TS Digest Issue | September 2023 | Fishing for Viruses

An illustration of a person on a unicycle juggling clocks to indicate a busy schedule.

From the Editor| Article

Handling the Extras in Academia

Researchers are expected to juggle several job duties with little to no training for those tasks. Is peer coaching an option?

Meenakshi Prabhune, PhD

Sep 1, 2023 | 1 min read

Above Image Credit:

To say that academics wear multiple hats is an understatement. A researcher must ideate, experiment, teach, analyze, collaborate, present, write, and more. As academics move up the ladder, their hat closets expand further. A new list of responsibilities adds to their already lengthy to-do lists: oversee budgets, mentor students, plan conferences, apply for funding, review papers, and serve on committees, to name a few.

Freshly minted faculty members have spent the couple of years leading up to their coveted positions just on the grueling application process. When academics focus all of their efforts on getting the desired job, they have little time to prepare for what it entails.

Given the minimal formal soft skills training, I am in awe of academics who succeed and empathetic towards those who struggle with these extra responsibilities. I wish that graduate students and postdoctoral scholars could undergo training for all of the functions they will handle as group leaders.

Researchers collaborate often to share their scientific expertise; wouldn’t it be great if they also set up soft skills collaborations? They could learn about mentorship from someone who wins over students; find out how a neighboring lab thrives on social media; or take notes from a senior researcher on handling a growing lab.

It would be valuable to rely on the strengths of peers in person and online. This month, we are proud to add bite-sized advice to our TS Digest. In this issue, Mark Emerson, a biologist at The City College of New York and an Undergraduate Research Mentor Award winner, offers tips for mentoring undergraduate students to ensure that everyone involved gets the most out of the experience.

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A depiction of a chain of amino acids arranged in a quaternary protein structure.

Technology Highlight| Article

Breaking Down Barriers to Protein Sequencing

Next-generation protein sequencing is becoming more powerful, streamlined, and accessible.

Sep 1, 2023 | 2 min read

Above Image Credit:

Protein and proteomic sequencing is an important part of understanding how organisms operate in health and disease. However, protein sequencing technology has lagged behind next-generation nucleic acid sequencing, largely due to the fact that proteins, unlike nucleic acids, cannot be easily amplified.¹ Novel strategies and technologies are aimed at making higher-resolution protein sequencing more streamlined and accessible.

A photo of the PlatinumTM Next-Generation Protein Sequencer, a small roughly cube-shaped blue-black-silver instrument.

Next-generation instruments such as the Platinum^™ Next-generation Protein sequencer enable single-molecule protein sequencing.

Quantum-Si

To study proteins, researchers commonly turn to mass spectrometry (MS) and antibody-based strategies such as western blotting and enzyme-linked immunosorbent assays (ELISA).¹ However, these techniques have their drawbacks. MS offers high-resolution protein characterization of known and unknown peptides, but the method is costly in terms of expensive instrumentation and the time required to learn and master its use. Indeed, many scientists choose to outsource their MS experiments to dedicated centers rather than maintain in-house instruments. By contrast, antibody-based strategies such as ELISA or immunohistochemistry are easily accessible and highly flexible but offer low resolution and can only detect known targets since each target must be matched with its own specific probe.

The development of next-generation protein sequencing technology is a priority. Scientists recently established an approach where peptides are immobilized in nanoscale reaction chambers on semiconductor chips.² Here, N-terminal amino acids—exposed one-by-one by aminopeptidases—can be detected in parallel with dye-labeled recognizers in real time. This method drives the Platinum^™ Next-generation Protein Sequencer from Quantum-Si, a benchtop device capable of interrogating proteoforms, post-translational modifications, and low-abundance proteins at single-molecule resolution. This sequencer’s workflow takes less than three hours of hands-on time, and data analysis does not require bioinformatics expertise. Instruments like this make single-molecule protein sequencing possible for any lab anywhere.

References

Alfaro JA, et al. Nat Methods. 2021;18:604-17.
Reed BD, et al. Science. 2022;378(6616):186-92.

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The figure shows two waves made of DNA double helixes representing gene expression changes in the malaria parasite and its human host. These changes reveal a synchronization between parasite and host.

Article

Malaria Parasites Sync with Hosts’ Molecular Rhythms

Evidence of malaria parasites aligning with their human hosts may pave the way for new antimalarial agents.

Mariella Bodemeier Loayza Careaga, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

A mosquito makes a high-pitched buzz, followed by an unnoticeable bite at night. As a result, the Plasmodium parasite enters the human body and trips off a cascade of events that leads to malaria. To better understand the pathogenesis of this life-threatening disease,¹ Steve Haase, a biologist at Duke University, studies Plasmodium biological clocks, which are internal timekeeping mechanisms that regulate many biological processes.²

In a recent Proceedings of the National Academy of Sciences paper, Haase’s team showed that one of the most common malaria parasites, Plasmodium vivax, syncs its gene expression to that of its human host. Evidence of parasite-human host alignment could help researchers identify new targets to fight malaria.

Haase’s group previously found that the malaria parasite has a biological clock that controls its 48-hour developmental cycle inside red blood cells.³ “In mouse models, the malaria parasites do appear to be aligned with the circadian clock of a host,” he said, but scientists were not sure if this was true in humans as well.

To find out, the researchers collected blood from 10 patients infected with P. vivax. After growing the parasites in culture dishes, they tracked gene expression changes in both the parasites and host cells over two days by using RNA sequencing.

The team found that hundreds of genes in both P. vivax and their hosts followed a clock-like rhythm, upregulating and downregulating their expression throughout the day. Based on these data, the team calculated the internal clock time for each parasite-human pair. They found that genes with rhythmic expression were in sync, meaning that if a person’s gene expression shifted by a few hours, the gene expression in the corresponding parasite also shifted to match its host’s rhythm.

Research on circadian rhythms in infections is novel and important, said Filipa Rijo-Ferreira, a molecular parasitologist at the University of California, Berkeley, who was not involved in the research. “This study recapitulated observations from mouse models and seems very supportive of this potential coevolution of all these Plasmodium species into being in sync with their hosts.”

A key next step is to identify the signals that enable the parasite-host synchronicity, said Haase. “If we can understand how the parasite and the host are talking to each other and disrupt that communication, that might be a way of making the disease less severe.”

References

World Health Organization. Malaria. 2023.
Motta FC, et al. Proc Natl Acad Sci U S A. 2023;120(24):e2216522120.
Smith LM, et al. Science. 2020;368(6492):754-759.

Illustration of a group scientists in medical or chemical laboratory.

Interview| Article

From Mentee to Mentor: Teaching Undergraduates in the Lab

Mark Emerson shares his secret for establishing a fruitful research experience for students and mentors alike.

Danielle Gerhard, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

Mark Emerson, a biologist at The City College of New York, studies the gene-regulatory networks that orchestrate retinal development.

Julianna LeMieux

Early career researchers often find themselves wearing two hats: the well-worn one of the mentee and the new, heavy with responsibility one of the mentor. With teaching, writing, and research demanding their time and attention, researchers may often wonder how they can best include undergraduate researchers on projects so that both get the most out of the experience. When Mark Emerson, a biologist at The City College of New York and recipient of a Council on Undergraduate Research Mentor Award, started his lab 11 years ago, he decided right from the start that he would equip young researchers with the skills they need to become great scientists.

What is a good project for an undergraduate in the lab?

It is important not to give students busy work because they need to feel a sense of ownership. I give each of my students a project based on his or her interests, but there are other considerations as well. I choose a project that is not time sensitive and that uses established techniques in the lab so that I can easily troubleshoot any problems. The goal is for students to slowly acquire independence, so I do not need to micromanage, but ultimately, I need a way to assess the outcome. Also, it is helpful to have shorter experiments in the project that can be done in stages—running gels, purifying fragments, doing minipreps—but that occur within the context of the project. This gives students the opportunity to have project continuity and scheduling flexibility.

Why do you think it is important to train undergraduate researchers?

Training undergraduate researchers is not necessary for many labs, but I think that we have a responsibility to make these opportunities available to students to prepare them to do good science. I find it very rewarding to see what they go on to do in the lab and community. Undergraduates also add a lot of unseen value to the lab. For example, when we dive into the details of a project or a technique, it reveals what we ourselves do not understand.

This interview has been edited for length and clarity.

A 96-well microplate containing serial dilutions of stained samples.

Technology Highlight| Article

Expanding Sample Analysis While Shrinking Instrument Footprints

Cutting-edge analytical instruments provide researchers with improved analysis capabilities in a compact design.

Sep 1, 2023 | 2 min read

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Bench space in most laboratories is scarce, owing to the abundance of bulky analytical instruments, including spectrophotometers, microplate readers, and fluorometers, which can perform overlapping functions. To avoid this overlap, scientists should use a microplate reader that measures absorbance, fluorescence, and luminescence all in one instrument.

A photo of the VANTAstar® ejecting a microplate containing samples.

The VANTAstar^® is a compact, upgradable microplate reader that can measure absorbance, fluorescence, and luminescence.

BMG LABTECH

Nevertheless, microplate readers are not without their faults. Most instruments acquire measurements at the well's center point.¹ This enables quick assessment of the entire microplate, which is important for time-course analyses. However, these instruments are inadequate for measuring heterogeneous samples or wells with nonhomogenous distributions. This highlights the need for microplate readers that scan the entire well and acquire multiple measurements.¹

Different microplate readers vary in their capabilities and features, which makes choosing the correct instrument challenging. Although scientists save money by buying microplate readers with fewer features, they may need to upgrade or buy a new instrument before the end of its life span if it no longer meets their needs.

Researchers can avoid these challenges by using a modular system such as the VANTAstar® microplate reader. In addition to its multimodal capabilities, well-scanning features, and ease of use, researchers can add additional components to the instrument, including environmental controls, reagent injectors, and shaking capabilities. Despite its small footprint, this instrument is compatible with microplate formats up to 384 wells, saving researchers time, space, and money. Compact microplate readers like this one provide scientists with the flexibility they need to answer new questions.

Reference

Pryor SW, et al. BioTechniques. 2007;42(2):168-172.

Illustration showing how this new novel nanotechnology simultaneously ‘fishes’ for multiple viruses or viral variants using different DNA nanobait that are designed to target specific viral sequences.

Article

Fishing for Viruses With DNA Nanobait

Scientists developed a novel nanotechnology that simultaneously detects multiple viruses from patient samples in less than an hour.

Danielle Gerhard, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

MODIFIED FROM © ISTOCK.COM, VikiVector; © ISTOCK.COM, urfinguss; © ISTOCK.COM, colematt; Designed by Erin Lemieux

A schematic showing the new DNA nanobait technology and associated experimental procedure.

Ulrich Keyser and his team designed their DNA nanobait technology to hunt down short viral RNA targets directly from patient samples.

License CC-BY 4.0, Bošković F, et al. Nat Nanotechnol. 2023;18(3):290-298.

Viral infections spread fast, but detection methods lag behind. Now, researchers at Cambridge University developed a novel nanotechnology that simultaneously detects multiple viruses directly from a small sample volume, which they hope will make a splash in rapid diagnostics.¹

Ulrich Keyser, a biophysicist from Cambridge University and coauthor of the paper, previously used nanopore sensors to detect isolated short nucleic acid species.² Like an electrocardiogram recording electrical signals from the heart, nanopore sensors transform biological signals into electrical currents. When molecules swim through the nanopore’s ionic current, they create ripples that carry information about their identities.

To adapt this technology for diagnostics, the team developed a one-pot approach that starts with the enzyme ribonuclease H, guided by DNA oligos, to selectively cut short sections of RNA that uniquely identify specific viruses.

To catch these viral RNA fragments, Keyser and his team developed DNA nanobaits. Using single-stranded bacteriophage DNA like a fishing rod, the researchers hung multiple lines baited with bespoke complementary DNA oligos, or nanobaits, bound to bulky placeholder oligos. Drawn to their complementary nanobaits, viral RNAs displaced the larger placeholder oligos and bound to the bait.

As the baited fishing rod flew through the nanopore sensor, it displaced ions, with larger molecules creating larger ripples in the ionic current. Unlike previous efforts that use additive signals for viral detection, Keyser’s approach causes a reduction in signal. “That is not only really innovative, but can be very powerful diagnostically because it reduces false positives,” said Adam Hall, a biomedical engineer at Wake Forest University School of Medicine who was not involved in the study.

The researchers used DNA nanobaits to fish for different respiratory viruses, including SARS-CoV-2 variants. “The nice thing about the test is that by adding more oligos, you can select for more pieces of the same viral genome for variant detection, or you can detect different viruses,” said Keyser.

References

Bošković F, et al. Nat Nanotechnol. 2023;18(3):290-298.
Bell NA & Keyser UF. Nat Nanotechnol. 2016;11(7):645-651.

Epic Fail| Article

The Oligo Was a No Go

As soon as Melanie McConnell added the wash buffer to her sample, she knew she had made a mistake.

Danielle Gerhard, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

Modified from © ISTOCK.COM, FoxysGraphic

Melanie McConnell conducted her graduate research at the University of Otago. She is now a professor at Victoria University of Wellington.

Victoria University of Wellington

In the mid-1990s, I was a first-year graduate student at the University of Otago in Michael Eccles’ laboratory. Our group studied transcriptional regulation in pediatric kidney cancer, and I was tasked with determining the DNA-binding site of a transcription factor involved in kidney development.

To do this, we decided to build a large oligonucleotide. My department had just purchased a brand-new oligonucleotide synthesizer—the first one in New Zealand. It was really complicated and took us ages, but we eventually designed a 100 base pair oligonucleotide. As the synthesis was going to take a few days, we programmed the machine to run over the weekend.

When I arrived in the lab on Monday, I was excited to validate the oligonucleotide. But first, I needed to wash the oligonucleotide to remove the chemicals that were added during synthesis. That’s when it all went wrong.

I’ll never forget that moment. As soon as I added the wash buffer to the spin column, I watched the tube and my precious oligonucleotide melt into a puddle of plastic. I misread the protocol. Instead of adding the wash buffer containing 0.1% trifluoroacetic acid (TFA), I accidentally added 100% TFA!

I was dumbstruck. The pungent fumes snapped me back to reality, and I sheepishly ventured to my advisor’s office to break the news. I knew that this was an expensive mistake. The synthesis cost NZ $1000, which was a lot of money at the time. After a long slow inhale, he said, “Okay, we can make another one.”

I learned two important lessons that day. First, read protocols and chemical safety sheets very carefully! Second, failure in science is normal and not a reflection of my potential. I wasn’t discouraged. We made more oligonucleotides, but eventually switched to a different technique and went on to discover the binding site for the transcription factor.¹ I still study regulation of gene expression in cancer, but now as a professor at Victoria University of Wellington.

This interview has been edited for length and clarity.

Reference

McConnell MJ, et al. Oncogene. 1997;14:2689-2700.

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Infographic

A New Kind of MINFLUX

MINFLUX brought record-breaking resolution to fluorescence microscopy in 2016. A new version perfected for protein tracking came out this spring.

Ida Emilie Steinmark, PhD

Sep 1, 2023 | 1 min read

Infographic showing the difference between the classic MINFLUX and the updated MINFLUX

References

Balzarotti F, et al. Science, 2016;355(6325): 606–612.
Wolff JO, et al. Science, 2023; 379(6636): 1004-1010.

A yellow, hairy caterpillar is sitting on a green leaf off a thin plant stem. — Microbes help plants fend off caterpillars.

Interview| Article

Deciphering Plants’ Biochemical Messages

Esther Ngumbi believes that chemical signals between plants, microbes, and insects hold the key to secure and sustainable food production.

Ida Emilie Steinmark, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

Photo of Esther Ngumbi, an entomologist at the University of Illinois Urbana-Champaign.

Esther Ngumbi, an entomologist at the University of Illinois Urbana-Champaign, studies chemical pathways in plants.

Jerri Caldwell Hammonds

Born in Kenya to teachers who farmed on the side to keep her in school, Esther Ngumbi has been fascinated by the relationships between plants, insects, and microbes since childhood. Now an expert on biochemical communication at the University of Illinois Urbana-Champaign, she strives to show how the chemical messaging that underpins interspecies interactions could help future-proof food production.

What’s your favorite example of how plants communicate biochemically with their surroundings?

When plants are attacked by insects, they recruit beneficial soil microbes by sending chemical signals below ground. The microbes help the plants activate their defenses in exchange for sugars. For example, they upregulate the Jasmonic acid pathway, which is critical for insect defense.¹ This cooperation helps fend off caterpillars. To me, that is amazing!

How does your work on chemical signals connect to food security and sustainability?

We have African projects that model this well. People border their fields with plants that release natural chemical signals to attract herbivorous insects and lure them away from the crop plants. Among their crops, they have other plants that push nutrients to beneficial microorganisms to keep them close. In fact, we can genetically engineer new plant varieties that do these jobs especially well, and we can design disease-suppressing soil by including specific microbes. Underground communities like rhizobacteria that live on plant roots can also mediate drought tolerance, for example, through enzymes and metabolites that they pass on to the plants.²

So, we can exploit the natural biochemical communication between organisms to design ecosystems that provide food sustainably without lots of fertilizers and pesticides. However, we must keep on our toes because we are dealing with insects and climate change. That keeps me grounded but also excited and connected to the work that I do.

This interview has been condensed and edited for clarity.

References

Disi JO, et al. Arthropod-Plant Interact. 2017;11:591-602.
Ngumbi E, Kloepper J. Appl Soil Ecol. 2016;105:109-125.

Article

A Stem Cell Zoo Reveals Surprising Differences in Embryogenesis

By comparing stem cells from six mammals of different sizes, scientists discovered stark differences in embryonic development paces.

Ida Emilie Steinmark, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

Human embryos take much longer to develop than mouse embryos.¹ Scientists thought that this was because humans are larger, but a new study has overturned that idea.² Researchers from the European Molecular Biology Laboratory found that species-specific gene expression likely regulates biochemical reaction speeds and consequently the developmental tempo.

The team first secured stem cells from rhinoceroses to test an animal bigger than humans. Because gestation includes both embryogenesis and a phase of fetal growth, the researchers had no clue if rhinos’ long gestation reflected their embryonic development speed. As the stem cells differentiated in vitro, the researchers followed the developmental pace through the expression of the segmentation clock,a set of genes that creates segments in the embryo from which the vertebrae, ribs, and skeletal muscles eventually form. The genes are expressed in waves, where each wave crest creates a segment.

They saw that rhino development was faster than that of humans. “That was very shocking,” said coauthor developmental biologist Jorge Lázaro. “We thought it was going to be super slow.” Testing rabbits, cattle, and marmosets confirmed that development pace didn’t correlate with body weight.

This is an image of a bioluminescent from gene expression reporter in stem cells from a rhinoceros.

Jorge Lázaro and his team measured the oscillation of the segmentation clock through a bioluminescent reporter, shown in this image of induced pluripotent stem cells from a rhinoceros.

European Molecular Biology Laboratory

Instead, the researchers found that the faster the segmentation clock ticked, the quicker reaction rates were for key embryogenesis proteins. The expression of thousands of genes involved in biochemical processes such as nuclear transport and RNA processing also correlated with segmentation clock speed. This corroborated the team’s earlier findings in mice and humans.¹ Consequently, they proposed that species-specific transcriptomic profiles control biochemical reaction speeds and ultimately developmental pace.

“The lengths they’ve gone to to analyze this evolutionary question is really beyond what anyone’s done before,” said developmental biologist Andrew Oates from École Polytechnique Fédérale de Lausanne, who was not involved in the study. However, to him, pinning the mechanism on “genes related to biochemical reactions” is too vague. “It’s still rather mysterious. [Although] probably the answer is in that set of genes.”

Lázaro might soon provide clarity. He plans to tweak the gene expression and speed up development in the stem cells. That should elucidate the mechanism and yield a blueprint for accelerating development elsewhere, for instance in organoids, some of which can take months to mature.

References

Matsuda M, et al. Science. 2020;369(6510):1450-1455.
Lázaro J, et al. Cell Stem Cell. 2023;30(7):938-949

Digital illustration of a brain, constructed by tiny dots and lines. Most dots and lines are teal-colored; others are green, yellow, red, and purple to denote areas of activity.

Just Curious| Article

What Was the First Animal to Evolve a Brain?

In the absence of a precise definition of brain, pinning down its origins is difficult. But scientists have a theory.

Ida Emilie Steinmark, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

Having a brain is so necessary to human experience that it’s almost impossible to imagine any life without it. However, many living organisms don’t have brains, and going back far enough in time will lead to an ancestor of our own that was equally brainless. So, when exactly did brains evolve?

That’s a difficult question, said evolutionary developmental biologist Sebastian Shimeld at the University of Oxford, because it’s hard to even distinguish between who has a brain and who doesn’t. “We don’t have a precise definition of a brain,” he said.

Scientists home in on the brain’s evolutionary origins by sorting out the animals without brains. Sponges have no neurons, so they are easy to discount, and while the more sophisticated jellyfish and sea anemones have a network of neurons, they have no central neural “headquarters” characteristic of a brain.

About 600 million years ago, another group of animals evolved that had bilateral symmetry, meaning that they had a front and a back. “The front is where the nervous system crystallizes because that’s the bit of the animal that’s meeting the environment head on,” Shimeld said. The first brain-like mass of neurons likely evolved at the front end of a long, thin, worm-like animal. “Everything else that descended from that is a descendent of that neural structure, we think.”

Today, there are many species, including some invertebrates such as the octopus, with brains that work similar to ours. These brains control perception, behavior, and higher functions like memory. They are complex and wondrous, and they all evolved from a clump of neurons in the head of a worm.

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Note: This article has been updated to correct a typo.

Human small and large intestines internal organ ?3D illustration.

Article

How Stress Inflames the Gut

In mice, chronically high levels of stress hormones worsen bowel inflammation.

Mariella Bodemeier Loayza Careaga, PhD

Sep 1, 2023 | 2 min read

Above Image Credit:

In 2017, 6.8 million people worldwide were affected by inflammatory bowel disease (IBD), a group of disorders that causes prolonged intestinal inflammation.¹ Scientists knew that chronic stress worsens IBD symptoms, but how exactly that happens was unclear.

In a recent Cell publication, Kai Markus Schneider, a physician scientist at RWTH Aachen University, and his colleagues reported that naturally occurring hormones secreted during stressful events, glucocorticoids, affect cells in the enteric nervous system (ENS) to cause intestinal inflammation and bowel problems.²

To investigate how stress worsens IBD symptoms, the team gave mice a chemical that damages their guts to simulate IBD and then stressed half of them by restricting their movement for seven to ten days. Stressed IBD mouse models showed increased inflammation compared to IBD mouse models without stress.

The image shows round-shaped glial cells in red and elongated neuronal cells in green surrounding the glial cells.

Researchers labeled the neurons (red) and glia cells (green) in the mouse gut nervous system.

Markus Schneider, Klaas Bahnsen, and Niklas Blank

“There are different pathways that can propagate the stress response into the guts,” Schneider explained. The release of glucocorticoids is one of them. When the researchers modulated glucocorticoid levels, they found that elevated levels worsened IBD inflammation, whereas reducing the levels prevented that effect.

Since glucocorticoids affect neuronal cells, the team next investigated how stress changed the gene expression in ENS cells. When stressed, enteric glia cells showed increased proinflammatory pathways, which worsened bowel inflammation by attracting more white blood cells. The team also found that stress increased the proportion of precursor-like neurons relative to mature neurons in the ENS, a shift that impaired the bowel’s movements.

Keith Sharkey, a gastrointestinal physiologist at the University of Calgary who was not involved in the research, said that the study provides a detailed mechanistic explanation for how stress affects the bowels. He also expressed curiosity about sex differences in those findings. “There is a clear difference in the way male and female [subjects] respond to stress, and we know that sex hormones are important in protection against colitis.”

The findings might point to ways to improve patient care, Schneider added. “Stress management is also essential for the management of IBD. We should offer this to patients, and patients should not be afraid to ask for psychological support to reduce stress levels.”

References

GBD 2017 Inflammatory Bowel Disease Collaborators. Lancet Gastroenterol Hepatol. 2020;5(1):17-30.
Schneider KM, et al. Cell. 2023;186(13):2823-2838.e20.