A trillion scents, one nose

The mammalian nose is a work of evolutionary art. Its millions of nerve cells, each tailored with just one of thousands of specific odor-chemical receptors encoded in the genome, can collectively distinguish a trillion distinct scents. Those sensations, in turn, inform many behaviors, from assessing food options to discerning friends from foes to sparking memories.

Today, in the journal Nature, a research team led by scientists at Columbia's Zuckerman Institute describes a previously undetected mechanism in mice -- starring the genetic molecule RNA -- that could explain how each sensory cell, or neuron, in mammalian noses becomes tailored to detect a specific odor chemical.

For example, there are sensory neurons in our noses that bear receptors uniquely tuned to detect ethyl vanillin, the main odorant in vanilla, and other cells with receptors for limonene, lemon's signature odorant.

"How sensory cells in the nose make their receptor choices has been one of the most vexing mysteries about olfaction," said Stavros Lomvardas, PhD, a Roy and Diana Vagelos Professor and Chair of Biochemistry and Molecular Biophysics and Herbert and Florence Irving Professor of Neuroscience at Columbia's Zuckerman Institute and the Vagelos College of Physicians and Surgeons, and corresponding author on the paper.

"Now, the story behind our sense of smell, or olfaction, is becoming clearer, and also more dramatic."

The sense-refining drama he is referring to unfolds entirely within the minuscule confines of each olfactory neuron's nucleus, where the cell's chromosomes and genes reside.

There, in a Squid Games-style, winner-takes-all competition, a developing cell's myriad olfactory receptor genes vie with each other in a process that winnow them down, in stages, first to handful of finalists and then to a single winner.

The prevailing gene is the one that determines the cell's odorant sensitivity.

In their study, Dr. Lomvardas and his team uncover details of the final stage of this process when the winner emerges from the finalist genes.

"It's basically a battle between a 1000 contenders," said Ariel Pourmorady, the paper's first author and an M.D.-Ph.D. candidate at the Zuckerman Institute in the Lomvardas lab.

The action is exceedingly complex and involves a dizzying cast of molecular characters.

Playing roles that either dial up or down each gene's ability to produce olfactory receptors are a variety of gene-regulating molecules.

By gathering into various alliances within the genome, these molecular players help turn specific genes on or off.

Also in the fray is another set of molecular hubs that reshape portions of the genome in ways that favor specific receptor genes.

When his team first observed these in the genome in 2014, Dr. Lomvardas dubbed them "Greek Islands" because they reminded him of islands in the Aegean Sea.

"It turns out that the genome has a certain spatial organization in the nucleus and changes in this structure are pivotal when it comes to which genes are expressed into proteins, like olfactory receptors," said Pourmorady.

"We are learning just how important this process is within maturing olfactory cells."

In their new Nature paper, the researchers summon a trove of data from mouse studies pointing toward RNA as the linchpin molecule in the olfactory system's gene-choosing mechanism.

RNA is most known as the go-between molecule that translates the genetic code embodied in DNA into protein molecules with specific cellular jobs, like detecting odorants.

Using sophisticated techniques for analyzing changes in genome structure as cells mature, however, the researchers say their evidence points to a pivotal second role for the RNA.

"It looks like the RNA the cell makes during gene expression also is altering the genome's architecture in ways that bolster the expression of one olfactory receptor gene while also shutting down all the others," Pourmorady said.

Big gaps in this genome-controlling story remain, but the researchers say the outline

is becoming more defined. It starts with maturing olfactory cells, which initially express many receptor genes at those genomic hubs where gene-regulating molecules and complexes, including Greek Islands, converge.

Then the RNA winnows the contending olfactory-receptor genes down to one.

The particular hub in each cell where the molecular stars align to produce the highest amount of RNA wins the competition.

At this hub, receptor-gene expression soars. But, like a slinky saboteur, RNA from that same hub may wind its way to all the other hubs.

In those locations, the RNA causes shape changes in the genome that shut down gene expression.

The result is a nose's worth of mature olfactory neurons, each of which bears on its surface only one odorant receptor.

Read more at Science Daily

In a first for 'sonogenetics,' researchers control mammalian cells with sound

Salk scientists have engineered mammalian cells to be activated using ultrasound. The method, which the team used to activate human cells in a dish and brain cells inside living mice, paves the way toward non-invasive versions of deep brain stimulation, pacemakers and insulin pumps. The findings were published in Nature Communications on February 9, 2022.

"Going wireless is the future for just about everything," says senior author Sreekanth Chalasani, an associate professor in Salk's Molecular Neurobiology Laboratory. "We already know that ultrasound is safe, and that it can go through bone, muscle and other tissues, making it the ultimate tool for manipulating cells deep in the body."

About a decade ago, Chalasani pioneered the idea of using ultrasonic waves to stimulate specific groups of genetically marked cells, and coined the term "sonogenetics" to describe it. In 2015, his group showed that, in the roundworm Caenorhabditis elegans, a protein called TRP-4 makes cells sensitive to low-frequency ultrasound. When the researchers added TRP-4 to C. elegans neurons that didn't usually have it, they could activate these cells with a burst of ultrasound -- the same sound waves used in medical sonograms.

When the researchers tried adding TRP-4 to mammalian cells, however, the protein was not able to make the cells respond to ultrasound. A few mammalian proteins were reported to be ultrasound-sensitive, but none seemed ideal for clinical use. So Chalasani and his colleagues set out to search for a new mammalian protein that made cells highly ultrasound sensitive at 7 MHz, considered an optimal and safe frequency.

"Our approach was different than previous screens because we set out to look for ultrasound-sensitive channels in a comprehensive way," says Yusuf Tufail, a former project scientist at Salk and a co-first author of the new paper.

The researchers added hundreds of different proteins, one at a time, to a common human research cell line (HEK), which does not usually respond to ultrasound. Then, they put each cell culture under a setup that let them monitor changes to the cells upon ultrasound stimulation.

After screening proteins for more than a year, and working their way through nearly 300 candidates, the scientists finally found one that made the HEK cells sensitive to the 7 MHz ultrasound frequency. TRPA1, a channel protein, was known to let cells respond to the presence of noxious compounds and to activate a range of cells in the human body, including brain and heart cells.

But Chalasani's team discovered that the channel also opened in response to ultrasound in HEK cells.

"We were really surprised," says co-first author of the paper Marc Duque, a Salk exchange student. "TRPA1 has been well-studied in the literature but hasn't been described as a classical mechanosensitive protein that you'd expect to respond to ultrasound."

To test whether the channel could activate other cell types in response to ultrasound, the team used a gene therapy approach to add the genes for human TRPA1 to a specific group of neurons in the brains of living mice. When they then administered ultrasound to the mice, only the neurons with the TRPA1 genes were activated.

Clinicians treating conditions including Parkinson's disease and epilepsy currently use deep brain stimulation, which involves surgically implanting electrodes in the brain, to activate certain subsets of neurons. Chalasani says that sonogenetics could one day replace this approach -- the next step would be developing a gene therapy delivery method that can cross the blood-brain barrier, something that is already being studied.

Perhaps sooner, he says, sonogenetics could be used to activate cells in the heart, as a kind of pacemaker that requires no implantation. "Gene delivery techniques already exist for getting a new gene -- such as TRPA1 -- into the human heart," Chalasani says. "If we can then use an external ultrasound device to activate those cells, that could really revolutionize pacemakers."

For now, his team is carrying out more basic work on exactly how TRPA1 senses ultrasound. "In order to make this finding more useful for future research and clinical applications, we hope to determine exactly what parts of TRPA1 contribute to its ultrasound sensitivity and tweak them to enhance this sensitivity," says Corinne Lee-Kubli, a co-first author of the paper and former postdoctoral fellow at Salk.

They also plan to carry out another screen for ultrasound sensitive proteins -- this time looking for proteins that can inhibit, or shut off, a cell's activity in response to ultrasound.

Read more at Science Daily

So-called junk DNA plays critical role in mammalian development

 Nearly half of our DNA has been written off as junk, the discards of evolution: sidelined or broken genes, viruses that got stuck in our genome and were dismembered or silenced, none of it relevant to the human organism or human evolution.

But research over the last decade has shown that some of this genetic "dark matter" does have a function, primarily in regulating the expression of host genes -- a mere 2% of our total genome -- that code for proteins. Biologists continue to debate, however, whether these regulatory sequences of DNA play essential or detrimental roles in the body or are merely incidental, an accident that the organism can live without.

A new study led by researchers at University of California, Berkeley, and Washington University explored the function of one component of this junk DNA, transposons, which are selfish DNA sequences able to invade their host genome. The study shows that at least one family of transposons -- ancient viruses that have invaded our genome by the millions -- plays a critical role in viability in the mouse, and perhaps in all mammals. When the researchers knocked out a specific transposon in mice, half their mouse pups died before birth.

This is the first example of a piece of "junk DNA" being critical to survival in mammals.

In mice, this transposon regulates the proliferation of cells in the early fertilized embryo and the timing of implantation in the mother's uterus. The researchers looked in seven other mammalian species, including humans, and also found virus-derived regulatory elements linked to cell proliferation and timing of embryo implantation, suggesting that ancient viral DNA has been domesticated independently to play a crucial role in early embryonic development in all mammals.

According to senior author Lin He, UC Berkeley professor of molecular and cell biology, the findings highlight an oft-ignored driver of evolution: viruses that integrate into our genome and get repurposed as regulators of host genes, opening up evolutionary options not available before.

"The mouse and humans share 99% of their protein coding genes in their genomes -- we are very similar with each other," He said. "So, what constitutes the differences between mice and humans? One of the major differences is gene regulation -- mice and humans have the same genes, but they can be regulated differently. Transposons have the capacity to generate a lot of gene regulatory diversity and could help us to understand species-specific differences in the world."

Colleague and co-senior author Ting Wang, the Sanford and Karen Loewentheil Distinguished Professor of Medicine in the Department of Genetics at the Washington University School of Medicine in St. Louis, Missouri, agrees.

"The real significance of this story is it tells us how evolution works in the most unexpected manner possible," Wang said. "Transposons were long considered useless genetic material, but they make up such a big portion of the mammalian genome. A lot of interesting studies illustrate that transposons are a driving force of human genome evolution. Yet, this is the first example that I know of where deletion of a piece of junk DNA leads to a lethal phenotype, demonstrating that the function of specific transposons can be essential."

The finding could have implications for human infertility. According to first author Andrew Modzelewski, a UC Berkeley postdoctoral fellow, nearly half of all miscarriages in humans are undiagnosed or don't have a clear genetic component. Could transposons like this be involved?

"If 50% of our genome is non-coding or repetitive -- this dark matter -- it is very tempting to ask the question whether or not human reproduction and the causes of human infertility can be explained by junk DNA sequences," he said.

Embryo implantation

He, the Thomas and Stacey Siebel Distinguished Chair Professor at UC Berkeley, studies the 98% or more of our genome that does not code for proteins. For most of He's career, she has focused on microRNAs and longer pieces of non-coding RNAs, both of which are potent gene regulators. Five years ago, however, her team accidentally discovered a microRNA regulator for a transposon family called MERVL (mouse endogenous retroviral elements) that was involved in cell fate determination of early mouse embryos. The unexpected abundance of transposon transcription in mouse embryos led He's team to investigate the developmental functions of transposons, which have taken up residence in the genomes of nearly every organism on Earth.

In a paper appearing this week in the journal Cell, He and her team identify the key regulatory DNA involved: a piece of a transposon -- a viral promoter -- that has been repurposed as a promoter for a mouse gene that produces a protein involved in cell proliferation in the developing embryo and in the timing of implantation of the embryo. A promoter is a short DNA sequence that is needed upstream of a gene in order for the gene to be transcribed and expressed.

Wild mice use this transposon promoter, calledMT2B2, to initiate transcription of the gene Cdk2ap1 specifically in early embryos to produce a short protein "isoform" that increases cell proliferation in the fertilized embryo and speeds its implantation in the uterus. Using CRISPR-EZ, a simple and inexpensive technique that Modzelewski and He developed several years ago, they disabled the MT2B2 promoter and found that mice instead expressed the Cdk2ap1 gene from its default promoter as a longer form of the protein, a long isoform, that had the opposite effect: decreased cell proliferation and delayed implantation.

The result of this knockout was the death at birth of about half the pups.

Modzelewski said that the short form of the protein appears to make the many embryos of the mouse implant with a regular spacing within the uterus, preventing crowding. When the promoter is knocked out so that the long form is present only, the embryos implant seemingly randomly, some of them over the cervix, which blocks exit of the fully developed fetus and sometimes kills the mother during the birthing process.

They found thatwithin a 24-hour period prior to embryo implantation, the MT2B2 promoter ramps up expression of the Cdk2ap1 gene so much that the short form of the protein makes up 95% of the two isoforms present in embryos. The long isoform is normally produced later in gestation when the default promoter upstream of the Cdk2ap1 gene becomes active.

Working with Wanqing Shao, co-first author of the study and a postdoctoral fellow in Wang's group at Washington University, the team searched through published data on preimplantation embryos for eight mammalian species -- human, rhesus monkey, marmoset, mouse, goat, cow, pig and opossum -- to see whether transposons are turned on briefly before implantation in other species. These online data came from a technique called single cell RNA sequencing, or scRNA-seq, which records the levels of messenger RNA in single cells, an indication of which genes are turned on and transcribed. In all cases, they had to retrieve the data on non-coding DNA because it is typically removed before analysis, with the presumption that it's unimportant.

While transposons are generally specific to individual species -- humans and mice, for example, have largely different sets -- the researchers found that different species-specific transposon families were turned on briefly before implantation in all eight mammals, including the opossum, the only mammal in the group that does not employ a placenta to implant embryos in the uterus.

"What's amazing is that different species have largely different transposons that are expressed in preimplantation embryos, but the global expression profiles of these transposons are nearly identical among all the mammalian species," He said.

Colleague and co-senior author Davide Risso, a former UC Berkeley postdoctoral fellow and now associate professor of statistics at the University of Padua in Italy, developed a method for linking specific transposons to preimplantation genes so as to weed out the thousands of copies of related transposons that exist in the genome. This method is crucial to identifying individual transposon elements with important gene regulatory activity.

"It's interesting to note that the data that we used were mostly based on the previous sequencing technology, called SMART-seq, which covers the full sequence of the RNA molecules. The current popular technique, 10x genomics technology, would not have shown us the different levels of protein isoforms. They're blind to them," Risso said.

Viruses are evolutionary reservoir

The researchers found that in nearly all of the eight mammalian species, both short and long Cdk2ap1 isoforms occur, but are switched on at different times and in different proportions that correlate with whether embryos implant early, as in mice, or late, as in cows and pigs. Thus, at the protein level, both the short and long isoforms appear conserved, but their expression patterns are species-specific.

"If you have a lot of the short Cdk2ap1 isoform, like mice, you implant very early, while in species like the cow and pig, which have none to very little of the short isoform, it's up to two weeks or longer for implantation," Modzelewski said.

Wang suspects that the promoter that generates the long form of the protein could be the mouse's original promoter, but that a virus that integrated into the genome long ago was later adapted as a regulatory element to produce the shorter form and the opposite effect.

"So, what happened here is a rodent-specific virus came in, and then somehow the host decided, 'OK, I'm going to use you as my promoter to express this shorter Cdk2ap1 isoform.' We see the redundancy that's built into the system, where we can take advantage of whatever nature throws at us and make it useful," he said. "And then, this new promoter happened to be stronger than the old promoter. I think this fundamentally changed the phenotype of rodents; maybe that's what makes them grow faster -- a gift of having a shorter pre-implantation time. So, they probably gained some fitness benefit from this virus."

"Whatever you look at in biology, you're going to see transposons being used, simply because there are just so many sequences," Wang added. "They essentially provide an evolutionary reservoir for selection to act upon."

Read more at Science Daily

How the brain ignores distracting information to coordinate movements

As you read this article, touch receptors in your skin are sensing your environment. Your clothes and jewelry, the chair you're sitting on, the computer keyboard or mobile device you're using, even your fingers as they brush one another unintentionally -- each touch activates collections of nerve cells. But, unless a stimulus is particularly unexpected or required to help you orient your own movements, your brain ignores many of these inputs.

Now, Salk researchers have discovered how neurons in a small area of the mammalian brain help filter distracting or disruptive signals -- specifically from the hands -- to coordinate dexterous movements. Their results, published in the journal Science on October 14, 2021, may hold lessons in how the brain filters other sensory information as well.

"These findings have implications not only for gaining a better understanding of how our nervous system interacts with the world, but also for teaching us how to build better prosthetics and robots, and how to more effectively repair neural circuitry after disease or injury," says Eiman Azim, assistant professor in Salk's Molecular Neurobiology Laboratory and the William Scandling Developmental Chair.

Scientists have long known that input from the hands is needed to coordinate dexterous movements, from throwing a ball to playing a musical instrument. In one classic experiment, volunteers with anesthetized, numb fingertips found it extremely difficult to pick up and light a match.

"There's a common misconception that the brain sends a signal and you just perform the resulting movement," says Azim. "But in reality, the brain is constantly incorporating feedback information about the state of your limbs and fingers and adjusting its output in response."

If the brain responded to every signal from the body, it would quickly become overwhelmed -- as happens with some sensory processing disorders. Azim and his colleagues wanted to identify exactly how a healthy brain manages to pick and choose which tactile signals to take into account to coordinate dexterous movements like manipulating objects.

They used a combination of tools in mice to study cells within a small area in the brainstem called the cuneate nucleus, which is the first area signals from the hand enter the brain. While it was known that sensory information passes through the cuneate nucleus, the team discovered that a set of neurons in this region actually controls how much information from the hands eventually passes on to other parts of the brain. By manipulating those circuits to allow more or less tactile feedback through, Azim's team could influence how mice perform dexterous tasks -- such as pulling a rope or learning to distinguish textures -- to earn rewards.

"The cuneate nucleus is often referred to as a relay station, as if information was just passing through it," says Staff Researcher James Conner, first author of the new paper. "But it turns out that sensory information is actually being modulated in this structure."

Conner and Azim went on to show how different parts of the cortex in mice -- the region responsible for more complex, adaptive behavior -- can in turn control the neurons of the cuneate to dictate how strongly they're filtering sensory information from the hands.

Today, despite decades of work, most prosthetics and robots struggle to be nimble-fingered and carry out small, precise hand movements. Azim and Conner say their work could help inform the design of better processes to integrate sensory information from artificial fingers into these kinds of systems to improve their dexterity. It also could have implications for understanding sensory processing disorders or troubleshooting what goes wrong in the brain when the flow of sensory information is thrown out of balance.

"Sensory systems have evolved to have very high sensitivity in order to maximize protective responses to external threats. But our own actions can activate these sensory systems, thereby generating feedback signals that can be disruptive to our intended actions," says Conner.

"We're constantly bombarded with information from the world, and the brain needs ways to decide what comes through and what doesn't," says Azim. "It's not just tactile feedback, but visual and olfactory and auditory, temperature and pain -- the lessons we're learning about this circuitry likely apply in general ways to how the brain modulates these types of feedback as well."

Read more at Science Daily

A peek inside a flying bat's brain uncovers clues to mammalian navigation

When driving up to a busy intersection, you probably pay more attention to where you will be in the near future than where you are at that moment. After all, knowing when you will arrive at the intersection -- and whether you need to stop or slow down to avoid a collision with a passing car, pedestrian or cyclist -- is usually much more important than knowing your current location.

This ability to focus on where we will be in the near future -- rather than where we are in the present -- may be a key characteristic of the mammalian brain's built-in navigation system, suggests a new study appearing online Thursday, July 8, in the journal Science.

Neuroscientists at the University of California, Berkeley, wirelessly tracked the brain activity of Egyptian fruit bats as they flew throughout a custom flight room. When the researchers compared the bats' flight paths with their neural readings, they found that the activities of the bats' "place cells" -- special type of neurons responsible for encoding an animal's spatial position -- were often more closely correlated with where the bats would be in the near future, rather than where they were in the moment.

"We wanted to find out: Does the neural activity at the present moment do a better job at representing a past or future position than it does the actual present position? And we found that, for some neurons, the neural activity actually does a much better job of representing a future position," said lead author Nicholas Dotson, who conducted the research as a postdoctoral scholar at UC Berkeley. "The finding shows that neural activity in this region is representing more than the bat's present position -- it's tentatively representing a full flight trajectory."

Place cells, located in a region of the brain called the hippocampus, work together to form an innate "GPS system" for a variety of land animals, including humans. As an animal explores a new environment, different place cells activate at different positions, creating an internal map of the territory that can be saved and stored.

"If you had access to neural activity in my hippocampus while I walked around a room, you'd be able to decode where I was in the room based on this neural activity," Dotson said.

The discovery of place cells in rodents was awarded the 2014 Nobel Prize in Physiology or Medicine, and many of the foundational experiments were conducted in the 1970s and '80s. However, a number of questions still remain about how this region of the brain operates during rapid movement and how it works to represent "nonlocal" positions.

"Because the hippocampus is involved in navigation, there have been several studies looking at coding in this brain region and asking: How does neural activity represent things that are going to happen in the future or that have happened in the past? And can this brain region exhibit activity that doesn't represent where we are right now, but actually represents a position that is far away?" said study senior author Michael Yartsev, an assistant professor of neurobiology and bioengineering at UC Berkeley.

Earlier experiments have been unable to conclusively answer this question, Yartsev said. This is likely because they were conducted using relatively slow-moving animals, like rats, that, in experimental enclosures, will only move about an inch or two in a second -- and also because when comparing the activity of individual neurons with an animal's position over time, a shift of a fraction of an inch will not make a huge difference.

Bats, however, are extremely speedy in flight.

"Bats move really, really fast. They fly at speeds of about 30 to 50 kilometers per hour in the laboratory, which is a huge advantage, because in the same fraction of a second, a rat might move a few centimeters, while a bat would move a few meters," Yartsev said.

To conduct the experiments, Yartsev and Dotson used wireless neural recording devices to monitor bats' brain activity as they flew freely throughout a custom-built room that had been outfitted with cameras to track the bats' precise flight paths. In one set of experiments, they recorded bats' position and brain activity while humans encouraged the animals to explore the full 3D volume of the room. In another set of experiments, the bats were left alone with a set of automatic feeders, located at different locations in the room, to entice the bats to fly around.

When Yartsev and Dotson compared the timing of neural activity with the bats' flight paths, they found that when shifting the bats' positions forward in time -- by comparing the neural activity with the locations where the bats would be in a few hundred milliseconds, or in a second -- suddenly, the neural activity correlated much more strongly with spatial position.

"Based on the data, you might assume that some neurons don't encode spatial information at all, because there is no correlation with the position at time zero or the present moment," Yartsev said. "But if you compare their activity to a position a second in the future, suddenly the correlation is incredibly sharp."

The findings suggest that place cells' activity doesn't just represent a single current position, but actually a trajectory that stretches into the near future, and into the past, as well.

"We can imagine walking down a hallway and picturing where we just were and where we will be shortly. What does that activity look like in the brain?" Dotson said. "Our findings suggest that as the bats are flying, they're representing in their mind not just where they are, but where they are along the path."

Though place cells and the basic components of this navigational system have been identified in a wide variety of mammals, it's not yet clear whether this ability to project a path up to a second into the future is unique to bats and their rapid flight pattern, or is shared by a wider variety of animals. However, the discovery opens up a variety of interesting questions about how we humans process our movement through time and space, Yartsev said.

Because the hippocampus is also a locus of many diseases, such as Alzheimer's, where a person's sense of location and memory is often disrupted, uncovering these basic neural computations could also give scientists a better understanding of disease-related impairment and help them devise more effective treatments.

Read more at Science Daily