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When Numbers Play Tricks: Unraveling The Brain’s Biases

Imagine living in a dark cave, with your entire understanding of the world based on shadows on the wall. Sounds unrealistic and terrifying, right? However, this allegory presented by Plato is an apt metaphor for our brain’s perception of the real world. While we might believe we perceive reality in its entirety, our brain can only provide a shadowy representation of the variables in our environment, and our decisions are based on these shadowy representations.

A comforting thought might be that numbers are a universal language for Western countries and, specifically, for those who use Arabic numerals. For instance, when the price of a good is marked as $14, it conveys an unambiguous and specific value, meaning that one would unequivocally expect to pay exactly that amount. However, experiments show that people make mistakes when dealing with Arabic numerals. For instance, under time constraints, the closer two numbers are in value, the more challenging it becomes for us to rapidly and accurately pinpoint the larger one. These mistakes are very similar to those made in psychophysics tasks involving physical stimuli, such as comparing the length of segments or averaging the orientation of tilted lines. These results, together with neurobiological studies, suggest the existence of a representation system for numbers that is similar to how we interpret physical stimuli.

A popular idea in theoretical neuroscience is that while the brain’s computational abilities have inherent limits, leading to imprecise representations, these representations are optimal within those constraints. This theory, called “efficient coding”, suggests that our brain’s perceptions are influenced by how often these magnitudes are encountered (i.e., the prior). For example, vertical and horizontal orientations are perceived with more clarity than oblique ones, likely because they’re more common in our environment.

A recent study (https://www.nature.com/articles/s41562-022-01352-4), led by Columbia postdoc Arthur Prat-Carrabin and published in Nature Human Behaviour, delves into whether our brain treats numbers the same way it treats physical stimuli. In their experiment, participants were asked to determine which series of numbers, red or green, had a higher average value. For instance, as shown in Figure 1, the number 79.60 would flash in red on the computer screen, followed by 44.92 in green, and so forth. Participants were tasked with rapidly and intuitively calculating the average of the red and green numbers to determine which sequence had the greater average. To investigate how the frequency at which numbers were encountered impacts their representation, numbers were drawn from different distributions: one, in which smaller numbers were more frequent (Downward prior); another, in which all numbers had equal chances (Uniform prior); and a third, in which larger numbers were more frequent (Upward prior).

Figure 1: Example trial of the task. Participants were presented with 5 red and 5 green numbers and had to choose the color with the higher average.

To analyze the participants’ decisions, the researchers compared their answers with multiple computational models characterized by two components: first, whether or not the numbers are encoded with a bias that depends on the value of the number, and second, whether or not the imprecision (the noise) with which numbers are represented varies with the number. Their results showed that the model that best aligned with the participants’ answers had to include both components. Notably, less common numbers are fuzzier in their perception. So, when using the Downward prior, bigger numbers are encoded with more noise, while with the Upward prior, the smaller numbers are the noisiest ones.

This discovery not only supports the “efficient coding” theory, which posits that the brain encodes and represents information in the most efficient way possible, but also showcases its applicability beyond just physical perceptions. Whether we’re assessing the speed of a car, the talent of a dancer, or the sweetness of a cake, our brain might use a universal mechanism to represent these variables. This mechanism dynamically adjusts to the statistical distribution of numbers that are expected or experienced, which implies that our understanding of numbers and magnitudes isn’t static but can be influenced by our prior experiences and expectations. In the near future, we might be able to design environments that help people refine their perceptions (such as by crafting digital games to enhance the consumer’s responsiveness to certain prices), allowing them to better discern specific value ranges and improve their decision-making.

Reviewed by: Emily Hokett, Trang Nguyen, Martina Proietti Onori

 

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Revealing How Cancer Cells Balance Immune Signals

The immune system constitutes a complex network of biological processes that serves to safeguard organisms from various diseases. An important role of the immune system is its ability to distinguish between normal cells in the body and foreign cells such as germs and cancer cells. This discrimination empowers the immune system to target foreign agents while preserving normal cellular integrity. If the immune system malfunctions, it mistakenly attacks healthy cells, tissues, and organs which is called autoimmune disease. These attacks can affect any part of the body, weakening bodily function and even turning life-threatening. Therefore, it is very critical to maintain a balance between combating tumor cells and avoiding autoimmune disease in the body. The immune system maintains such balance by using checkpoint proteins on immune cells. The checkpoints act like switches that need to be turned on (or off) to ensure an optimal immune response. However, cancer cells sometimes find ways to use these checkpoints to avoid being attacked by the immune system.

Medicines known as monoclonal antibodies can be designed to target these checkpoint proteins. These drugs are called immune checkpoint inhibitors. Checkpoint inhibitors don’t kill cancer cells directly, but they help the immune system to better find and attack the cancer cells in the body. Immune checkpoint inhibitors such as anti-PD-1 antibodies, are now used to treat some types of cancers such as metastatic melanoma or brain tumors due to their ability to boost the immune response against cancer cells. It normally acts as a type of “off switch” that helps keep the T cells from attacking other cells in the body. It does this when it attaches to PD-L1, a protein on some normal (and cancer) cells. When PD-1 binds to PD-L1, it basically tells the T cell to leave the other cell alone. Some cancer cells have large amounts of PD-L1, which helps them hide from an immune attack (Figure 1).

 

Figure 1: The interaction between tumor cells and T cells. Tumor cells contain PD-L1 receptors that bind to PD-1 in T cells to hide an immune attack from T cells (left panel). However, through the application of an immune checkpoint inhibitor (anti-PD-L1 or anti-PD-1), this binding is hindered, empowering T cells to eliminate tumor cells (right panel). Image is created via Biorender.

The Izar group at Columbia University Medical Center, led by Patricia Ho identified a mechanism by which CD58 loss or downregulation contributes to cancer immune evasion and immune checkpoint inhibitor (ICI) therapy. CD58 is known as an adhesion molecule facilitating the initial binding of effector T cells. By sequencing the whole genome of patient samples, specifically those who underwent anti-PD-1 antibody treatment and later developed drug resistance, they illuminated the role of CD58. Notably, CD58 facilitates early T-cell infiltration into tumors within a tumor differentiation. Moreover, the study shed light on CD58 loss potentially expediting tumor progression by upregulation of PD-L1, consequently intensifying resistance to ICI therapies.

To better understand the CD58/PD-L1 co-regulation, the research group performed a genome-wide knock out screen to identify the regulator of CD58. Among the top hits of cell population with low CD58 protein, authors identified CMTM6 as an important regulator for the T cell activation and antitumor responses. CMTM6 binds to CD58 and promotes its recycling from cell surface to endosomes, thus preventing its lysosomal degradation (Figure 2). Interestingly, CMTM6 is also known as an important regulator of PD-L1 maintenance. Thus, loss of CD58 results in increased PD-L1 levels by stabilizing CMTM6-PD-L1 interactions and therefore reducing lysosomal PD-L1 degradation (Figure 2).


Figure 2: Both CD58 and PD-L1 require and directly bind to CMTM6. CD58 forms a direct binding with CMTM6, enhancing recycling and curtailing lysosomal degradation (left). CD58 knockout fosters PD-L1 up-regulation by interacting with CMTM6 (right). Figure is adopted from the original paper.

This work provides a molecular basis for clinically relevant roles of CD58 to facilitate initial infiltration into the tumor. Loss of CD58 confers resistance to immune checkpoint inhibitors which may provide important insights and inform rationale for the next generation of combinations of cancer immunotherapies.

Read more about this novel finding here: The CD58-CD2 axis is co-regulated with PD-L1 via CMTM6 and shapes anti-tumor immunity.

Reviewed by: Martina Proietti Onori and Carlos Diaz Salazar Albelda

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Masters of disguise: unlocking the secrets of color-changing cuttlefish

Cuttlefish, octopus, and squid are fascinating marine creatures that have perfected the art of camouflage underwater. With uncanny speed, these soft-bodied cephalopods can change the color, pattern, and texture of their skin to blend seamlessly into their surroundings, impressively adapting to their environment. Unlike many animals, cuttlefish don’t rely on fur or feathers to hide in the background. Instead, they actively manipulate thousands of pigment cells in their skin to acquire the color of the environment around them. This intricate disguise process starts in their brains, as camouflage is a response to the animal’s perception of the external world. To conceal their bodies, cephalopods convert visual inputs into neural representations within their brain, ultimately transmitting signals all the way to the skin, where thousands of tiny structures called chromatophores adjust to allow color changes. However, camouflage is just one captivating aspect of cuttlefish biology. These marine animals present a rich repertoire of signaling behaviors for mating and communication and they are proficient learners, with memory capabilities not often seen in invertebrates. Collectively, these attributes position cephalopods as extraordinarily captivating subjects within the realm of biological studies.

Researchers at Columbia University’s Zuckerman Institute, led by Dr. Richard Axel, have made significant progress in comprehending the shape-shifting abilities of cuttlefish. In a recent article published in Current Biology, they generated a detailed neuroanatomical brain map, revealing insights into how their skin transformation is controlled. Tessa Montague, PhD and colleagues focused on the dwarf cuttlefish (Sepia bandensis), a small tropical species found around coral reefs in the Indo-Pacific Ocean (Figure 1A). The dwarf cuttlefish is an intriguing model for neuroscience research with its rich array of dynamic social behaviors and camouflage. Through an advanced imaging technique called MRI (magnetic resonance imaging), computer programming and web design they constructed a 3D atlas illustrating the dwarf cuttlefish’s brain anatomy (Figure 1B).

Figure 1. (A) Adult dwarf cuttlefish. (B) 3D template brain, based on magnetic resonance imaging (MRI) of 8 cuttlefish brains, (i) anterior view, (ii) right view, and (iii) posterior view. The abbreviations in color indicate the different brain structures that were identified and mapped.  Modified from T. G. Montague et al., 2023

By scanning the bodies and brains of male and female cuttlefish, the researchers identified 32 distinct lobes or functional units within the cuttlefish brain (Figure 1B). Each lobe is densely packed with neurons and performs specialized tasks. The two largest lobes, making up 75% of the total brain volume, are the optic lobes (O in Figure 1B). They receive direct projections from the eyes and process visual information, a crucial step in enabling cuttlefish camouflage. Notably, other key lobes in the camouflage pathway include those controlling the chromatophores, the pigment-filled saccules in cuttlefish skin that provide the color. When the lobes send signals to the chromatophores, these rapidly expand or contract to alter skin shades on a millisecond timescale. The lateral basal lobe (LB in Figure 1B) for example, is the lobe involved in establishing the most appropriate skin pattern components for camouflage. Another brain area highlighted by the atlas is the vertical lobe complex (V and surrounding lobes in Figure 1B), which previous studies suggest plays a key role in learning and memory. Unlocking the functions of this lobe could reveal the neural basis for complex behaviors like camouflage.

While past studies have mapped the brains of related cephalopods like squid and octopus, this is the first complete atlas for a cuttlefish species and it provides the neuroscience field with a valuable comparative perspective.  The researchers found strong similarities in the anatomy of the dwarf cuttlefish with the common cuttlefish, despite differences in size and camouflage strategies between the species. This suggests that fundamental aspects of brain organization are conserved, at least among close cephalopod relatives. It also highlights how flexible cuttlefish brains are: they can generate very different camouflage patterns using essentially the same basic circuit layout. Exploring what accounts for such flexibility on the microscopic scale will be the next challenge.

To maximize the utility of the brain atlas, for both educational and research purposes, the authors built an interactive freely available web tool, called Cuttlebase, where users can identify specific brain regions in the histological 2D atlas (that shows the cellular components in the cuttlefish brain)  and use the 3D model of the brain to explore the different lobes (Figure 2).

Figure 2. Cuttlebase is an interactive scientific web toolkit for the dwarf cuttlefish that includes a histological brain atlas in 2D and a 3D body and brain atlas to visualize and explore the different organs and brain lobes. Obtained and modified from https://www.cuttlebase.org/

This atlas serves as an invaluable tool for the scientific community to explore the basic anatomical components of complex behaviors and can give us insight into how brains are capable of representing information. It also offers an invaluable anatomy lesson, giving scientists a privileged peek inside the ingenious brains of these undersea masters of disguise. With a detailed understanding of their sophisticated neural systems, unraveling cuttlefish mysteries seems closer than ever.

Reviewed by: Trang Nguyen, Maaike Schilperoort

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Copy & Paste – an essential mechanism to repair damaged DNA

Maintaining genome integrity is crucial for cell survival and genome instability is one of the hallmarks of cancer. The genomes of many living organisms including humans are composed of different numbers of DNA molecules that are folded into structures called chromosomes. Each chromosome represents one double stranded DNA molecule. Most of our cells harbor two sets of each chromosome, which are denoted as homologous chromosomes (Figure 1, left panel). The homologous chromosome pairs can contain identical or different versions (alleles) of the genes that they carry since they are inherited from each parent. In this sense homologous chromosomes are genetically non-identical.

Most of the cells in our body are constantly renewed and replaced in a process that involves cellular division. During cell division the genome needs to be duplicated so that each of the newly formed daughter cells receives a genome copy. The process of DNA duplication is called DNA replication. During replication, each of the two strands of the DNA double helix serves as a template for the synthesis of a new complementary DNA strand. These chromosome copies that are produced during cell division are named sister chromatids (Figure 1, right panel), and they are found transiently in the cells just before the division. Unlike the homologous chromosomes, the alleles of the genes in the sister chromatids are genetically identical since they are produced by DNA replication.

Figure 1: Schematic representation of the homologous chromosomes and the sister chromatids produced during cellular division. Created with BioRender.com

During DNA replications, as well as under the influence of diverse environmental and endogenous agents, lesions constantly occur on DNA. These DNA “injuries” can trigger mutations, compromise the genome integrity, or even cause cell death. DNA double-strand breaks (DSBs) are one of the most deleterious types of DNA lesions that can lead to gross chromosome rearrangements. Similar rearrangements are very common in cancer.  One of the major mechanisms for DNA DSBs repair is homologous recombination (HR), a process explained below. Mutations in different genes from the HR pathway have been associated with diseases like cancer of Fanconi anemia, as well as to hypersensitivity to DNA-damaging agents which increase mutation accumulation.

One of the versions of HR involves the use of the intact sister chromatid as donor of information. Since the sister chromatid has an identical sequence to the damaged DNA molecule, this repair system faithfully restores the genetic information and is considered as being error free. Homologous recombination can also occur between the homologous chromosome pairs in the case of non-dividing cells. In HR, the DNA DSB ends are processed and a long single-stranded DNA (ssDNA) overhang is left to serve as platform for the assembly of the protein machinery mediating the repair (Figure 1, a). This ssDNA is coated by the protein RPA to prevent degradation and folding. Subsequently RPA is replaced by RAD51, into a structure termed the presynaptic nucleoprotein filament. This filament is capable of searching for a homologous undamaged DNA molecule that will serve as a template for repair. This exchange between RPA and RAD51 on ssDNA in the presynaptic filament assembly is facilitated by mediators of HR. One of those mediators is a complex of four proteins that are paralogues of RAD51: the BCDX2 complex. However, the exact mechanism of action of the BCDX2 complex in HR remained elusive.

Figure 2. Double-strand breaks (DSBs) can be repaired by several homologous recombination (HR)-mediated pathways. Represented is a simplified version of the “synthesis-dependent strand annealing HR pathway”. In all pathways, the repair is initiated by resection of a DSB to provide 3′ single-stranded DNA (ssDNA) overhangs (a and b). The ssRNA is rapidly coated with RPA to prevent damage and folding (c). RPA is exchanged with RAD51 with the help of HR mediator complexes, which are composed from different RAD51 paralogues, one of which is four protein complex BCDX2 (d). RAD51 coated DNA can search for the non-damaged homologous chromosome / sister chromatid pair and invade that intact DNA duplex (e). After strand invasion and “copying” of the information from the intact DNA on one of the damaged strands from the blue DNA duplex, the reaction can proceed to hybridization to the ssDNA on the other break end of the blue DNA duplex, followed by DNA synthesis (f) and restoring  of blue DNA molecule (g). Created with BioRender.com

Recent studies from Columbia postdoc Aviv Meir and colleagues revealed the structure of the human BCDX2 complex. Cryogenic electron microscopy, a technique that allows the high-resolution structure determination of biomolecules in solution, was used to resolve both the  free and single-strand DNA-bound states of BCDX2. This provided the first structural information of one of the RAD51 paralogues complexes. This structural information provides insight into how the complex assembles and disassembles, which in turn is linked to the regulation of its function. The scientists also discovered by single molecule analysis that the association of BCDX2 with RPA–ssDNA enhances the rate of assembly of the RAD51–ssDNA filament. In humans, BCDX2 binds the RPA–ssDNA prior to the arrival of RAD51 and then promotes the RAD51 filament assembly. This novel mode of action for the proteins of the BCDX2 complex is different from what was previously observed in other organisms, where BCDX2 only transiently associates with the ssDNA. The work by Dr. Meir and colleagues, recently published in the prestigious journal Nature, not only elucidates how BCDX2 mediates RPA–RAD51 exchange on ssDNA but also provides a foundation for deciphering how alterations in BCDX2 subunits that were found in patients with cancer can impact genome repair and can lead to the pathogenesis. This valuable information opens the doors for future targeting of those “defectuous” BCDX2 parts for therapeutic developments.

Reviewed by: Maaike Schilperoort, Apurva Limaye, Giulia Mezzadri , Trang Nguyen

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What will capture your attention?

Ever wonder how your brain directs your attention to certain stimuli or events? For example, if you are walking into a crowded restaurant you will likely direct your attention to finding an empty seat or finding your friends rather than the color of the tablecloths or the number of tables and chairs in the restaurant. Our brains have to navigate and distinguish what is important in complex environments. The term “salience” is defined as a noticeable or important object that stands out from the surroundings or background. Salience processing has been studied in neuroscience and psychology in order to understand how our brains distinguish important stimuli. This phenomenon involves two general mechanisms: bottom-up processing in which sensory information can be amplified or filtered, or top-down processing which focuses on goal-directed behaviors and cognitive control. Prior research has identified areas of the brain that are involved in salience processing using functional magnetic resonance imaging (fMRI) studies. FMRI is a technique that measures brain activity by measuring minute changes in blood flow. The regions identified include the regions in the cortex: dorsal attention network (DAN), salience network (SN), sensory cortex, primary somatosensory cortex (S1), and subcortex (Figure 1). However, understanding how these regions communicate with one another in salience processing is still unknown. In addition to these cortical networks, the locus coeruleus (LC), which is the primary source of the neurotransmitter norepinephrine (NE) and located in the brain stem (Figure 1), is also associated with salience processing. 

Figure 1. Cortical and non-cortical regions of the brain involved in salience processing.

Researchers commonly use MRI or FMRI (functional magnetic resonance imaging) as a technique to detect active areas of the brain while performing an activity. This is complemented with EEG (electroencephalogram) scans to measure electrical activity in the brain while performing different tasks.  One of the widely used experimental designs to evaluate salience processing includes the oddball task. For these experiments, subjects are instructed to detect infrequent target stimuli (a sound of a laser gun)  in a stream of standard stimuli (standard tones). FMRI and EEG measurements were obtained to determine activation of the different areas of the brain while hearing the sound of the target laser gun as compared to hearing the consistent standard tone. Columbia postdoc Linbi Hong and colleagues in their study, published in PLOS Computational Biology on May 2023, wanted to further understand the interaction between the non-cortical, LC-NE system and the cortical networks using simultaneous recordings of pupillometry which measures changes in pupil diameter and has been used as a marker for LC activity, electroencephalography (EEG) and fMRI in an oddball experiment. 

The study first utilized EEG-informed fMRI analysis to map the neural cascade underlying salience processing and identify the order of specific regions which are activated during the oddball task. The researchers defined areas in the brain to understand the organization of regions involved in salience processing. Next, the researchers wanted to understand the functional connectivity between the distinct cortex regions and the LC-complex (non-cortex regions) during salience processing by analyzing EEG signals at different times during the oddball experiment. It was determined that the prefrontal cortex and dorsal attention network are the main players in processing salient stimuli. 

Next, the researchers characterized the directional interactions between the functional network regions including the cortical and non-cortical regions. Specifically, they determined that non-cortical regions showed significant functional connectivity with dorsal auditory attention networks and salience networks. These results indicate that the non-cortical regions (LC network) are involved in switching between the different cortical networks during salience processing.

Overall, this study identified spatiotemporally the connectivity of different cortical regions to the non-cortical regions (LC complex) in salience processing. The study also provides insights into utilizing noninvasive pupillary response, which measures non-cortical region (LC) activity, in combination with EEG and fMRI signals, which measure cortical activity, as a valid approach to understand connectivity within the different regions in the brain. As a result, these methods could be used to determine any decline in connectivity of the cortical and non-cortical regions which can provide further understanding in neurological diseases such as Alzhiemers in which the LC network is the first region affected. Future studies from this group can provide additional input to how our brains can distinguish important information such as finding your friends in a crowded restaurant and whether this information is captured the same way in diseased conditions. So, next time you look around in a crowded restaurant, you can think, what captures your attention?

Reviewed by: Trang Nguyen, Martina Proietti Onori, Maaike Schilperoort

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How many candles on the cake this year?

Every year as we celebrate our birthdays, we mark the addition of a year to our lives. Our birthdays determine our chronological age measured in days, months and years since the day we were born. Biological aging on the other hand is another measure of aging that accounts for the gradual accumulation of cellular and tissue damage that occurs in the body as we grow older. Aging is a natural process and various factors contribute to biological aging, including our chronological age, genetics, lifestyle, nutrition, and physical activity. Research has shown poor nutrition and low physical activity can accelerate biological aging. Accelerated biological aging is marked by increased levels of certain hallmarks of cellular damage, leading to chronic diseases. Poor nutritional habits and sedentary lifestyles have been associated with increased risk of heart diseases, high blood pressure, cholesterol and type 2 diabetes. Additionally, over 60% of the aging population (>65 years) is expected to be affected by more than one chronic disease by 2030. Research has also shown that lifestyle interventions may reduce or delay the progress of biological aging. In this regard, Aline Thomas and colleagues obtained real life data from a large cohort of US adults to study the association between lifestyle behaviors and biological aging using mathematical models. They assessed signs of aging in individuals who engaged in some form of moderate to vigorous physical activity in their leisure time and followed a diet that resembled a mediterranean diet compared to individuals who followed a less-healthy lifestyle. A Mediterranean diet focuses on plant-based foods and healthy fats. It includes vegetables, fruits, whole grains, fish and extra virgin olive oil as a source of healthy fats. The researchers studied diet, exercise, and variations in healthy lifestyle behaviors across different age groups, genders, and body mass indices (BMI).

Dr. Thomas and colleagues combined data collected over a period of 20 years from 1999-2018 for their study. The study included 42,625 participants between the ages of 20-85 and assessed the adherence to the Mediterranean diet and an exercise regimen using a point based system. Inclusion of fruits and vegetables, legumes, cereals, fish and a ratio of mono-unsaturated to saturated fats were each awarded one point. A healthy Mediterranean diet also includes a mild-moderate amount of alcohol, which is 0-1 glass for women and 0-2 glasses for men. So, a point was given if a mild-moderate amount of alcohol was consumed. Dairy products and meat are not part of the Mediterranean diet. If participants had consumed these foods but had consumed it less than a specific amount, they were still awarded a point. The points were totaled and found to be between 0 and 9. Higher scores meant a higher adherence to the Mediterranean diet. Leisure time physical activity (LTPA) describes any physical activity performed during participants free disposable time. The researchers assessed LTPA based on the frequency, duration and intensity to calculate points / scores for each activity. They categorized the activity levels based on the scores per week into four groups ranging from – sedentary (0 points), low (<500 points), moderate (500-1000 points) and high (>1000 points). Biological age was calculated using an algorithm called PhenoAge. The algorithm calculates biological age based on chronological age and 8 biomarkers obtained from blood samples.

The study included individuals across different races, socio-economic backgrounds, marital statuses, income to poverty ratios, and with various lifestyle-related factors (e.g., smoking, BMI category, total energy intake), making it a representative population of US adults. The researchers found very interesting observations relating to diet and exercise. They discovered that adherence to a relatively healthy diet and engagement in  physical activity were independently associated with a lower biological age. Participants with a healthy diet and some level of activity were on average 1 biological year younger than the participants with the least healthy diet and sedentary lifestyle. Another very interesting finding was that individuals who had a less healthy diet but who were active even at a low level showed delayed biological aging. However, delayed biological aging was not found in participants with a healthy diet and a sedentary lifestyle, suggesting that moderate physical activity is a key component of healthy biological aging.

The findings of this study reiterates the need for better lifestyle choices across all strata of the population as the results were consistent regardless of age, sex and BMI category. A nutritious diet and moderately active lifestyle can have a positive impact on health, aging and quality of life. Getting older is inevitable, but you may be one year younger with a healthy diet and an exercise routine.

Reviewed by: Trang Nguyen, Giulia Mezzadri, Erin Cullen, Maaike Schilperoort 

 

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Unveiling the secrets of pain: decoding the structure of a human receptor for effective relief

Pain is an essential sensation for the survival of organisms. It acts as a protective mechanism, signaling potential harm and prompting animals to recognize noxious stimuli for avoiding future harm. For example, individuals unable to feel pain sensations (a rare congenital disease) seem advantaged at first glance; however they are actually at a high risk of unknowingly injuring themselves. While acute pain serves a crucial role in preserving well-being, chronic or persistent pain can severely impact the quality of life. Unfortunately, the mechanisms underlying the transition from acute to chronic pain remains poorly understood, making diagnosis and treatment challenging. Although opioids are commonly used to manage chronic pain, they carry the risk of addiction and interference with normal brain activity. As an alternative, targeting directly the receptors involved in pain pathways, such as the transient receptor potential (TRP) family, offers a promising avenue for developing analgesic therapies.

A recent study (https://www.nature.com/articles/s41467-023-38162-9) led by Dr. Arthur Neuberger, a postdoctoral research scientist in the department of Biochemistry and Molecular Biophysics within Dr. Alexander Sobolevsky’s lab, investigated the structure and inhibition mechanism of human TRPV1 and its interaction with a potential analgesic compound known as SB-366791.

TRPV1, a receptor found on nerve cells, plays a pivotal role in sensing pain and heat. It is a temperature-sensitive TRP ion channel, commonly referred to as the vanilloid receptor 1 or capsaicin receptor (pungent compound from chili pepper). TRPV1 functions as an ion channel involved in pain sensation, becoming activated in response to noxious signals. When activated, TRPV1 opens, leading to a decrease in the electrical resistance of the cell membrane and the subsequent flow of ions. If the strength of the noxious signal and the response of the ion channel are sufficient, a change in the membrane potential occurs, allowing the signal to be transmitted from the peripheral nervous system to the spinal cord and ultimately to the brain (Figure 1).

Figure 1. Overview of the pain-processing pathway. When exposed to noxious stimuli, such as the burning sensation of a flame on the skin or the heat sensation of capsaicin on the tongue, TRPV1 ion channels are activated in the peripheral nervous system. The resulting heat signal is transmitted through nerve fibers and follows spinal pathways until it reaches the brain, where it triggers the generation of a pain response. Figure created using Biorender.

The study, published in Nature Communications, significantly advanced our understanding of the three-dimensional structure of TRPV1, providing crucial insights into its functional properties. The researchers employed advanced techniques, such as cryo-electron microscopy (cryo-EM), to determine the precise arrangement of human TRPV1. Additionally, whole-cell patch clamp electrophysiology (technique that measures the electrical signals as flow of ions across the cell membrane providing information on how the cell responds to certain stimuli or drugs) was used to propose the inhibition of TRPV1 by SB-366791. These cutting-edge techniques provided valuable insights into the architecture and functional characteristics of TRPV1 (Figure 2). Through their investigations, the researchers successfully identified the specific binding site of SB-366791 on the vanilloid site of TRPV1, utilizing cryo-EM and mutagenesis techniques. This discovery suggests that the analgesic compound may exert its effects by selectively inhibiting TRPV1 activity. Furthermore, the study revealed the structural changes that occur within TRPV1 upon binding to SB-366791. These structural alterations affect the conformation of the protein, demonstrating that even in the closed state of TRPV1, SB-366791 can bind and stabilize the closed conformation. This mechanism has the potential to reduce pain signaling. Understanding these structural modifications is important for the development of novel analgesic drugs targeting TRPV1.

Traditionally, pharmacological and physiological investigations of TRPV1 have primarily focused on rodent and squirrel orthologues. However, since the TRPV1 channel plays a central role in almost every aspect of human physiology and disease, including pain and temperature perception, the recent study by Neuberger and colleagues is significant. By specifically examining the human TRPV1 structure and its interaction with SB-366791, this research opens up new possibilities for the development of effective analgesic therapies and sets the stage for future investigations in the field of pain management.

Figure 2. Overview of cryo-electron microscopy (cryo-EM) and the structure of human TRPV1 in complex with the inhibitor SB-366791. In cryo-EM, a beam of electrons is directed at a frozen solution containing the protein of interest. The resulting electrons pass through a lens system, creating a magnified image on a detector. From this image, a three-dimensional model of the protein’s structure is generated. The structure of hTRPV1 in complex with SB-366791 is visualized from two perspectives: parallel to the membrane and extracellularly. In the images, the TRPV1 subunits are color-coded as green, yellow, pink, and blue, while the lipids are represented as purple sticks. (Figure modified from Neuberger et al., 2023)

Reviewed by: Maaike Schilperoort, Giulia Mezzadri and Trang Nguyen

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A new advance in understanding the development of binocular vision

Have you ever wondered how two eyes can work together to create a single image? This occurs through a complicated mechanism called binocular vision, or stereopsis. When light hits the eye, photosensitive cells called rods and cones translate that light into an electrical signal, which they communicate to retinal ganglion cells (RGCs). RGCs have long axons that transmit visual signals to neurons in the brain. These axons are initially carried from the eye to the brain in a bundle called the optic nerve. Each eye has an optic nerve, and the two optic nerves meet at an intersection called the optic chiasm. At the optic chiasm, some of the RGC axons from each optic nerve cross to the opposite, or contralateral, side of the brain and the rest stay on the same, or ipsilateral, side. This process allows the part of the world that the left eye can see and the part that the right eye can see to overlap in the center, forming one cohesive image. 

Figure 1. The role of the CMZ in ipsilateral RGC generation. On the left, a diagram shows axons either remaining on the ipsilateral side or crossing to the contralateral side of the brain at the optic chiasm, a process which makes binocular vision possible. Ipsilaterally projecting RGC axons are shown in green, and contralaterally projecting RGC axons are shown in black. On the right, CMZ progenitors are shown differentiating into ipsilaterally projecting RGCs (modified from Slavi et al., 2023).

When binocular vision does not develop normally, depth perception can be impaired. This is often the case in albinism, a genetic condition characterized by low levels of melanin that occurs in humans and other mammals. While melanin is most widely known for its contributions to skin color in humans, it also plays an important role in the development of the visual system. The exact nature of this role is still being uncovered. Recent work emerging from a collaboration between the Mason-Dodd lab and the John lab at Columbia University’s Zuckerman Institute, led by Postdoctoral Research Scientist Dr. Nefeli Slavi and Associate Research Scientist Dr. Revathi Balasubramanian, used albino mice to identify why deficits in depth perception might occur in albinism. The study, published in the leading neuroscience journal Neuron, also advanced our understanding of the development of binocular vision generally. 

Most RGCs are formed in a location called the neural retina during development, but recent research has uncovered a prominent role for another source of RGCs called the ciliary margin zone (CMZ). The CMZ produces mostly ipsilaterally projecting RGCs, which are present at abnormally low levels in albinism. Based on this existing research, Drs. Slavi and Balasubramanian and their co-authors decided to investigate why these cells are not produced at typical levels in albino mice. First, the group found that there were fewer cells in the actively dividing phases of the cell cycle in the CMZ in albino mice. Further investigation revealed that this deficiency in cell division led to a lower number of ipsilaterally projecting RGCs in these mice. They traced these deficits to decreased expression of a protein called CyclinD2, which is known to encourage cell division. To further investigate the role of CyclinD2 in cell division in the CMZ, the researchers used mice that were pigmented (not albino) but did not express CyclinD2 in the CMZ. These CyclinD2-deficient mice had the same impairments in cell division as the albino mice.

Next, the researchers tested the impact of this impaired cell division on depth perception. They assessed the depth perception of albino and CyclinD2-deficient mice compared to control pigmented mice using an established setup called the visual cliff task. In the task, mice are placed on a narrow platform on a smooth glass surface with a shallow drop below the glass on one side and a far drop on the other side. They assessed each mouse’s depth perception based on how often they chose the shallow, safer side. This task showed that albino and CyclinD2-deficient mice were unable to recognize the difference between the shallow and deep sides, demonstrating impaired depth perception. 

Figure 2. Diagram of the platform used for the visual cliff task to assess binocular vision in mice. A mouse with impaired depth perception won’t recognize the difference between the side of the platform with surface directly under the glass and the side that looks like a long drop off a cliff (Slavi et al., 2023).

Once the researchers discovered that the deficit in the number of ipsilaterally projecting neurons was important for visual function, they turned to attempting to correct it. They found a clue in a set of nearly 25 year old studies, which indicated that CyclinD2 might be able to be increased by activating calcium channels in the neuronal membrane. To test the effect of calcium channel activation on neurogenesis and depth perception,  the researchers injected pregnant albino mice with a calcium channel-activating drug called BayK-8644. The drug increased CyclinD2 activity during a critical period for RGC neurogenesis in the mid-gestational mouse pups. In those pups, ipsilateral neurogenesis was restored significantly, though not normalized to the level of the control pigmented mice. When these mice were adults, the visual cliff task was used to assess their depth perception. Remarkably, the increase in CyclinD2 activity and subsequent increase in RGC neurogenesis during gestation dramatically improved binocular vision in the albino mice. However, the drug did not rescue these deficits in CyclinD2-deficient mice, suggesting that the presence of CyclinD2 is necessary for the development of binocular vision. 

This paper describes a novel role for the CMZ in the development of the retina and binocular vision, expanding our collective understanding of how the complex mammalian visual system develops. It also identified a role for CyclinD2 in this process, further showing that the presence of CyclinD2 is necessary for the development of binocular vision. These discoveries uncovered new directions for further understanding the development of the visual system and treating vision-related disorders.

 

Reviewed by: Trang Nguyen and Flavia Dei Zotti

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Using exposomics to manage sex- and gender-specific healthcare

It is no secret that gender bias is prevalent in biomedical and population health research. Not only is basic science research biased against females, but so are clinical trials. The exclusion of non-white, non-male, and non-cis-gender research subjects in clinical trials is justified by the need to “minimize” extraneous variables. This has led to the systematic exclusion of women, non-cis-gender, and Black, Indigenous, and People of Color (BIPOC) individuals in research and created gaps in managing the health of individuals of different races and ethnicities across both the gender and sex spectra.

A person’s state of health is a product of an interaction between their genetic composition and their collective lifelong environmental exposures. Since men and women are largely genetically similar except for differences due to their sex chromosomes, environmental exposures are the main determinants of sex-specific health outcomes. Environmental exposure is determined by cultural norms, which are in turn dictated by heterosexism, classism, misogyny, patriarchy, and racism. Therefore, biological sex and gender identity may influence the types and patterns of environmental exposures that an individual experiences.

A recent review by Dr. Meghan L. Bucher and colleagues, from the Department of Environmental Health Sciences, presents the exposome as a tool to analyze both environmental exposures and the associated biological effects of those exposures to understand how the intersection of environmental health and biological sex and gender identity impact health.

Exposome, a word introduced 15 years ago, is a new field that aims to provide an environmental complement to the genome. While there have been substantial advances in genomics over the past decade, the environment remains mysterious from a scientific standpoint because it does not lend itself to a systematic evaluation of its constituent components. Historically, we have not been able to comprehensively analyze the environment in a way such that it fits into the biomedical framework. That is what the exposomics sets out to deliver. Further, it also aims to provide an analysis of how our biology responds to those environmental exposures.  In conventional analyses, only a few exposures or markers are targeted, whereas, exposomics characterizes all exposures in an untargeted and comprehensive manner. For example, exposomics use high-resolution mass spectrometry (HRMS) technologies that facilitate high-throughput detection of compounds or chemical patterns from complex and dynamic exposures.

Exposure patterns may vary significantly depending on biological sex and gender identity. Further, exposures may exert sex-specific effects by interacting with biological factors such as sex hormones. The ability to define and characterize one’s exposome based on gender identity and biological sex would provide critical insight into factors influencing an individual’s health.

An example of how biological sex determines exposure patterns can be seen in how specific occupations and household responsibilities are historically segregated – men tend to outnumber women in professions such as law enforcement, the military, and politics; women on the other hand tend to do more household chores than men. This can potentially cause different environmental exposures in men and women. Furthermore, biological sex can influence the use of personal care products, such as menstrual and intimate care products and hormonal contraceptives.

Figure 1: Proposed framework to integrate exposomic analysis in healthcare. (Adapted).

Gender identity can likewise influence the use of personal care products including cosmetics and other beauty products. This is partly because such products are marketed mainly to women, female-identifying, and feminine-presenting consumers, who are the primary targets of unrealistic beauty standards. Studies show that women use approximately twice as many personal care products each day compared with men, which results in higher exposure to chemicals and toxic substances, including exposure to carcinogens, nanoparticles, and metals.

Since the environment itself as well as the environmental exposure are constantly changing, assessing the environmental factors at a specific point in time provides limited insight into collective exposures or exposures during key developmental periods. In contrast, exposome-wide characterization, which features the use of HRMS-based assays, can profile a variety of biospecimens in an untargeted and unbiased way to simultaneously identify both exogenous factors and endogenous responses to those exposures.

Exposomics as a field is still in its early stages. However, there have been some initial studies that looked at the differences in the metabolomic profiles of men and women, as defined by biological sex, in healthy and disease states. These studies have revealed baseline differences based on biological sex such as creatinine content (a chemical waste product produced by our body), steroid hormones, and branched-chain amino acids.

Going forward, new frameworks, as summarized in Figure 1, can systematically incorporate exposomic characterizations in health outcomes and interventions. For example, new digital databases (that include e.g., environmental data, chemicals, and toxicokinetics), biobanking (e.g., BioBank procedures), analytical platforms such as HRMS, and computing power (e.g., cloud computing services) need to be constructed in order to characterize the exposome on multiple levels depending on the research questions asked. Once this has been achieved, questions can focus on how the differences in exposome profiles identified lead to altered health outcomes, and further incorporate these findings into basic science research, clinical trial design, and data science approaches.

Such a parallel exposomic platform may initially seem intractable. However, the National Institutes of Health (NIH) recently launched a new initiative titled “All of Us”, aimed at gathering data from more than one million US citizens. This database could be an excellent resource of data from which to build a more complete understanding of the exposome.

An intersectional approach – sex- and gender-specific environmental exposure and its evaluation for impact on health –  with a rigorous effort to not only include but center women, sexual and gender minorities, and BIPOC individuals in health research will help to remedy our current dearth of understanding regarding sex- and gender-specific health outcomes. The findings can be translated into educational efforts, among stakeholders, scientists, and the public, to increase awareness of the role of the environment in sex- and gender-specific health. This can in turn inform policymaking regarding the regulation of environmental factors and exposures. Ultimately, such research will help to manage individual health risk assessment and precision medicine, where individual behaviors may be geared to improve health.

Reviewers: Trang Nguyen, Maaike Schilperoort

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​How a virus invading a cell limits another virus access to the same cell

Alphaviruses can infect both vertebrate and invertebrate animals.  Their transmission between species and individuals occurs mainly via mosquitoes. These viruses are small, spherical, and have a genome composed of a single strand ribonucleic acid (RNA) in the “positive-sense”. The alphaviral life cycles and their RNA genome amplification (replication) have been studied since their discovery in 1953. However, the very initial events of viral genome replication have remained unknown.

Positive-strand RNA viruses genome can be directly translated into viral proteins with the participation of factors and structures provided by the invaded host cell. However, in order to amplify the viral genome and to produce new viral particles during the virus propagation, the positive RNA strand has to be converted to its complementary negative strand by an enzyme that is encoded in the viral genome. This enzyme uses RNA as a template to synthesize RNA, a so-called RNA-dependent RNA polymerase (RdRp). RdRp are used during replication of the genome to synthesize a negative-sense antigenome that is then used as the template to create a new positive-sense viral genome, necessary for the future viral progeny and viral propagation (Figure 1).

Figure 1. Overview of the alphavirus life cycle. Alphaviruses enter the cell by recognizing a cell receptor, followed by release into the host cell of the viral plus-strand genome (1). The genome serves as a template carrying the information for production of a fused version of viral proteins (viral polyprotein, 2). This polyprotein is cleaved to different combinations (not shown) constituting an RNA-dependent RNA polymerase, and two forms of protein complexes required for viral replication (3 and 4). The consecutive cleavage of the polyprotein has been shown to influence transitions in production between the full-length minus-strand RNA, the genomic plus strand, as well as of another form of viral RNA (not shown) required for subsequent viral particles (nucleocapsid) assembly and release (5 and 6). Figure adapted from the original paper.

A phenomenon known as superinfection exclusion has been previously observed, where infection by one virus can block the infection of a subsequent homologous virus. This form of viral competition protects the virus to complete its reproduction without the need to share the cell’s resources with homologous viruses or with its own progeny. One of the mechanisms of superinfection inclusion can be by reducing the host cell receptors that the virus uses to recognise and enter into the cell. However, such changes in the cells are thought to take place several hours upon infection and for some viruses the phenomenon of superinfection exclusion has been observed as soon as just 15 minutes of the first infection (Figure 1). This rapid competitive behavior was observed over 40 years ago. This mechanism providing such rapid protection over a secondary infection is very beneficial, especially considering the ability of the virus to enter cells within minutes. However, its causes, as well as the very earliest stages of alphaviral replication and whether the two processes are linked has remained unclear.

Previous studies of the alphavirus’s life cycle have mainly used populations of infected cells. The use of recently developed single cell-based methods allows to overcome several limitations of population-level studies. For example, the classic population-based studies have shown the average growth of the virus over time across millions of cells and have revealed that the first release of viral progeny can be detected as early as 3–4 hours post infection (Figure 1). However, there is an inherent cell-to-cell variability in the infection spreading in a group of cells. Use of single-cell analyses in biology has shown how the variability of individual cells can be masked by the overall population’s behavior and how variability between individual cells contribute to viral growth and spreading kinetics. An important challenge on how the dynamics of early replication could affect the competitive interactions is the lack of sensitivity on low-abundance targets during early infection. In order to capture the dynamics of the earliest stages of replication, it is necessary to utilize an approach with sufficient sensitivity to simultaneously measure individual molecules of multiple viral RNA species at low abundance.

The recent work published from Columbia postdoc Zakary Singer and his colleagues presents a new quantitative detailed characterization of the initial replication activity of members of the alphavirus genus, Sindbis virus. The study consists in analyzing the viral genome biology at the level of individually affected cells and not in a group of cellular population. The authors used quantitative live single cell imaging technique to follow and measure the viral replication in real time upon infection as well as to elucidate how these contribute to the rapid exclusion of a superinfecting alphavirus. Singer and colleagues observed that the rapid onset of viral RNA synthesis as a passive superinfection exclusion mechanism could contribute to this advantage. Furthermore, a mathematical model of exponential viral growth in a resource-limited environment appeared consistent with the measurements of viral replication. The authors also investigated whether there is a bidirectional inhibition between two viruses in the same cell, by experimental measurements and a mathematical modeling of competitive growth using parameters estimated from single-virus infection experiments. The results from both methods suggested that the superinfecting virus is equally able to reduce replication levels of the first virus and that the cell appears to have fixed carrying capacity that sets up the combined replication level of the two viruses. Due to the speed of Sindbis replication would strongly disadvantage the second virus and reduce the second virus’s replication, showing the importance of intrinsic growth kinetics in alphaviral superinfection exclusion.

The work by Singer and colleagues also allowed to shed light on classic questions remaining in alpha virology and suggested a revised model of early replication wherein both plus- and minus strands are made at a similar rate during early infection in contrast to previous claims that initially the positive strand RNA production is predominant. Additionally, the paper provides one of the earliest detections of alphaviral replication, as well as a new framework for understanding early replication and the resulting exclusionary phenomenon. Finally, the work hints on how in the future the complex interplay with innate immunity and stochasticity will be broadly relevant to the study of many infectious diseases, and how quantitative models might lead to improved antivirals. Check out more from the original publication.

Edited by: Sam Rossano and Trang Nguyen

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