse and start to rupture within a fraction of a millisecond. This effect was seen in simulations of the virus in air and in water.

The results are preliminary, and based on limited data regarding the virus’ physical properties. Nevertheless, the researchers say their findings are a first hint at a possible ultrasound-based treatment for coronaviruses, including the novel SARS-CoV-2 virus. How exactly ultrasound could be administered, and how effective it would be in damaging the virus within the complexity of the human body, are among the major questions scientists will have to tackle going forward.

“We’ve proven that under ultrasound excitation the coronavirus shell and spikes will vibrate, and the amplitude of that vibration will be very large, producing strains that could break certain parts of the virus, doing visible damage to the outer shell and possibly invisible damage to the RNA inside,” says Tomasz Wierzbicki, professor of applied mechanics at MIT. “The hope is that our paper will initiate a discussion across various disciplines.”

The team’s results appear online in the Journal of the Mechanics and Physics of Solids. Wierzbicki’s co-authors are Wei Li, Yuming Liu, and Juner Zhu at MIT.

A spiky shell

As the Covid-19 pandemic took hold around the world, Wierzbicki looked to contribute to the scientific understanding of the virus. His group’s focus is in solid and structural mechanics, and the study of how materials fracture under various stresses and strains. With this perspective, he wondered what could be learned about the virus’ fracture potential.

Wierzbicki’s team set out to simulate the novel coronavirus and its mechanical response to vibrations. They used simple concepts of the mechanics and physics of solids to construct a geometrical and computational model of the virus’ structure, which they based on limited information in the scientific literature, such as microscopic images of the virus’ shell and spikes.

The 3D image of the collapsing virus, right, captured at the instant of the maximum vibration amplitude. Spikes were removed from the color-coded plot, on left, for clarity. Credits: Courtesy of the researchers

From previous studies, scientists have mapped out the general structure of the coronavirus — a family of viruses that s HIV, influenza, and the novel SARS-CoV-2 strain. This structure consists of a smooth shell of lipid proteins, and densely packed, spike-like receptors protruding from the shell.

With this geometry in mind, the  team modeled the virus as a thin elastic shell covered in about 100 elastic spikes. As the virus’ exact physical properties are uncertain, the researchers simulated the behavior of this simple structure across a range of  elasticities for both the shell and the spikes.

“We don’t know the material properties of the spikes because they are so tiny — about 10 nanometers high,” Wierzbicki says. “Even more unknown is what’s inside the virus, which is not empty but filled with RNA, which itself is surrounded by a protein capsid shell. So this modeling requires a lot of assumptions.”

“We feel confident that this elastic model is a good starting point,” Wierzbicki says. “The question is, what are the stresses and strains that will cause the virus to rupture?”

A corona’s collapse

To answer that question, the researchers introduced acoustic vibrations into the simulations and observed how the vibrations rippled through the virus’ structure across a range of ultrasound frequencies.

The team started with vibrations of 100 megahertz, or 100 million cycles per second, which they estimated would be the shell’s natural vibrating frequency, based on what’s known of the virus’ physical properties.

When they exposed the virus to 100 MHz ultrasound excitations, the virus’ natural vibrations were initially undetectable. But within a fraction of a millisecond the external vibrations, resonating with the frequency of the virus’ natural oscillations, caused the shell and spikes to buckle inward, similar to a ball that dimples as it bounces off the ground.

As the researchers increased the amplitude, or intensity, of the vibrations, the shell could fracture — an acoustic phenomenon known as resonance that also explains how opera singers can crack a wineglass if they sing at just the right pitch and volume. At lower frequencies of 25 MHz and 50 MHz, the virus buckled and fractured even faster, both in simulated environments of air, and of water that is similar in density to fluids in the body.

“These frequencies and intensities are within the range that is safely used for medical imaging,” says Wierzbicki.

To refine and validate their simulations, the team is working with microbiologists in Spain, who are using atomic force microscopy to observe the effects of ultrasound vibrations on a type of coronavirus found exclusively in pigs. If ultrasound can be experimentally proven to damage coronaviruses, including SARS-CoV-2, and if this damage can be shown to have a therapeutic effect, the team envisions that ultrasound, which is already used to break up kidney stones and to release drugs via liposomes, might be harnessed to treat and possibly prevent coronavirus infection. The researchers also envision that miniature ultrasound transducers, fitted into phones and other portable devices, might be capable of shielding people from the virus.

Wierzbicki stresses that there is much more research to be done to confirm whether ultrasound can be an effective treatment and prevention strategy against coronaviruses. As his team works to improve the existing simulations with new experimental data, he plans to zero in on the specific mechanics of the novel, rapidly mutating SARS-CoV-2 virus.

“We looked at the general coronavirus family, and now are looking specifically at the morphology and geometry of Covid-19,” Wierzbicki says. “The potential is something that could be great in the current critical situation.”

Featured image: Ultrasound has potential to damage coronaviruses, a new MIT study finds. Credits: Image: MIT News, with images from iStockphoto

Reference: Tomasz Wierzbicki, Wei Li, Yuming Liu, Juner Zhu, Effect of receptors on the resonant and transient harmonic vibrations of Coronavirus, Journal of the Mechanics and Physics of Solids, Volume 150, 2021, 104369, ISSN 0022-5096, https://doi.org/10.1016/j.jmps.2021.104369. (https://www.sciencedirect.com/science/article/pii/S0022509621000600)

Provided by MIT

Dietary Fats Interact With Grape Tannins to Influence Wine Taste (Food)

Wine lovers recognize that a perfectly paired wine can make a delicious meal taste even better, but the reverse is also true: Certain foods can influence the flavors of wines. Now, researchers reporting in ACS’ Journal of Agricultural and Food Chemistry have explored how lipids –– fatty molecules abundant in cheese, meats, vegetable oils and other foods –– interact with grape tannins, masking the undesirable flavors of the wine compounds.

Tannins are polyphenolic compounds responsible for the bitterness and astringency of red wines. Wine testers have noticed that certain foods reduce these sensations, improving the flavor of a wine, but scientists aren’t sure why. Some studies have indicated that tannins interact with lipids at the molecular level. In foods, lipids are found as fat globules dispersed in liquids or solids. Julie Géan and colleagues wanted to investigate how tannins influence the size and stability of lipid droplets in an emulsion. They also wondered how the prior consumption of vegetable oils would impact the taste of tannins for human volunteers.

The researchers made an oil-in-water emulsion using olive oil, water and a phospholipid emulsifier. Then, they added a grape tannin, called catechin, and studied the lipids in the emulsion with various biophysical techniques. The team found that the tannin inserted into the layer of emulsifier that surrounded the oil droplets, causing larger droplets to form. In taste tests, volunteers indicated that consuming a spoonful of rapeseed, grapeseed or olive oil before tasting a tannin solution reduced the astringency of the compounds. Olive oil had the greatest effect, causing the tannins to be perceived as fruity instead of astringent. Combining the biophysical and sensory results, the researchers concluded that tannins can interact with oil droplets in the mouth, making them less available to bind to saliva proteins and cause astringency.

The authors acknowledge funding from the Conseil Interprofessionnel du Vin de Bordeaux.

“New Insights into Wine Taste: Impact of Dietary Lipids on Sensory Perceptions of Grape Tannins”
Journal of Agricultural and Food Chemistry

Provided by American Chemical Society

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Overlooked Helpers: Membrane Building Blocks Play a Decisive Role in Controlling Cell Growth (Botany)

Lipids are the building blocks of a cell’s envelope – the cell membrane. In addition to their structural function, some lipids also play a regulatory role and decisively influence cell growth. This has been investigated in a new study by scientists at Martin Luther University Halle-Wittenberg (MLU). The impact of the lipids depends on how they are distributed over the plasma membrane. The study was published in “The Plant Cell”.

If plant cells want to move, they need to grow. One notable example of this is the pollen tube. When pollen lands on a flower, the pollen tube grows directionally into the female reproductive organs. This allows the male gametes to be delivered, so fertilisation can occur. The pollen tube is special in that it is made up of a single cell that continues to extend and, in extreme cases, can become several centimetres long. “This makes pollen tubes an exciting object for research on directional growth processes,” says Professor Ingo Heilmann, head of the Department of Plant Biochemistry at MLU.

For the current study, Heilmann’s team focused on the phospholipids of pollen tubes, which, as the main component of the plasma membrane, are responsible for separating the cell’s interior from its surroundings. “Lipids are generally known to have this structuring function,” says Dr Marta Fratini, first author of the study. It has only recently come to light that some phospholipids can also regulate cellular processes. The scientists from Halle have now been able to show that a specific phospholipid called phosphatidylinositol 4,5-bisphosphate (“PIP2”) can control various aspects of cell growth in pollen tubes – depending on its position at the plasma membrane. They did this by labelling the lipid with a fluorescent marker. “We found it is either distributed diffusely over the entire tip of the pollen tube without a recognisable pattern, or is concentrated in small dynamic nanodomains,” Fratini explains. One can imagine a group of people on a square: either individuals remain 1.5 metres apart as currently prescribed, or they form small groups.

Pollen tube with swollen tip (green: one of the enzymes responsible for lipid production, magenta: the lipid nano domains discovered and described in the study) / Foto: Marta Fratini

It appears that different enzymes are responsible for the varying distribution of PIP2. “Plant cells have several enzymes that can produce this one phospholipid,” explains Heilmann. Like the lipids, some of these enzymes are widely distributed over the membrane and others are concentrated in nanodomains, as shown by the current study. Depending on which of the enzymes the researchers artificially increased, either the cytoskeleton – a structure important for directed growth – stabilised and the pollen tube swelled at the tip, or more pectin – an important building material for plant cell walls – was secreted. This made the cell branch out at the tip. To make sure that the distribution of the lipids was indeed responsible for these growth effects, the biochemists artificially changed the arrangement of the enzymes at the plasma membrane – from clusters to a wide scattering or vice versa. It turns out they were able to control the respective effects on cell growth. 

“As far as I know, our study is the first to trace the regulatory function of a lipid back to its spatial distribution in the membrane,” says Heilmann. Further research is now needed to clarify exactly how the membrane nanodomains assemble and how the distribution of PIP2 at the membrane can have such varying effects. 

The research was funded by the Deutsche Forschungsgemeinschaft (German Research Association, DFG) through various programmes, including the Research Training Group 2498 “Communication and Dynamics of Plant Cell Compartments”, and supported by the Centre for Innovation Competence HALOmem at MLU. 

Featured image: A pollen tube that grows out of a pollen grain (green: one of the enzymes responsible for the production of lipids that control cell growth, magenta: actin cytoskeleton). / Foto: Marta Fratini

Study: Fratini et al. Plasma membrane nano-organization specifies phosphoinositide effects on Rho-GTPases and actin dynamics in tobacco pollen tubes. The Plant Cell (2020). doi: 10.1093/plcell/koaa035

Provided by Martin Luther University Halle-Wittenberg

How Lipids Distribute Proteins Within Cells? (Biology)

An international team of scientists, coordinated by the Seville Institute of Biomedicine (IBiS) and the University of Seville has solved one of the hitherto unresolved enigmas of basic biology: how exactly do lipids distribute proteins within a cell? To do this, they used a new, completely innovative microscopy technology, which they applied to “mutant” cells they designed in their laboratory.

This discovery represents a major advance in understanding how proteins are distributed in cells to perform their vital functions, and could open the door to understanding the causes of diseases associated with failures in protein distribution at the cellular level: from cancer to neurodegenerative diseases, such as Alzheimer’s.

The study was carried out by the Department of Cell Biology’s “Membrane Trafficking” research group, part of the Faculty of Biology of the University of Seville and the IBiS, led by Professor Manuel Muñiz Guinea, in collaboration with the universities of Hiroshima (Japan), Geneva and Fribourg (Switzerland). The RIKEN Institute in Japan, where the “Super-resolution Living Cell Microscopy” Laboratory is located, also participated in this project. This is a unique facility in the world which conducted analyses using a high resolution fluorescence microscope that allows the study of very fast and dynamic processes in living cells on an incredibly small scale.

As Manuel Muñiz explains, “the cell is the basic unit of life and, at the same time, an extremely complex and sophisticated machine in which thousands of proteins, among other components, are strategically located in different compartments where they carry out cellular functions”. The cell must ensure that its proteins are properly distributed to their place of function, because if this fails and they do not reach their destination, the proteins either stop working or get out of control, causing diseases ranging from genetic syndromes to cancer or neurological diseases. Therefore, it is important to investigate how proteins are distributed towards their functional destination.

Many years ago it was suggested that, in addition to cells’ conventional protein transportation machinery (whose discovery received the Nobel Prize for Medicine in 2013), the lipids that make up the cell membranes may also play an additional role in the distribution of proteins within cells. This work by the Seville researchers solves this enigma of basic biology, demonstrating for the first time how lipids can distribute proteins at the cellular level.

Molecular exit doors

The proteins are manufactured in a compartment of the cell and then have to be distributed correctly by exiting through specific “doors”. In this study, scientists from Seville discovered that membrane lipids are responsible for selecting and directing certain proteins to the correct exit doors.

To make this discovery, they designed a “mutant cell” that was programmed to manufacture a shortened version of cellular lipids called ceramides. The researchers suspected that the length of these lipids could be a determining factor in choosing the appropriate exit door.

“And that’s exactly what we found,” explains the IBiS researcher. “Thanks to the short ceramides we generated, we were able to demonstrate for the first time that lipids are only able to guide proteins during transport if they are the right length. Moreover, by using such a powerful ‘super microscope’ we were able to capture for the first time on an ultra small scale and in vivo how proteins exit through these molecular doors”.

Yeast model

As a curiosity, this study was carried out using yeast cells (the same unicellular fungus used to make bread, beer and wine) as a model organism, “because, being eukaryotic cells just like ours, they perform the same basic cellular processes in a very similar way, so the observations can be extrapolated to human cells”, explains the University of Seville professor.

However, because they are also simpler and can be genetically manipulated very effectively, “yeast cells are an excellent model to understand the fundamental workings of the human cell and what causes disease, as demonstrated by the fact that several Nobel Prize for Medicine have been awarded to researchers who used this microorganism in their studies, including Paul Nurse or Randy Schekman”.

In conclusion, Manuel Muñiz explains that the article published in Science Advances “has also served to demonstrate that lipids and proteins influence each other to self-organise together within the cell”, and points out that the mechanism they have discovered and used for this “could be used in other processes, such as the entry and exit of certain viruses from the cell, as well as in the formation of exosomes (extra-cellular lipid vesicles involved in communication between cells, particularly in cancer)”.

Featured image: Newly synthesized C26 ceramide–based GPI-AP cargos form clusters in the ER membrane adjacent to specific ERES. © Sofia et al.

Reference: Sofia Rodriguez-Gallardo, Kazuo Kurokawa et al., “Ceramide chain length–dependent protein sorting into selective endoplasmic reticulum exit sites”, Science Advances 11 Dec 2020: Vol. 6, no. 50, eaba8237 DOI: 10.1126/sciadv.aba8237 https://advances.sciencemag.org/content/6/50/eaba8237

Provided by University of Seville

How a Large Protein Complex Assembles in a Cell (Biology)

A team of ETH re­search­ers led by Karsten Weis has de­veloped a method that al­lows them to study the as­sembly pro­cess for large pro­tein com­plexes in de­tail for the first time. As their case study, the bio­lo­gists chose one of the largest cel­lu­lar com­plexes: the nuc­lear pore com­plex in yeast cells.

The nuc­lear pore com­plexes (or­ange struc­tures), some of which are in the pro­cess of as­sembly, are among the largest pro­tein com­plexes in a cell. (Visu­al­isa­tion: Olga V Posukh, In­sti­tute of Mo­lecu­lar and Cel­lu­lar Bio­logy, Nov­os­ibirsk)

Cells produce a great number of different protein complexes, each of which is made up of many individual proteins. These protein complexes, like ribosomes for example, are what regulate almost all of a cell’s life-sustaining biological functions.

Biologists have succeeded in determining the structure of many of these complexes, but there is less research so far on how the individual proteins assemble and then change over time. Conventional approaches have thus far proved insufficient for studying the exact course that these reactions in cells take, especially where large complexes are concerned.

A group of ETH researchers led by Karsten Weis and research associate Evgeny Onischenko at ETH Zurich’s Institute of Biochemistry are now presenting a new approach. Their method makes it possible to track the dynamics of protein complex assemblies, even for very large ones, with high temporal resolution. The study has just been published in the journal Cell.

Inspired by metabolic analysis

The ETH researchers call their new approach KARMA, which stands for kinetic analysis of incorporation rates in macromolecular assemblies and is based on methods for investigating metabolic processes. Scientists researching metabolism have long used radioactive carbon in their work, e.g., to label glucose molecules, which cells then take up and metabolise. The radioactive labelling enables researchers to track where and at what point in time the glucose molecules or their metabolites appear.

“This type of research inspired us to apply a similar principle in exploring the reactions that take place in the assembly of protein complexes,” Weis explains. In their approach, the ETH researchers work with labelled amino acids, the fundamental building blocks of proteins, which contain heavier carbon and nitrogen isotopes. In a culture of yeast cells, the team replaces the lightweight amino acids with their heavier counterparts. The yeast uses these heavy amino acids in protein synthesis, which shifts the molecular weight of all newly produced proteins.

A time scale for the assembly of a complex

To isolate protein complexes, the researchers remove yeast cells from the cultures at regular intervals and employ mass spectrometry to measure the tiny weight difference between molecules with heavier amino acids and those without. This indicates the age of a protein in a complex. Basically, the older the protein, the earlier it was incorporated into the complex. Based on these age differences, the researchers apply kinetic state models to ultimately reconstruct the precise assembly sequence of a given protein complex.

As a case study to validate their method, Weis and his team chose the nuclear pore complex in yeast cells. This structure has some 500 to 1,000 elements composed of about 30 different proteins each in multiple copies, thus making it one of the largest known protein complexes.

Using KARMA, the ETH biochemists were able to obtain a detailed map of which modules are integrated into the structure and when. One of their findings was a hierarchical principle: individual proteins form subunits within a very short time, which then assemble from the centre out to the periphery in a specific sequence.

Durable scaffold

“We’ve demonstrated for the first time that some proteins are used very quickly in the assembly of the pore complex, while others are incorporated only after about an hour. That’s an incredibly long time,” Weis says. A yeast cell divides every 90 minutes, which means it would take almost a whole generation to complete assembly of this vital pore complex. Precisely why the assembly of new pores takes so long in relation to the yeast reproduction cycle is not known.

The ETH researchers also show that once assembly of the pore is complete, parts of the complex are highly stable and durable – in the inner scaffold, for example, hardly any components are replaced during its lifetime. By contrast, proteins at the periphery of the nuclear pore complex are frequently replaced.

Defective nuclear pores facilitate disease

Nuclear pores are some of the most important protein complexes in cells, as they are responsible for the exchange of substances and molecules between the cell nucleus and cytoplasm. For example, they transport messenger RNA from the nucleus to the cellular machinery outside the nucleus, which needs these molecules as blueprints for new proteins.

Moreover, nuclear pores play direct and indirect roles in human disease. Accordingly, changes in the nuclear pore and its proteins can impact the development of conditions like leukaemia, diabetes or neurodegenerative diseases such as Alzheimer’s. “Generally speaking, though, the reasons why pore defects cause these disease patterns are not well understood,” Weis says, explaining that KARMA might help to gain deeper insight into such issues in the future.

Versatile platform

“Although we applied KARMA to only one protein complex in this study, we’re excited about its future applications. Our method will now enable us to decipher the sequence of a whole host of biological processes,” Weis says. Their technique can be used, for example, to study molecular events that occur during the infection cycle of viruses such as COVID-19 and potentially help to find new drug candidates that break that cycle.

The new method can also be applied to other biological molecules besides proteins, such as RNA or lipids.

Reference: Onischenko E, Noor E, Fischer JS, Gillet L, Wojtynek M, Vallotton P, Weis K: Maturation Kinetics of a Multiprotein Complex Revealed by Metabolic Labeling, Cell, Available online 16 December 2020. DOI: 10.1016/j.cell.2020.11.001

Provided by ETH Zurich