Crispr – Page 2 – BioQuakes

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Fighting Cancer with CRISPR

On April 27, 2023

In Student Post

For many years the treatment of cancer has remained difficult and uncertain. Though there are many treatment methods such as chemotherapy and bone marrow transplants, these methods are never guaranteed to work. However, a teenage girl named Alyssa diagnosed with T-cell acute lymphoblastic leukemia (T-ALL) has been successfully treated with a new experimental treatment. T-ALL is a type of cancer where cancerous T-cells overpopulate healthy T-cells, leaving the patient susceptible to disease. In this form of cancer, T-cells also mistake each other as threats. Due to the nature of this cancer, in order for treatment to be effective, T-cells would have to appear foreign to the patient’s immune system. This is made possible through the gene editing system, CRISPR. For Alyssa’s treatment, doctors utilized and altered donated T-cells. Using CRISPR, the donated T-cells were stripped of CD7 protein, a common T-cell protein, and CD52 protein, a protein recognized by cancer treatment. Additionally, donated T-cells received a receptor that gave them the ability to target cancerous and healthy T-cells by having the ability to recognize CD7. All of these changes were made through a process called base editing with CRISPR. During base editing, individual letters, or bases, in the T-cells’ DNA code were altered. These minor alterations have the ability to change the nature of the cell. Thanks to this new treatment, Alyssa’s cancer is now undetectable.

I found T-ALL cancer and its destructive nature relatable to the way that viruses take over human body cells, however, our adaptive immunity uses antigens to recognize an intruder. T-cells contain specific proteins which make them recognizable to other T-cells, including cancerous ones. T-ALL destroys the body’s own T-cells which is why this specific treatment needed to use altered T-cells that did not contain recognizable proteins. When the body is infected by a virus, memory T and B cells use antibodies, a little piece of the virus, to remember and recognize the virus if it were to enter the body again.

CRISPR May Be the Cure!

On April 23, 2023

In Student Post

There are still many disorders and diseases in this world that cannot be cured, and Huntington’s disease (HD) is one of them.

HD is a neurological disorder that causes individuals to lose control of movement, coordination, and cognitive function. HD occurs because of a mutation in the Huntingtin (HTT) gene where a specific codon sequence repeats, creating a long, repetitive sequence that turns into a toxic, expanded protein clump. These clumps form in a part of the brain that regulates movement called the striatum and prevent the neurons in the striatum from functioning properly. As of now, HD still has no cure, but CRISPR gene editing (Clustered Regularly Interspaced Short Palindromic Repeats) might just be the solution.

Dr. Gene Yeo of UC San Diego School of Medicine, along with his team and colleagues from UC Irvine and Johns Hopkins University, researched RNA-targeting CRISPR/Cas13d technology as a way to possibly eliminate HD and its negative effects on the brain. CRISPR gene editing, as its name suggests, enables scientists to “edit” – add, remove, or alter – existing genetic material. The group desired to see if RNA-targeting CRISPR would be able to prevent the creation of the protein clumps that damage the function of the striatum. As we learned in AP Biology, the addition, removal, or substitution of a base of a codon can drastically change the structure and function of a protein. Each codon codes for a specific amino acid, and if multiple codons have changed due to a mutation, it is likely that the protein will fold differently than it is supposed to and will lose its function.

Yeo and his team desired to develop an effective therapy for HD, hoping to stop the formation of toxic protein clumps and alter the course of the disease. However, they did not want to create permanent changes in the human genome as a precaution. The team instead engineered a therapy that alters the RNA that turns into the protein clumps. They conducted testing on mice and found that RNA-targeting CRISPR therapy reduced toxic protein levels in a mouse with HD, improving motor coordination. In connection with the molecular genetics unit in AP Biology, since the RNA that causes HD is altered, the protein that is translated will change since different amino acids correspond to different codons.

Further testing will be necessary to confirm the benefits of this therapeutic strategy, but CRISPR does look like a promising medical treatment for HD and many other diseases in the future.

Progress in Treating Huntington’s Disease Thanks to CRISPR Technology

On April 23, 2023

In Student Post

Scientists have discovered a new way to treat Huntington’s disease, thanks to CRISPR technology. Their research has reduced symptoms of the disease in the mice that they tested on.

Huntington’s Disease, which is a neurological disorder, is caused by a genetic mutation in the HTT gene. More specifically, repetitive and damaging sequences in the HTT gene cause Huntington’s disease. It causes progressive loss of movement, coordination, and cognitive functions.

Researchers have discovered a possible solution to these symptoms: CRISPR technology.

According to the article, “CRISPR is a genome-editing tool that allows scientists to add, remove or alter genetic material at specific locations in the genome.” One of the risks of CRISPR use is that it can affect off-target genes and molecules, causing unwanted alterations in chromosomes and genes.

Study author Gene Yeo, PhD, explains how our cells struggle to copy repetitive DNA, which can lead to errors that cause repetitive sequences to increase with each generation. As we learned in class, the process to copy DNA is a complex one where there are many factors at play. DNA is replicated in a semi conservative manner, meaning that the old DNA strands are conserved and combined with the new, complementary strands. There is a replication fork, with a leading and lagging strand, on which DNA is replicated in the 5’-3’ direction. For replication on the leading strand, RNA primase adds RNA, DNA polymerase III adds nucleotides to the open end of the RNA, then a sliding clamp attaches to the DNA polymerase III and slides it along the strand, resulting in the leading strand being synthesized.

The scientists directly targeted the RNA involved in the DNA replication process to remove toxic protein buildup that is responsible for the mutation in the HTT genes. They were able to complete this process using CRISPR, and without disrupting other important genes.

After testing on mice, they reported that their research has resulted in improved motor coordination, less striatal degradation and reduced toxic protein levels. These improvements on the mice’s condition lasted for up to 8 months, and had no on other RNA molecules, making scientists optimistic that this treatment could be effective for humans.

A Potential Cure: We’ve Waited 151 Years For This!

On April 23, 2023

In Student Post

CRISPR, which stands for Clustered Regularly Interspaced Short Palindromic Repeats, is a gene editing tool comprised of DNA sequences from prokaryotes, that is becoming more commonly used to treat and potentially cure life-threatening diseases that have previously been viewed as a death sentence; in December of 2022, a study was conducted at the University of California San Diego School of Medicine, where it was discovered that CRISPR technology can be used to target the gene that causes Huntington’s Disease.

First, we must understand what exactly Huntington’s Disease is. Huntington’s Disease, which was discovered in 1872, is a rare neurological disorder characterized by the gradual destruction of nerve cells in the brain. It is caused by a single defective gene, and this mutation is as dangerous and tragic as it is rare; the disease has no cure, and patients typically do not survive beyond 20 years post-diagnosis.

However, thanks to CRISPR, it is a very real possibility that that will soon change.

The study that was conducted at U.C. San Diego involved the experimentation of Cas13d – an RNA editing technique – against toxic RNA and protein buildup that is associated with the HTT gene mutation that causes Huntington’s Disease, and the trial was found to be successful in terms of eliminating that buildup. The experiment was conducted on mice, and it was also discovered that only one injection of the Cas13d therapy was necessary to yield results, and the benefits (improved motor function, lessened symptoms) lasted for eight months.

This discovery is especially fascinating as it connects to our AP Biology units in terms of mutations: The most common genetic mutations are insertions, substitutions, and deletions. The mutation that causes Huntington’s Disease, however, fits into neither one of these categories: if anything, the mutation is considered a duplication, as it is characterized by the unwanted repetition of cytosine, adenine, and guanine; these repetitions are what lead to the protein buildup, and damage the HHT gene.

In previous years leading up to the U.C. San Diego experiment, trials conducted to target the gene that causes Huntington’s Disease have mostly been unsuccessful, but we can hope that this new discovery is a step in the right direction and may provide the key to figuring out how to treat this disorder that has historically been viewed as a death sentence.

CRISPR technology may be the key to treating Huntington’s Disease

On April 23, 2023

In Student Post

Huntington’s disease is a well-known neurological disorder that is characterized by a loss of movement, coordination, and cognitive function. More than 200,000 people live with Huntington’s disease and more than a quarter million Americans are at risk of inheriting the diseases, but there is currently no cure. Scientists have recently been trying to develop a treatment using RNA-targeting CRISPR/Cas13d technology to eliminate toxic RNA that causes Huntington’s disease. CRISPR allows scientists to edit, add, and remove genetic material from specific places in the genome. This tool is based on a immune-defense mechanism from bacteria. Since there is a risk of editing a part of the genome unintentionally, studies have focused more on targeting RNA directly instead.

Huntington’s disease is caused by a mutation in a gene for the protein huntingtin. This mutation, known as a trinucleotide repeat expansion, causes cytosine, adenine, and guanine to be repeated many more times in the gene that normal. As a result, the protein that is produced can form toxic clumps in the part of the brain responsible for movement, which is called the striatum.

In AP Bio, we learned how genes are used to produce proteins that our bodies use. First, the RNA polymerase creates mRNA from transcribing DNA. A guanine nucleotide is added to the 5′ end, a poly-A tail is added to the 3′ end, and the introns are cut out. Then, the mRNA leaves the nucleus and goes to a ribosome for translation. The anticodon on the tRNA matches with the codon on the mRNA, which brings along the corresponding amino acid. The amino acid connects with the next amino acid to create a protein molecule. The additional CAG sequences in the huntingtin gene are transcribed onto the mRNA, which is then used to create a polypeptide. Since it is longer than normal, this protein’s shape will be deformed and will be toxic to the brain.

In neuronal cultures from patients with Huntington’s disease, scientists have used CRISPR to destroy mutant RNA molecules and clear out toxic protein buildup. Other genes were not affected by this treatment. When tested in mice, scientists found that the mice had better motor coordination and less toxic protein levels.

I chose this topic because I am very interested in how scientists can use mechanisms seen in other organisms to help treat human diseases.

The Quest for the Cure: Could Modified CAR T-Cells Using CRISPR Technology Be the Key?

On April 23, 2023

In Student Post

Cancer: difficult to talk about, but even harder to cure. Unfortunately, most, if not all of us have lost loved ones due to cancer, the disease which took the lives of over 600 thousand Americans in 2022. Lymphoma, one of the most common types of cancer, is cancer of the lymphatic system, which is part of the body’s immune system and includes the lymph nodes, bone marrow, thymus gland, and spleen. Although scientists have not yet discovered a cure for it, scientists across the globe have made significant amounts of discoveries which could help treat lymphoma.

One promising treatment option is CAR (chimeric antigen receptor) T- cell therapy, discovered in 2002 by scientists at Memorial Sloan Kettering Cancer Center. As we learned in AP Biology, T-cells are part of the body’s immune system, and some (helper T-cells) recognize antigens while others (T-Killer cells) kill infected or cancerous cells. Helper T-cells recognize fragments of the antigen on the surface of macrophages and signal for the immune system to destroy the infected cell, a task that is often carried out by T-killer cells. When scientists modify T-cells in a lab by inserting a gene for a chimeric antigen receptor (CAR), the T-cells can better identify and bind to cancerous antigen fragments and signal for the body to destroy the cancer cells. After modifying the T-cells, they are inserted back into the patient.

However, one major problem with this treatment is that the T-cells often get “T-cell exhaustion,” which is when they are effective and efficient at first but quickly become worn-out and ineffective. To combat this problem, Dr. Michel Sadelain altered the T-cells’ genes through the use of CRISPR technology.

CRISPR (clustered regularly interspaced short palindromic repeats) technology is a cutting-edge method of genome editing which physically cuts and/or replaces nucleotides in DNA.

As we learned in AP Biology, DNA, the genetic “code” in every living thing, is composed of two strands, in the shape of a double helix, which are made of a phosphate and deoxyribose sugar “backbone,” and bases which attach to each sugar. There are four bases: A (adenine), T (thymine), G (guanine), and C (cytosine). A base, sugar, and phosphate together form what is called a nucleotide, and these are what CRISPR replaces or deletes.

DNA codes for all characteristics of a living organism, so when it is altered, the genetic makeup and traits change. This is an essential fact when considering CRISPR technology, since it can alter the base sequence within DNA, causing potentially life-saving gene alterations. Dr. Sadelain used this technology to insert a gene in T-cells which makes them more resilient and less prone to T-cell exhaustion. Dr. Park of Memorial Sloan Kettering recently proved Dr. Sadelain’s discovery to be effective in a clinical trial which used Sadelain’s method to use CRISPR to insert CARs and a molecule called 1XX into T-cells which were then injected into patients with lymphoma. The alterations Sadelain made were intended to create T- cells that work efficiently for longer periods of time and therefore will be more effective in destroying lymphoma cells. The results of the trial were very promising since it was very successful, safe, and used a relatively small amount of modified T-cells, meaning that the treatment could be accessible to more people if it is approved. Do you think this treatment method will be a success?

As confirmed through this trial, the capabilities of CRISPR and CAR T-cell therapy are evolving to a mind-blowing extent and are providing safer and more effective treatments for cancer and other diseases every year. Although a universal cure for cancer has not yet been discovered, the discoveries of this study could alter the future of lymphoma treatment and save the lives of thousands.

Overcoming a Critical Limitation of CRISPR

On April 23, 2023

In Student Post

Recent research demonstrates that CRISPR Spherical Nucleic Acids (SNAs) can be delivered across the cell membrane and into the nucleus, all while retaining bioactivity and capability of gene editing. Gene editing is technology which allows a scientist to change an organism’s DNA.

The work displayed in this article builds on a 25-year study to uncover the properties of SNAs and the factors that distinguish them from the blueprint of life. SNAs are structures typically composed of spherical nanoparticles covered with DNA or RNA, giving them chemical and physical properties different from those forms of nucleic acids found in nature.

A variety of SNAs exist, with cores and shells of different chemical compositions and sizes. SNAs are also now being evaluated as potent therapeutics in human clinical trials, such as trials for brain cancer and skin cancer.

According to nanotechnology pioneer Chad A. Mirkin, “these novel nanostructures provide a path for researchers to broaden the scope of CRISPR utility by dramatically expanding the types of cells and tissues that the CRISPR machinery can be delivered to.” “We already know SNAs provide privileged access to the skin, the brain, the eyes, the immune system, the GI track, heart and lungs. When this type of access is coupled to one of the most important innovations in biomedical science in the last quarter-century, good things will follow.”

Mirkin’s team used Cas9 (protein required for gene editing) as the core of the structure, and attached DNA strands to the surface to form a new type of SNA. These SNAs were also preloaded with RNA capable of performing gene editing and fused with peptides to control their ability to navigate compartmental barriers of the cell, making it the most efficient. In AP Biology, we learned that peptides are molecules containing two or more amino acids. Peptides that contain several amino acids are called polypeptides or proteins. These SNAs effectively enter cells without the use of transfection agents, and display high gene editing efficiency between 32% and 47% across several human and mouse cell lines.

If You Give A Mouse…Sight!

By Allisteric Site

On April 23, 2023

In Student Post

In a recent study published in the Journal Of Experimental Medicine, researchers in China successfully used CRISPR Gene-Editing technology to restore sight to mice with retinitis pigmentosa.

That’s a lot of vocabulary all at once, so let’s establish some definitions first and foremost. According to the National Eye Institute, retinitis pigmentosa is a “genetic disease that people [and animals] are born with…that [affects] the retina (the light-sensitive layer of tissue in the back of the eye)”. As for CRISPR Gene-Editing technology, YG Topics defines it as, “a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence”.

Most inherent forms of blindness and loss-of-vision stem from genetic mutations, and thus retinitis pigmentosa is one of many forms of genetically caused blindness. However, through CRISPR technology, the researchers in the study successfully edited the DNA of mice who had the mutation to eliminate retinitis pigmentosa and give them the ability to see. The results of the study are very promising, as not only does retinitis pigmentosa affect mice, but human beings. Thus, there is evidence that CRISPR could be used to cure blindness among everyday people. Kai Yao, a professor from the Wuhan University of Science and Technology who contributed to the study said, “The ability to edit the genome of neural retinal cells, particularly unhealthy or dying photoreceptors, would provide much more convincing evidence for the potential applications of these genome-editing tools in treating diseases such as retinitis pigmentosa”.

In AP Biology, we discussed how DNA factors into the traits of a living being. DNA is made up of 3 base codons that form up to 20 different amino acids. These amino acids code for specific proteins. Through a process of DNA transcription and translation, the DNA uses various forms of RNA to code for proteins, which help tell the cell what to do. Thus, the way the cell acts is largely determined by its DNA. Essentially, DNA codes certain traits through various amino acid sequences. Mutations and alternations to amino acid sequences cause different traits, such as red hair, blue eyes, or blindness.

Thus, successfully altering the DNA of mice has huge implications for the human race. CRISPR could potentially be used to edit the DNA of humans, and thus help limit and prevent certain genetic conditions. Many diseases are based on genetic mutations, and if CRISPR Gene Editing technology is proven successful, we could potentially eliminate genetic diseases in a few decades. While “much work still needs to be done to establish both the safety and efficacy” of CRISPR technology, some groundbreaking scientific treatments could be coming sooner than you think (Neuroscience News).

The “Most Complicated” Cancer Treatment EVER

On April 23, 2023

In Student Post

There are many approaches to treating cancer, ranging from invasive surgeries to extremely damaging radiation and chemotherapy. The teeny-tiniest clinical trial ever began at UCLA in yet another attempt to find another way to eradicate cancer. With only 16 participants, this trial combined two areas of research: gene editing and T-cell engineering. The reason for the miniscule sample size is the intensely customized nature of the treatment. Each patient’s tumor had completely unique mutations, so each patient needed equally unique T-cell engineering through gene editing.

One reason cancer is so hard to treat is because they have adapted to be resistant to the body’s own immune response. The patients that have cancers, especially ones in the later stages, have lost the battle against their cancer with their own immune system, so a new super-immune system must now be built. This army of new T-cells (white blood cells, which identify and kill bad cells, seen below) will need “training” for its difficult battle ahead. First, however, the researchers must determine how to train these cells so they will actually be successful. They used algorithms to find identifiable mutations in the tumor, something that the T-cell can seek out to differentiate the cancerous cells from the normal cells.

After testing to make sure that the T-cells can actually identify these mutations, T-cell receptors are designed specifically to their tumor. Then, each patient’s blood is taken so that the DNA code for the new receptors can be inserted using CRISPR, a genome editing technology at the cutting edge of genetic medical research. The DNA code is transcribed to mRNA, which is then used in the ribosome to build polypeptides, in this case, the receptor proteins for the T-cells. In order to ensure that these new T-cells (with the special receptors) are received, the patients had to take medication that suppressed the number of immune cells, so that the ones they are given can take hold.

One month into treatment, 5 of the patients’ tumors stopped growing, and only 2 of the participants had associated side effects. Although only 5 patients had the desired results, Dr. Ribas, one of the researchers, says that they “need to hit it stronger the next time” because they were limited to a small dosage of T-cells to start in order to establish safety. Additionally, the technology will only get better and better as the research progresses and the T-cells can have more and more mutation targets to look for in a tumor.

CRISPR: how one tool can change an entire generation (of invasive mice).

On April 23, 2023

In Student Post

In recent years, technology has heavily impacted scientists abilities to change the world. CRISPR is a recently discovered gene editing tool that is revolutionizing the way scientists are treating patients and curing diseases. Recent research has also found that CRISPR can be used to help mice infestations in certain parts of the world (random, but cool)!

Mice infestations are a problem in many islands, and CRISPR is here to help. Scientist believe they had found a way to make an entire mice population extinct (in a few decades) by using gene editing via CRISPR. In order to understand exactly how scientists plan on doing this, it is important to understand what a “haplotype” is. A haplotype is a set of genes that are inherited by the next generation together. The “regular house mouse” has what’s called a “t-haplotype,” and it’s passed down roughly 95% of the time (a lot compared to the normal 50%). The study states that male house mice with two t-haplotype copies become infertile, and females with two t-haplotype copies will become sterile as well. As we know from AP-Biology, when an organism has two copies of some gene, it is known as homozygous- meaning it has two of the same alleles of some gene. In this case the phenotype that makes the mouse homozygous would be the altered t-haplotypes. If a mouse has two of these altered t-haplotype genes, it becomes sterile and cannot reproduce.

CRISPR plays a crucial role here – by using gene editing through CRISPR technology, scientists are able to edit the t-haplotype of the M. musculus house mouse so that next time a male M. musculus mates with a female, the offspring will become infertile. That’s right, CRIPSR can be used to completely alter and wipe an an entire M. musculus population over the course of a few years. By using computer technology, scientists predict that by adding just 256 “altered” mice to a certain island population of mice, an island of 200,000 mice can be fully wiped out within about 25 years.

Scientists are hopeful, optimistic, and invested in CRISPR technology. The “25 years later” prediction is a long time to wait, and scientists hope that sometime in the future, CRISPR will be able to work faster, allowing problems to be solved more quickly and more efficiently. I think that this study is an important part of CRISPR potential, and it makes me very curious to see what CRISPR has in store for the future, and what other kinds of animal related issues it can help solve.

New CRISPR Technique can Potentially be a Treatment for Leukemia

On April 23, 2023

In Student Post

An article published on December 11, 2022 on newscientist, shares fascinating information on a 13 year old patient with leukemia, having no detectable cancer cells after being the first person to receive a new type of CRISPR treatment, to attack cancer.

The 13-year-old leukemia patient, Alyssa, has had many treatments that have been unsuccessful in helping her condition. Leukemia is caused by immune cells in the bone marrow dividing and growing rapidly. This relates to what we learned about in Biology class in how cancer cells become cancerous by cells dividing uncontrollably. It is also related to how cancer is caused by changes to the DNA (mutations) that alter important genes and change the behavior of them. Leukemia is also caused by the mutations in DNA.

The most common treatments for leukemia are known as killing all bone marrow cells with chemotherapy and then replacing it with a transplant. If this treatment is unsuccessful, an approach known as CAR-T therapy is used. This involves adding a gene to a type of immune cell known as a T cell that causes it to destroy cancerous cells. This also relates back to how in biology class we learned about the functions of T- cells being vital because they protect us from infection. The modified cells are called CAR-T cells. Alyssa’s leukemia was caused by T cells so if they used this technique to modify CAR-T cells to attack other T cells, it would lead to these cells killing each other. Wasseem Quasim at the University College London Great Ormond Street Institute of Child Health, has discovered many drawbacks with this treatment. Due to the many problems conventional gene editing can cause, Qasim and his team used a modified form of the CRISPR gene-editing protein, and Alyssa is the first person ever to be treated with. Alyssa received a dose of immune cells from a donor that had been altered to attack the cancer, and tests revealed 28 days later she had no signs of cancer cells. CRISPR is technology that can be used to edit genes. It finds specific DNA inside a cell and then changes that piece of DNA. It has also been discovered that CRISPR can be an effective tool for cancer treatment. This new approach to CRISPR treatments could be hugely beneficial to cancer patients and Many other treatments involving CRISPR base editing are being developed.

Analyzing Viruses in One Step Using LUNAS

On April 23, 2023

In Student Post

In this article from the American Chemical Society, Maarten Merkx and his colleagues research how to use and combine CRISPR-related proteins with a bioluminescence technique whose signal could be detected with a digital camera. This new technique can diagnose illnesses faster while being much more efficient and practical. Bioluminescence is a chemical reaction involving the luciferase protein that causes the luminescent, glow-in-the-dark effect. The luciferase protein has been incorporated into sensors that emit an easily observed light when they find their target, but they lack the incredibly high sensitivity required of a clinical diagnostic test.

CRISPR, a gene-editing technique, has the ability to increase sensitivity, but it requires many steps and additional specialized equipment. With the new technique, called LUNAS (luminescent nucleic acid sensor), two CRISPR/Cas9 proteins specific for different neighboring parts of a viral genome each have a distinct fragment of luciferase attached to them. If a specific viral genome that the researchers were testing for was present, the two CRISPR/Cas9 proteins would bind to the targeted nucleic acid sequences and come close to each other, allowing the complete luciferase protein to form and shine blue light in the presence of a chemical substrate. RPA-LUNAS successfully detected SARS-CoV-2 RNA within 20 minutes, even at concentrations as low as 200 copies per microliter.

This is similar to the process of gene regulation that uses an inducible operon as we learned in class. An inducible operon is a type of negative regulation that turns on when it interacts with an inducer. It is usually off which means there is an active repressor that binds to the operator to block the RNA polymerase from transcribing the DNA. When there is the inducer, such as the virus, the inducer inactivates the repressor by binding to the allosteric site which allows the RNA polymerase, such as the CRISPR/Cas9 proteins, to transcribe and eventually produce the protein, such as the luciferase protein.

As we are recovering from the devastating COVID-19 Pandemic, how can new medical advancements and technology help us prepare for future outbreaks?

The Fluorescent Frontier: Glow in the Dark Proteins in Disease Research

On April 23, 2023

In Student Post

We all know that although science is improving rapidly on a global scale, diagnostic tests for diseases remain sensitive and require complicated techniques. One evident example is the tests for COVID-19. This complexity can range from their preparation to an interpretation of their results. However, recent research from the American Chemical Society has developed a method that is able to analyze viral or infected nucleic acids in less than 30 minutes and in just one step. This is all due to “glow in the dark” proteins.

Bioluminescence is a scientific phenomenon that powers many animals: a firefly’s flash, an anglerfish’s glowing head, and even phytoplankton’s blue color. Here a chemical reaction occurs, involving the luciferase protein. This protein essentially causes the “glow in the dark” effect. The protein is incorporated into sensors which emit a light when a target is located. Although the simplicity of these sensors is idyllic for clinical diagnostic testing, they still lack the sensitivity

One solution to this problem is presented by a particular gene editing technique: CRISPR. The Broad Institute defines CRISPR as; Clustered Regularly Interspaced Short Palindromic Repeats. It is essentially an efficient and customizable alternative to other existing genome editing tools. With this new technique, Maarten Merkx and his coworkers wished to use CRISPR-connected proteins while combining them with a bioluminescence form whose glow could be seen by humans, through a digital camera for example.

To ensure that there was an ample amount of DNA or RNA to analyze, they used a technique known as Recombinase Polymerase Amplification, or RPA. This is a simple method which works continuously at a temperature of 100 F. With this two CRISPR proteins specific for different parts of a viral genome each have a different fragment of luciferase attached. In other words, the new treatment known as LUNAS (Luminescent Nucleic Acid Sensor), takes two CRISPR proteins for different parts of a viral genome and has a distinct fragment of luciferase added to each.

Moreover, if one specific viral genome that the researchers were testing was present, the two CRISPR proteins would bind to the targeted nucleic acid sequence. This would allow them to come together and promote the full luciferase protein to form and glow. Additionally, to account for the luciferase being depleted, the researchers used a control reaction which turned green. In the event of a positive viral detection the color would change from green to blue. To prove the validity of this method, the researchers tested LUNAS on clinical samples of nasal swabs testing COVID-19. The method successfully detected the virus in less than 20 minutes, even at low concentrations. With this, the LUNAS method holds great potential in detecting other viruses in a concise and efficient manner.

To connect to our AP Bio class, we learned about how specific proteins code for specific actions or results in our bodies. At their tertiary and quaternary structures, proteins have a myriad of functions ranging from acting as a receptor to interacting with an enzyme. This parallels with the luciferase’s specific function of creating a glow affect. Additionally we learned about cell communication and how interaction with a receptor would result, or cause a specific occurrence. This connects to luciferase’s binding to its sensor, causing the glow affect. This cell communication also connects to the two CRISPR proteins attaching to a specific nucleid acid sequence. If the nucleid acid holds the viral genome and the luciferase, it would connect and form a glow response – a direct example of intercellular communication. Continually, we learned about DNA manipulation and alteration and how segments can be added in, substituted, or even removed. This occurs in CRISPR gene editing’s nature as a genome editing tool. It exemplifies all these manipulations to both DNA and RNA. We also learned about ideal protein function at a variety of temperatures, pHs, and environmental settings. This idyllic setting in seen in RPA’s function at a continuous 100 F.

To close, I feel that the use of luciferase, or “glow in the dark” proteins fronts an entirely new way of combating diseases and supporting disease identification. It would provide a new way for doctors and scientists to diagnose patients in a time efficient manner. And frankly, the idea of being diagnosed by something “glow in the dark” is entirely lightening and provides some relief to the gravity of the situation. I invite any and all comments regarding this specific method of disease identification or any other relevant discussion points.

Biological Warfare: Bacterial Edition

On March 15, 2023

In Student Post

In February 2023, a study was published announcing that bacteria possess something similar to humans that can activate and deactivate immune pathways, and therefore this “something” could be used to cure diseases; that “something” is called the ubiquitin transferase enzyme.

Biological warfare, the use of infectious agents to kill diseases caused by other infectious agents, has been considered as a potential solution in the past. In fact, years prior, a family of DNA sequences now referred to as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) were discovered in bacteria, and it was determined that these sequences were capable of killing other phages and being used to cure infections.

Our immune pathways, as we learned in our immunity unit in AP Biology, is crucial for our survival as a species. Our immune system consists of innate immunity, involving natural killer cells that serve as our first line of defense against pathogens, and adaptive immunity, involving B cells and T cells that need to be trained to fight these pathogens. Our immune pathways alone, however, cannot rid us of neurodegenerative diseases, and these diseases still unfortunately have no cure..

One may be wondering now, how can the ubiquitin transferase enzyme work to treat diseases like Parkinson’s? How does it help our immune pathways? Well, the answer to that is protein editing. The enzyme contains two proteins, CD-NTase-associated protein 2 and 3 (also known as Cap2 and Cap3); these proteins are what serve as the activation and deactivation for immune pathways: they can direct old, unnecessary, or damaged proteins to be broken down.

When the potential of CRISPR was discovered, scientists used genome editing to direct the machine so it would kill its targeted diseases. A similar attempt could be made with the ubiquitin transferase enzyme.

Finding the existence of this in bacteria especially is an amazing discovery, as not only does it propel us in the right direction in terms of potentially curing Parkinson’s or other neurodegenerative disorders, but it connects back to our other lesson in AP Biology that humans and bacteria are not so different after all. We share about a thousand genes!

It is particularly interesting knowing how biological warfare could be used to help us.

The Science of Love

On March 6, 2023

In Student Post

The neurobiology of love is not as simple to figure out as one may think. Throughout time, researchers have studied the prairie vole when looking to discover more about what exactly is going on in our brains when we fall in love. The prairie vole is used because they form monogamous relationships, in which they show empathy for and display actions that we would describe as love. The monogamy of prairie voles was tested and proven nearly 50 years ago, and since then, we have been researching prairie voles to discover the neurobiology of their love.

When first looking at exactly what happens in a prairie vole’s head when they “fall in love” the hormones oxytocin and vasopressin were found to play a key role in the bonding of prairie voles. These hormones work just as any hormone we have learned about in this AP Biology class does, they are peptides that bind to receptors, resulting in the change of the shape of the receptor, resulting in a change within the cell. This signal caused by oxytocin and vasopressin has been shown have the ability to change a solitary or polygamous prairie vole, into one that forms a monogamous bond with another prairie vole. This is unique to other voles, as the receptors for these hormones in prairie voles are in a separate location to those within other types of voles. This discovery opened up a doorway into understanding how the location and abundance of hormones receptors can affect the bonding of animals or even ourselves. In order to truly understand this though, researchers had to find a way to manipulate the ways that genes encode these receptors.

Vasopressin Oxytocin

Although this phenomenon has been well studied in the past, the invention of CRISPR technology has opened many more doors into studying the ways that these hormone receptors work. Throughout time, it has been accepted that oxytocin in the true hormone that controls the bonds of these prairie voles. But, after a research team used CRISPR to “delete the gene that encodes the oxytocin receptor in prairie vole embryos,” it was found that the genetically modified prairie voles were still able to form bonds as easily as their non-modified brethren. Scientists are now trying to really figure out what the biology of the love of these voles is, as their original hypothesis that oxytocin controls it has been found incorrect. Scientists are now looking to hormones such as vasopressin, to see if that was the missing part of the puzzle they need. This discovery that oxytocin is not the entire basis of the bonding of prairie voles does show that as usual, love is less complex than we may think, and that they must look at the neurobiology of love as having a much bigger scale than before thought of.

Currently, scientists are aiming to look at the activity of genes in the brain to further look to what causes love bonds. Recent studies like this have shown that after the voles have mated, genes important to memory and learning are turned on, and that their brain’s reward structure turns on after a stable amount of time with a bond. These studies have also proven that the voles brains become “activated,” once they have created a bond, as their neural activity “lights up” and their brains “re-wire themselves.”

“How does this relate to us humans?” you may ask. Well, the ways that prairie voles react to their bonds can show us what an evolved brain, which neurologically is wired to have a partner. The studies done on prairie voles inspired scientists to look back closer at the reward structures pf the brain looked at in studies for prairie voles, and it was found that humans had similar responses to the voles. Overall, with much more time to let research and technology develop, studies on prairie voles have definitely taken us one step closer in understanding the science of love.

CRISPR Provides New Hope for those with Huntington’s Disease

On March 1, 2023

In Student Post

Intro

Neuron with mHtt inclusion

Neurons transfected with a disease-associated version of huntingtin

Huntington’s disease is a neurological disorder that affects the basal ganglia and cerebral cortex of the brain. These areas of the brain are associated with movement, learning, thinking, planning, motivation, and emotion. Huntington’s disease is caused by a single mutation in the huntingtin (HTT) gene, afflicting more than 200,000 people worldwide and 30,000 in the United States. There was believed to be no cure, however, novel research regarding CRISPR gene editing is giving those who suffer from this condition new hope.

Identifying the problem (and connection to AP Bio)

“Our cells have a hard time copying repetitive DNA, and these copying errors can cause repetitive sequences to grow longer with each generation,” (Gene Yeo, PhD).

Huntington’s disease is caused by repetitive and damaging sequences in the HTT gene. Within the cell cycle, in order for the cell to divide into two daughter cells during mitosis, the cell’s DNA must be replicated in the synthesis phase. In Huntington’s disease, the damage done to the HTT gene is carried through the synthesis phase, causing everlasting effects on future generations of cells. These repeated genes amass to many times their normal length and result in toxic clumps which aggravate the striatum of the brain which is important in regulating movement; thereby leading to Huntington’s Disease.

Inventing a solution

CRISPR is a tool that edits genomes by precisely cutting DNA and then letting natural DNA repair processes take over. The system consists of two parts: the Cas9 enzyme and a guide RNA. In this new study, Gene Yeo and his team of researchers at the University of California San Diego School of Medicine are using RNA-targeting CRISPR/Cas13d technology to develop a new therapeutic strategy that specifically eliminates toxic RNA that causes HD. Yeo delivered the CRISPR therapy through viral vehicles to neuronal cultures grown from the stem cells of an individual with Huntington’s syndrome. His team has found that the approach not only targeted and destroyed mutant RNA molecules but also cleared out toxic protein buildup without disrupting other genes.

Predictions for the future

Yeo’s team collaborated with Wenzhen Duan’s team at Johns Hopkins to conduct preclinical testing in mice. They found that the CRISPR therapy improved motor coordination, attenuated striatal degradation and reduced toxic protein levels in a mouse model of HD. The therapy lasted for at least 8 months and caused minimal effects on other RNA molecules. Although a mice’s anatomy is nowhere near as complex as a human’s, this new research gives incredible insight into the future that CRISPR holds and how impactful its use can be.

TMEM251: A Gene Come True?

On October 19, 2022

In Student Post

Have you ever finished a painfully long math problem, only to realize you made a mistake in the very first step? Although you probably panicked because your final answer was embarrassingly far away from the correct one, fixing your one small mistake may have actually revealed that the mess was not truly all that complicated. Identifying the root of a seemingly drastic problem quite frequently uncovers the underlying simplicity of the matter, but who would have thought that this basic concept could save generations of victims of a viciously lethal disease?

Mucolipidosis type II is an extremely rare inherited disease that causes physical, mental, and visual deformities and usually claims its victims before they turn seven years old (oftentimes much sooner). What causes Mucolipidosis type II? Simply put, it is when lysosomes do not receive the enzymes necessary to digest materials, making them not only ineffective, but also dangerous. When lysosomes are unable to perform the necessary recycling functions that you have learned about in AP Biology, materials are instead stored in the cell, causing Mucolipidosis type II and its seemingly infinite list of tragic symptoms such as scoliosis, neurological disabilities, ectrodactyly, enlargement of the heart, and more.

In fully functional cells, mannose-6-phosphate biosynthetic pathway, or M6P, signals the transport of hydrolytic enzymes into the lysosomes. Inversely, when M6P is either not functional or not present, the hydrolytic enzymes do not make it to the lysosomes, causing Mucolipidosis type II. Now that we know what happens when M6P is dysfunctional, let’s take one more step back. What causes M6P to stop working in the first place? Well, it turns out that a team of scientists at the University of Michigan had the same question.

Using CRISPR technology, the team tested individual genes’ effects on cellular functions, and they found the answer to our previous question: TMEM251. This gene is responsible for creating an enzyme called GNPT, which signals M6P to transport the enzymes to the lysosomes, and when this one singular gene fails to work, it causes the lifelong adversities of Mucolipidosis type II. Also, TMEM251 is located in the Golgi apparatus, and you (hopefully) already learned in AP Biology that lysosomes are created from this organelle. Therefore, this fact further supports the validity of the new finding.

The disease currently has a 100% fatality rate because scientists have not yet discovered a cure. Or have they? Now that scientists have identified the root of the problem, non-functional TMEM251 genes, they are experimenting with enzyme replacement therapy- the supplementing of missing lysosomal enzymes into the cell through endocytosis– to rehabilitate the cells to their proper, functional forms. It may be too early to place bets, but this groundbreaking discovery could turn out to be the hope that scientists around the globe have been searching for.

Can CRISPR Gene Editing Cause Problems in the Embryos it is Meant to Customize?

On March 25, 2022

In Student Post

Researchers from around the Tri-State area came together in 2020 to examine the effectiveness of the Crispr-Cas9 double stranded DNA break (DSB) induction on human embryos to repair a chromosomal mutation. The study, which was published in Cell, began with sperm from a mutated male patient at the EYS locus, which causes retinitis pigmentosa blindness. The researchers then attempted to use CRISPR-Cas9 technology to repair the blindness gene in a number of fertilized embryonic stem cells that carried the EYS mutation. The results showed that about half of the breaks in the experiment went unrepaired, which resulted in an undetectable paternal allele. After mitosis, the loss of one or both the chromosomal arms was also common. This study shows that using CRISPR-Cas9 technology is still in its early days, and needs to be further vetted before it is used to treat patients.

Instead of correctly and consistently editing the genome of the embryos, the Crispr-Cas9 wreaked havoc, leaving behind chromosomal trauma. The data shows that the embryos started to tear apart and get rid of big pieces of the chromosome that had the EYS mutation, some losing the entire chromosome. The promise of Crispr technology is about changing one gene, but how can that be done when a larger, untargeted part of the genome is also being altered? Dr. Egli, the paper’s main author, brought up a more likely use for the Crispr editing: deploying it as a form of “moleculure bomb”, sent in to shred unwanted chromosomes. An important part of using gene editing is the ability to consistently predict the outcome, However, the resulting “mosaicism prevents inferring the genotype of the fetus from a biopsy and is thus incompatible for clinical use”, according to the Cell authors.

There were many rarities that appeared in the alleles of the embryos used. With a small sample size, due to the difficulty to acquire human embryos, there was no ability to rule out rare events. Although there were combinations of maternal and paternal alleles that showed interhomolog events, they occurred after the two-cell-stage injections, all mosaic. A single Cas9-induced break can result in outcomes in the human embryo that suggest species-specific differences in repair. In on-target sequencing of the cells, the detection of only a wild-type maternal allele might have been because of the unrepaired breaks and the loss of the chromosomal arm or the loss of the entire chromosome. This study shines light on the dangers of Crispr gene editing. The quotes from researchers, doctors, and genealogists all echo the same risk, we must walk before we can run. Testing and ensuring the safety of using Crispr on an embryo before the first round of DNA replication happens is crucial to the ultimate promise of gene repair. If it can’t be done safely with no off target effects, then Crispr “would be deeply unethical”, according to Dr Faraheny from Duke University.

CRISPR can help in the detection of Kidney Rejection sooner than you thought

On March 20, 2022

In Student Post

Kidney Transplant

Diagram of a Transplanted Kidney

As of now, the only way to diagnose acute rejection is through a biopsy. This procedure can only detect problems when they are in a late stage. Doctors would be able to begin anti-rejection medication sooner if there was a way to non-invasively diagnose kidney rejection at an early stage. Well, there might be now….

Researchers have found an early warning sign of rejection in the urine of kidney transplant patients, a cytokine protein called CXCL9. Currently, the method used for measuring the protein (an enzyme-linked immunosorbent assay, or ELISA) has been unsuccessful. However, Jonathan Dordick and colleagues have been working to develop a better technique.

Kidney Transplant

How CRISPR works

They have based their new detection method on a gene-editing technology called CRISPR/Cas12a. The CRISPR/Cas12a enzyme cuts a probe to produce a fluorescent signal when in the presence of the CXCL9 protein. Then by attaching a DNA barcode that aggregates a large number of CRISPR/Cas12a molecules, they were able to boost the fluorescent signal. This then led to an antibody that recognizes CXCL9.

Another essential thing to note is that, unlike different CRISPR-based detection methods, the use of PCR amplification is not required. This makes it easier to modify to a device that could be used in more accessible ways, like in a doctor’s office or at home. When tested, the new system accurately measured CXCL9 levels for 11 kidney transplant patients. Since the immuno-CRISPR system is about 7 times more sensitive than an ELISA, kidney rejection can now be detected early.

Can Cancer Cell’s Medication Immunity Be Stripped?

By biodougversity

On March 20, 2022

In Student Post

Cancer is one of the hardest diseased to fight. If a tumor begins to grow inside of a patient, they may be given drugs to fight off the corrupt cells. The problem with this is that the cancer cells could become immune to these drugs. Through the use of CRISPR. In Novel Crispr imaging technology reveals genes controlling tumor immunity, a new way of fighting cancer is revealed. Instead of targeting the whole tumor, Perturb-map marks cancer cells and the cells around cancer cells. Once this is completed, it is able to identify genes controlling cancer’s ability to become immune to certain drugs.

To fight cancer cells, scientists use thousands of CRISPRs at the same time. This identifies every gene in a sequence and allows them to be studied. Through Perturb-map, scientists can now dive deeper and find where the cell immunity to drugs originates. A certain pathway in the cell is controlled by the cytokine interferon gamma or IFNg, and a second is by the tumor growth factor-beta receptor or TGFbR. When the cell had a gene with TGFbR2 or SOCS1, the latter of which regulates IFNg, tumor cells grew. When the cell lacked one of these, it shrunk. Moreover, it was discovered that tumors with SOCS1 were susceptible to attacks by T cells, but TGFbR cells had immunity against them. This stayed true even when both types of cells lived in the same environment. With findings like these emerging more and more, the future of cancer treatment is looking brighter than ever.

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