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16th September 2024Protein Changes in the Brain Could Explain How Neurodevelopment Goes Awry
Aprocess as intricate as brain development requires precisely controlled expression of crucial genes, but it’s the different flavors of a protein, or isoforms, that the genes encode that do the heavy lifting.
“One thing that hadn’t been systematically studied is how those genes [involved in brain development] are spliced into different transcripts,” said Michael Gandal, a physician scientist at the University of Pennsylvania. “We think of genes as the main output of our genome, but they really aren’t. It’s the specific transcript of a gene and how it gets spliced together that ultimately is what makes a functional protein or a regulatory molecule.”
While researchers have shown how isoform diversity regulates synapse formation in the brain and contributes to the risk of developing neuropsychiatric disorders, less is known about their role in shaping neurodevelopment. Now, in a paper published in Science, Gandal and his team sequenced regions of the developing brain and discovered a number of different types of isoforms linked with neurodevelopmental disorders. These findings provide a framework that allows scientists to understand how isoforms and the way they change could potentially orchestrate and influence neurodevelopment.
“We were quite interested in understanding how genes regulate [development], how that process may go awry in individuals with neurodevelopmental disorders, and whether we can connect the two together through this study,” said Gandal.
Previous attempts to study isoforms in neurodevelopment used a technique called short-read sequencing. But this approach failed to provide a complete picture of the isoforms since they showed up as small fragments that the researchers had to stitch together into a meaningful picture. Gandal and his team bypassed this tedious task by using long-read sequencing, which gives researchers the ability to sequence entire, uninterrupted genetic sequences. With this approach, the researchers set out to get a better picture of the different isoforms during a critical window of development.
ByClaudia Lopez-Lloreda, PhD
Article can be accessed on: The Scientist
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10th September 2024With AI, extreme microbe reveals how life’s building blocks adapt to high pressure
High-pressure limited proteolysis (Hi-P LiP) deeply interrogates protein structure under pressure. Credit: PRX Life (2024). DOI: 10.1103/PRXLife.2.033011 An assist from a Google Artificial Intelligence tool has helped scientists discover how the proteins of a heat-loving microbe respond to the crushing conditions of the planet’s deepest ocean trenches, offering new insights into how these building blocks of life might have evolved under early Earth conditions.
The findings, published in PRX Life, will likely prompt further studies into the inner workings of proteins and life on other planets, and serve as a successful case study on how artificial intelligence was able to accelerate such research by decades.
“This work gives us a better idea of how you might design a new protein to withstand stress and new clues into what types of proteins would be more likely to exist in high-pressure environments like those at the bottom of the ocean or on a different planet,” said Stephen Fried, a Johns Hopkins University chemist who co-led the research.
Fried’s team subjected Thermus thermophilus—a microorganism widely used in scientific experiments owing to its ability to withstand heat—to lab-simulated pressures mimicking those of the Mariana Trench. The tests revealed some of its proteins resist those stress levels because they have a built-in flexibility with extra space between their atomic structures, a design that allows them to compress without collapsing.
By Johns Hopkins University
Article can be accessed on: phys.org
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10th September 2024A Gene Editing System Corrects Cystic Fibrosis Mutation
In cystic fibrosis (CF), patients’ organs become overwhelmed by thick mucus, which affects breathing and can result in serious bacterial infections. The outlook for patients with CF has improved significantly since scientists identified the gene mutation responsible for the disease in 1989. A majority of cases are caused by a three-base-pair mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. This mutation, called F508del, results in ion channel defects throughout the patients’ bodies.
Gene therapies have huge potential for diseases like CF because they provide long-lasting treatments. However, genetic editing techniques used to develop these promising treatments, including the Nobel Prize-winning CRISPR-Cas9, have proved inefficient at correcting F508del. Now, published in Nature Biomedical Engineering, researchers used an optimized version of the genome editing technology prime editing to alter this mutation in lung cells from patients with CF. The approach restored function to the same degree as a powerful CF drug combination therapy. The findings could lead to CF treatments that permanently correct the disease’s genetic origin.
David Liu, a geneticist at the Broad Institute of MIT and Harvard and coauthor on the study, recognized that complex genetic diseases require flexible and precise gene therapies. In 2019, he developed prime editing, a gene editing technique that takes a different approach from CRISPR. The latter technique cuts the genome like scissors, snipping through both strands of the DNA, which can lead to unintended, off-target edits. In contrast, prime editors, said Liu, are like “DNA word processors in that we develop them to do true search and replace editing.” He added that this allows the technology to make more precise and controllable changes, making it ideal for the repair job needed to correct the F508del mutation: the addition of only a few DNA letters at specific locations.
In their 2019 paper, Liu’s team used prime editing to alter the gene mutations causing sickle cell disease and Tay-Sachs disease. Liu explained that they attempted to edit mutated CFTR as well but with limited success. “That was a very difficult correction to make,” said Liu. The genetic machinery that guided the prime editor proved unstable and corrected less than one percent of mutations.
Researchers have made improvements to prime editing over the last five years. In the latest paper, Liu’s team took another crack at editing CFTR using what he calls a “kitchen sink approach” that utilized six major enhancements to prime editing. These changes improved the accuracy and stability of the molecular machinery.
Liu said much effort went into incorporating an enhancement that allowed the system to make edits to the DNA without attracting the attention of cells’ built-in repair pathways. This protein machinery—the DNA mismatch repair system—reversed the prime editor’s changes if it noticed the rewriting. By adding in silent mutations near the edits they made to F508del, Liu’s team found that the mismatch repair system was less likely to undo edits.
By RJ Mackenzie
Article can be accessed on: The Scientist
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5th September 2024RNA editing plays critical role in fruit flies’ sense of smell and social interactions
The recoding of Ile27 to Val is conserved across Drosophila species. Credit: Science Advances (2024). DOI: 10.1126/sciadv.adi9101 Imagine your DNA as a set of instructions or a recipe book that tells your body how to make everything it needs to function, from proteins to cells. Every time the body needs to build something, it reads these instructions. But sometimes, the body can make small edits to these instructions—this is where RNA editing comes in.
RNA editing is like a proofreading process that happens after your DNA’s instructions are copied. Instead of just following the recipe exactly, your cells can make tiny changes to the instructions. These changes can help the body adapt to different situations by creating new versions of proteins that might be better suited to certain tasks.
In humans and fruit flies alike, RNA editing prevents autoimmune responses and adjusts protein functions. However, in humans, most editing occurs in non-coding regions, with only a small fraction leading to changes in protein function. In contrast, in flies, the majority of RNA editing events occur in sequences that directly produce proteins.
Given the abundance of RNA editing events that lead to changes in protein-coding sequences in flies, a major challenge is determining which of these thousands of events is biologically important and worth investigating.
Researchers from Bar-Ilan University in Israel have now pinpointed one such event and determined its pivotal role in the sense of smell and social interactions of Drosophila (fruit flies). The findings of their study appear in the journal Science Advances.
By Bar-Ilan University
Article can be accessed on: phys.org
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5th September 2024Tiny Test Tubes Sort Stem Cells for Improved Therapy
With the power of self-renewal and multipotency, mesenchymal stem cells (MSCs) offer the possibility of regenerative treatment for a variety of medical conditions, including spinal cord injury and damage following a heart attack.
A leading hypothesis is that the therapeutic benefits of MSCs are conferred by their secretion of extracellular vesicles (EVs): membrane-derived structures that contain a variety of important bioactive molecules. Therapeutic cells like MSCs have variable EV production, contributing to inconsistent outcomes that hinder their clinical translation. Yet, scientists are unable to select cells based on their EV secretion levels.
“How do you know how many extracellular vesicles the [MSCs are] secreting? How do you know this batch of cells is better than this batch?” said Dino Di Carlo, a bioengineer at the University of California, Los Angeles.
In a new study published in Nature Communications, Di Carlo and his colleagues applied their expertise in microfluidics and nanotechnology to answer these questions. They developed a method to identify subpopulations of MSCs that secrete high levels of EVs, an approach that could allow for the selection of more therapeutically active cells for clinical applications.
Previously, Di Carlo and his team developed hydrogel-based microcontainers, which they called nanovials, that act as test tubes for single cells. By coating the inside of the bowl-shaped nanovials with antibodies that are specific to proteins released by a cell of interest, the researchers could capture individual cells and quantify their secretions. In an earlier study, the team used this technology to measure protein secretion from individual B cells, which they then linked to cell surface markers and gene expression data from the same cell.
By Rebecca Roberts, PhD
Article can be accessed on: The Scientist
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27th August 2024Closing the RNA loop holds promise for more stable, effective RNA therapies
Engineering ocRNAs and icRNAs. Credit: Nature Biomedical Engineering (2024). DOI: 10.1038/s41551-024-01245-z New methods to shape RNA molecules into circles could lead to more effective and long-lasting therapies, shows a study by researchers at the University of California San Diego. The advance holds promise for a range of diseases, offering a more enduring alternative to existing RNA therapies, which often suffer from short-lived effectiveness in the body.
The work is published in Nature Biomedical Engineering.
RNA molecules have emerged as powerful tools in modern medicine. They can silence genes through small interfering RNAs (siRNAs) or serve as templates for making therapeutic proteins, as seen with messenger RNAs (mRNAs). Unlike gene editing technologies, which make permanent changes to DNA, RNA therapies offer a temporary but highly targeted approach.
However, one major challenge is that RNAs do not last long in the body, which limits their effectiveness. The concept of circular RNAs (cRNAs) has gained traction as a solution to this challenge. Circular RNAs, unlike their linear counterparts, have a closed-loop structure that renders them more resistant to degradation. The problem is that existing methods for creating circular RNAs are complex and inefficient.
By Liezel Labios, University of California – San Diego
Article can be accessed on: phys.org
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27th August 2024Rebuilding the Genome of Woolly Mammoths
Over time, DNA can degrade into numerous short fragments that wiggle around. Eventually, the chromosome loses its original structure and genetic information to the ravages of time. However, researchers pondered whether fossil chromosomes that retained their original three-dimensional configuration existed.
To find out, an international team of researchers set off on an expedition in the frosty tundra of Siberia. In 2018, they discovered a promising, well-preserved skin sample from a giant titan of the Ice Age: a 52,000-year-old woolly mammoth.
Using a high-throughput technique to capture chromatin conformation (Hi-C) and DNA sequencing, molecular geneticists Erez Aiden and Olga Dudchenko at Baylor College of Medicine reassembled the three-dimensional structure of DNA and its genome—the first reconstruction of its kind. These findings, published in Cell, uncovered new biology and ushered in an exciting new chapter in paleogenomics.
Sampling from Fossil Chromosomes to Uncover Genetic Insights
Typically, ancient DNA fragments yield short snippets of DNA and provide an incomplete picture of the genomic puzzle. However, the woolly mammoth skin sample showed promise. Dudchenko observed that the sample was exceptionally well-preserved from macroscopic down to nanometer scales, with no molecular movement, and could potentially contain a more complete genomic picture.
To explore the 3D morphology of the existing chromosomes, Aiden modified an in situ Hi-C protocol, dubbed PaleoHi-C. This technique, adapted for ancient samples and combined with DNA sequencing, enabled the researchers to map the chromosome features, assemble a reference genome, and determine activation patterns across genes.
With this data, the team assembled the first 3D reconstruction of the woolly mammoth’s genome, which had 28 chromosomes; the order of genes was very similar to that of the Asian elephant. Then, upon analyzing the X chromosomes, they confirmed the specimen was a female woolly mammoth. They also observed inactive X chromosome (Xi) superdomains, a pattern resembling the bipartite architecture seen in humans and mice. However, to the researchers’ surprise, they uncovered new biology. Dudchenko noted that the mammoth “was a bit of an overachiever”, as its Xi exhibited a tetradic architecture. This pattern is also seen in modern elephants.
Next, they compared the degree of transcriptional activity of DNA between mammoth and elephant tissue. “We saw interesting differences in the genes related to hair follicle development and, more broadly, hair maintenance,” Dudchenko remarked.
By Laura Tran, PhD
Article can be accessed on: The Scientist
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20th August 2024Biochemists engineer proteins that can transition between assembly and disassembly via allosteric control
Design strategy for building switchable oligomers. Credit: Nature (2024). DOI: 10.1038/s41586-024-07813-2 A team of biochemists at the University of Washington has developed a means for engineering proteins that can transition between assembly and disassembly via allosteric control. In their paper published in the journal Nature, the group details their engineering process and how well it has worked thus far during testing.
A. Joshua Wand, with Texas A&M University, has published a News and Views piece in the same journal issue explaining why being able to get a protein to assemble or disassemble in the presence of an effector has been an important goal of chemists and outlines the work done by the team on this new effort.
Prior research has shown that if chemists could build proteins that assemble themselves into desired shapes on command, the results could be used for highly specific purposes, such as building a cage-shaped protein to carry a drug to a certain part of the body for therapeutic purposes.
Prior research has also shown that if such a mechanism could be developed, via a process known as allosteric regulation, a specific trigger would be needed—one that chemists refer to as an effector. To achieve this feat, the research team used several techniques they previously developed.
One such technique involved using an AI app to help predict a protein structure given a list of attributes. Another involved creating a protein with a hinge that was able to take on two different forms. And a third involved a way to connect certain proteins together.
By Bob Yirka
Article can be accessed on: phys.org
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20th August 2024Fast Tracking Protein Delivery into the Nucleus
Rush hour never ceases at the nucleus’ border. Gene products begin their lives in the nucleus as strands of mRNA that ship out into the cytoplasm, where they serve as templates for protein synthesis. Many of these proteins, such as transcription factors, subsequently return to sender, crossing back into the cell’s central organelle. To traverse the nuclear envelope, proteins must transit through a nuclear pore complex (NPC). This protein channel serves as a gatekeeper to the nucleus, restricting passage to select proteins that carry a nuclear localization signal (NLS)—an amino acid sequence that stamps the protein for delivery to the nucleus.
In a recent study published in Nature Physics, researchers found that proteins harboring a flexible domain near the NLS enter the nucleus faster. To mimic this limber protein element, the biophysicists designed a bendable protein tag that expedited delivery of protein cargo into the cell’s core organelle.
“People are certainly looking at how to deliver various therapeutics, diagnostics, or just simply research tools to the nucleus, and this could be rather important in very significantly improving the efficiency of that process,” said Michael Rout, a cell biologist at the Rockefeller University who was not involved with the work.
Scientists previously found that the NPC shapeshifts to allow cargo to cross the nuclear threshold, but scientists know little about how structural alterations to the protein parcels themselves affect transport. “Cargoes have been to some degree seen as like the corpse at a funeral—they’re the purpose for the whole ceremony, but they don’t take an active part in the process,” said Rout.
To study the relationship between a protein’s molecular makeup and its movements, Sergi Garcia-Manyes, a biophysicist at the Francis Crick Institute and study coauthor, developed a system to time protein transport into the nucleus. He and his team chose a common protein motif called the immunoglobulin domain (Ig) as their test subject. They worked with two Ig mutants: one that was more flexible and another that was more rigid compared to wild type Ig. However, Ig domains don’t have an NLS, so the researchers gave each version the nuclear postage stamp. Fusing these mutant constructs to a fluorescent protein allowed the researchers to monitor protein distribution under a microscope and time their shipments to the nucleus. The researchers were ready to take their souped-up proteins to the races. When they pitted the mutants against one another, they found that the flexible Ig domain took less time to enter the nucleus than the stiff variety.
By Kamal Nahas, PhD
Article can be accessed on: The Scientist
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13th August 2024Chemists synthesize plant-derived molecules that hold potential as pharmaceuticals
The oligocyclotryptamines were originally isolated from Psychotria leaves in New Caledonia. Credit: Jose-Luis Olivares, MIT; molecule provided by Scott et al. MIT chemists have developed a new way to synthesize complex molecules that were originally isolated from plants and could hold potential as antibiotics, analgesics, or cancer drugs.
These compounds, known as oligocyclotryptamines, consist of multiple tricyclic substructures called cyclotryptamine, fused together by carbon–carbon bonds. Only small quantities of these compounds are naturally available, and synthesizing them in the lab has proven difficult. The MIT team came up with a way to add tryptamine-derived components to a molecule one at a time, in a way that allows the researchers to precisely assemble the rings and control the 3D orientation of each component as well as the final product.
“For many of these compounds, there hasn’t been enough material to do a thorough review of their potential. I’m hopeful that having access to these compounds in a reliable way will enable us to do further studies,” says Mohammad Movassaghi, an MIT professor of chemistry and the senior author of the new study.
In addition to allowing scientists to synthesize oligocyclotryptamines found in plants, this approach could also be used to generate new variants that may have even better medicinal properties, or molecular probes that can help to reveal their mechanism of action.
By Anne Trafton, Massachusetts Institute of Technology
Article can be accessed on: phys.org