Resources » Technical Resource Centers » Product Resources Center » Protein Analysis Library
Active and purified proteins play crucial roles in nearly all biological processes, especially in the fields of functional and structural analysis, assay development, and drug candidate discovery.
Trypanosome parasites are transmitted to mammals by the blood-sucking tsetse fly. The parasites' stopover in the insect-host has two stages. They live in the insect's gut for two to three weeks and then migrate to the saliva glands. When the fly has its next meal, the parasites are transferred via the saliva to the prey, infecting its bloodstream. In this way, the mammal now becomes host to the parasite, and the disease is spread. To complete their two-stage stay in the insect, the parasites must undertake an epic journey of active migration fraught with perils, such as the fly's digestive enzymes, immune system and the need to cross the intestinal barrier.
Research has shown that these insect-stage parasites are capable of group behavior, using cell-cell signaling to promote collective migration. By moving in numbers they are better able to transverse the fly's intestines, complete the journey to the saliva glands, and proliferate the disease. But how do these parasites communicate in order to coordinate their movements in response to signals from neighboring parasites?
Scientists interfered with the parasite communication system by inducing "stress" in parasite cells causing them to release exosomes. They found that the presence of these exosome-secreting cells disrupted the normal migration of the parasite cells. Parasite "scouts" which monitor the environment in the insect-host pick up "keep away" messages from these damaged cells, and, in turn, communicate with the migrating parasite population, messaging them to avoid contact with the "unfit" ones. When exosome secretion was inhibited no effect on the migration was observed. These results strongly suggest that exosomes act as a repellent that drives the fit parasites away from either damaged cells or an unfavorable environment. Read more »
Salmonella is the lauding cause of bacteria-associated foodborne illnesses in the United States, according to a previous study. Thus, early detection of the pathogen by rapid and sensitive testing is important for preventing the illness.
In a newly published study, researchers artificially contaminated food with salmonella. They then tested the food samples using Salmonella-specific antibodies combined with the tyramide signal amplification technique. They found that salmonella was present after 15 hours and removed other microorganisms to clarify the laboratory results. This time frame is shorter than the 2–3 days required to detect salmonella in a culture.
The scientists compared their assay results with those of a sandwich-type enzyme-linked immunosorbent assay (ELISA) and an ELISA coupled with IMS; the developed assay was 25- and 4-fold more sensitive than standard ELISA and IMS-ELISA, respectively. Tyramide signal amplification combined with IMS can improve the sensitivity and specificity for detecting salmonella. The developed assay can be easily adapted for other foodborne pathogens and will contribute to improved food safety and public health. Read more »
As some individuals produce antibodies that can recognize a virus even two years after infection, scientists are working to train the immune system to produce these antibodies. When produced naturally, these antibodies cannot cure systemic infection, but may be sufficiently strong to prevent infection if induced by a vaccine. As a conceptual test, using mice genetically engineered to simulate the human immune system, the authors of the "sequential immunization" paper devised a method for training the immune system and producing a class of antibodies known as PGT121, which react to diverse strains of HIV.
The human immune system contains multiple different precursors, although few can give rise to PGT121 antibodies; thus, the researchers first genetically analyzed the antibodies to determine their naive states. Next, authors of the "tailored immunogens" paper created a series of viral protein structures, starting with HIV and working backwards, to eventually teach the antibodies to recognize multiple forms of natural HIV.
Based on their established principles for HIV vaccine development, the next step is to develop immunogens that exhibit high affinity for the antibody precursors present in humans (which may differ from those engineered into the mouse model). This next advance will enable the vaccine to locate and train the correct parts of the human immune system in a naturally diverse environment.
Source of the publication:
All cells eventually die. Without cell death, cells become immortal, and cell immortality leads to cancer. Cancer cells counteract the breakdown of telomeres by building them up as quickly as they are degraded. A cancer cell does this by spackling a telomere with the enzyme telomerase. When the telomerase finds and attaches to a telomere, it adds a repeating DNA sequence to the repeating DNA sequences that are already in place, thereby lengthening the telomere and adding to the chromosome's protective ends. Oncologists and cancer researchers wish that telomerase would not do this. Without this constant telomere repair, chromosomes would eventually degrade and cancer cells would die.
Scientists at University of Colorado Cancer Center used CRISPR gene editing technology and live cell, single molecule microscopy to observe this essential interaction between the telomerase and telomeres, in a real-time manner. The group was able to visualize this process by using the CRISPR DNA editing technology to insert a code into the gene that synthesizes telomerase. This inserted code manufactured a fluorescent protein, which was attached to telomerase. The group then used nanoscopy to visualize the fluorescent protein. Read more »
The recently published manuscript by PNAS stated that new research tools and other developments have made it possible to decipher in detail how different cancers develop, as well as how benign or precancerous tissues become malignant and deadly.
The body's immune system is capable of intercepting pre-malignancies and preventing cancer. It does so countless times every day, and we wish to leverage this natural ability. Building upon our innate defenses against cancer is the foundation of new immunotherapies, which have shown great promise in a very short time; however, additional studies are needed. Efforts such as The Cancer Genome Atlas (TCGA) and genome-wide association studies (GWAS), which identify common genetic variants in different people to determine whether a variant is associated with a trait or, in this case, cancer, have significantly advanced the basic research of immune oncology and precision therapy. According to the authors, both should be dramatically expanded for cancer prevention.
The authors also cited several cancers for which immediate prevention efforts are required, including Lynch syndrome, an inherited condition that increases the risk of many cancers, including those of the digestive and gynecologic tracts; clonal hematopoiesis, an age-related precursor to leukemia; and cervical intraepithelial neoplasia, abnormal growth of cervix tissue caused by human papilloma virus that can progress to cervical cancer.
If the goal is to eradicate cancer, a radically new focus, investment and approach to
premalignant diseases,
and cancer prevention are requires and should be supported and sustained by broad, deep efforts such as the
Cancer Moonshot and Human Vaccines Project.
Read more »
Researchers at BUSM have been studying a protein called PTRF (Cavin-1) whose absence in model organisms and humans results in a nearly complete loss of fat cells. Due to this lack of fat cells, their fat is mis-targeted to other tissues, thereby causing insulin resistance, and eventually leading to the development of type 2 diabetes. Paradoxically, most obese individuals have an abundance of fat cells; however, they develop insulin resistance and their fat is mis-targeted to other tissues.
The researchers analyzed normal model organisms and compared them with those lacking PTRF. They also studied fat cells grown in the lab that either had PTRF or lacked it. Cells respond to rapid nutritional challenges by synthesizing new proteins to enable efficient storage of fat. These challenges correspond to those of the nutrient-rich diet common in Western society, and increasingly, worldwide. In the absence of PTRF, cells were unable to synthesize sufficient new proteins in response to cycles of fasting and refeeding, the equivalent of the human dietary cycle. Further studies described how PTRF functions at the molecular level, and the inefficiency of this process in most instances of obesity might be the link between the similar deleterious consequences of too little and too much fat.
Describing the role of PTRF and gaining a better understanding of how fat can be distributed in these models may eventually offer new opportunities for the treatment of diabetes in humans. However, the authors caution that type 2 diabetes is a complex condition and proteins other than PTRF can also contribute to the development of the disease. Read more »
In a report that appeared in the July 12 issue of JAMA, an HIV/AIDS themed issue, Huldrych F. Gunthard, M.D., of University Hospital Zurich, Switzerland, and colleagues, along with the International Antiviral Society-USA panel, updated the recommendations for the use of antiretroviral therapy in adults with established HIV infection, including when to start the treatment, initial regimens, and modified regimens, as well as providing recommendations for using antiretroviral drugs for preventing HIV among those at risk, including pre-exposure and post-exposure prevention.
There have been substantial advances in the use of antiretroviral drugs (ARVs) for the treatment and prevention of HIV infection since the last version of these recommendations were published in 2014, warranting an update to the recommendations. A panel of experts in HIV research and patient care convened by the International Antiviral Society-USA reviewed data published in peer-reviewed journals, presented by regulatory agencies, or presented as conference abstracts at peer-reviewed scientific conferences since the 2014 report for new data or evidence that would affect previous recommendations or their ratings. A comprehensive literature search was conducted. Recommendations were made by consensus, and each recommendation was rated by strength and quality of the evidence. Read more »
The microenvironment of tumors is a mix of cell types, mostly comprising inflammatory cells. White blood cells, recruited from the blood and bone marrow, represent a significant portion of these inflammatory cells and influence nearly all steps of tumor progression. One type of white blood cell, called as tumor-associated neutrophils (TANs), predominates in the tumor microenvironment; however, the role of TANs in tumor development remains largely unexplored in humans.
Researchers from the Perelman School of Medicine at the University of Pennsylvania have now identified a unique subset of TANs that exhibit hybrid characteristics of two immune cell types—neutrophils and antigen-presenting cells—in samples from early-stage human lung cancers. This is the first study to describe this phenomenon in a human tumor.
The goal of this research was to characterize TANs and determine their specific roles in the regulation of T cell responses in patients with early-stage lung cancer. This is particularly important because cytotoxic T lymphocytes are the chief effector cells mediating anti-tumor immunity. The findings demonstrate that the microenvironment of early-stage lung tumor can stimulate neutrophils to differentiate into a cell subset with enhanced anti-tumor capabilities. Read more »
Chimeric antigen receptor (CAR) therapies have emerged as a potent tool for treating hematologic malignancies. A major scientific challenge with engineered T cells is that this technology cannot be applied to solid tumors, as CARs mostly target shared antigens found on both normal and cancerous cells.
Recently, Scientists have developed and characterized a monoclonal antibody, 5E5, that specifically recognizes a sugar modification, Tn glycan on MUC1, widely expressed by adenocarcinomas. 5E5 is able to eliminate Tn-MUC1-expressing tumors in mouse models of leukemia and pancreatic cancer. Therefore, cancer-specific neo-epitopes formed by aberrant glycosylation might serve as potential targets for CAR T cell therapy in the treatment of a variety of adenocarcinomas and bone-marrow-derived cancers.
The potential of CAR T cells engineered to target aberrantly glycosylated cancer-specific proteins might provide a safer alternative as compared to targeting shared, overexpressed tumor antigens. In near future, CAR T cell therapy might undergo clinical trials, to test its safety, and efficacy in treating wide variety of solid tumors that express Tn-MUC1.
Phenolic compounds have antioxidant activity and increasing evidence suggest that this antioxidant activity affects the biochemical pathways linked to inflammation in mammals.
Researchers at the University of Illinois have identified candidate genes that control the accumulation of phenolic compounds in broccoli. They crossed two broccoli lines and tested their progenies for total phenolic content and their ability to neutralize oxygen radicals in cellular assays. Then, in the most promising progeny they identified the genes involved in generating phenolics, by using quantitative trait locus analysis.
By identifying the genes involved in accumulating the phenolic compounds, researchers are now one step closer to breeding broccoli and related Brassica vegetables like kale and cabbage with mega-doses of phenolic compounds.
The good news is that phenolic compounds are flavorless and stable; therefore, these vegetables can be cooked without losing the health-promoting qualities of phenolic compounds.
On consuming these vegetables, the phenolic compounds are absorbed and targeted to specific areas of the body, or are concentrated in the liver. Flavonoids spread through the bloodstream, reducing inflammation owing to their antioxidant activity.
Recently, a group of researchers developed a mathematical model based on evolutionary theories to demonstrate differences in sub-populations of tumors. Using this model, they discovered that cancer cells at the surface of a tumor, close to the surrounding environment, are predictably different from the cells within the interior of the tumor. Cells at the tumor surface invest their limited resources in developing cellular characteristics that promote invasion, and their ability to utilize resources from the surrounding environment, such as blood vessels. Tumor cells at the exterior surface develop these characteristics despite high risk of cell death.
The researchers confirmed these data by showing that the cells within the interior and exterior of breast cancer tissue display distinct gene expression patterns. Cells within the interior of the tumor have more static characteristics, such as less proliferation and more cell death. In contrast, cells on the surface of the tumor have higher proliferation rates and are more likely to produce an acidic environment, which is consistent with the need for cells on the tumor surface to grow and invade into the surrounding normal tissue.
This is an interesting finding and by promoting the non-invasive characteristics, tumor growth can be slowed down.
For the first time in history, researchers have isolated the parts of the human genome that could explain the differences in how humans experience happiness. These are the findings of a large-scale international study in over 298,000 people, conducted by VU Amsterdam scientists. The researchers found three genetic variants for happiness, two variants that can account for differences in symptoms of depression, and eleven locations on the human genome that could account for varying degrees of neuroticism. The genetic variants for happiness are mainly expressed in the central nervous system and the adrenal glands and pancreatic system. The results were published in the journal of Nature Genetics.
These findings, which resulted from a collaborative project with the Social Science Genetic Association Consortium, are available for follow-up research. This will create an increasingly clearer picture of what causes differences in happiness. The genetic overlap with depressive symptoms that have been found is also a breakthrough. This shows that research into happiness can also offer new insights into the causes of one of the greatest medical challenges of our time: depression. The research effort headed by the VU scientists is the largest ever study into the genetic variants for happiness. It was successfully completed thanks to the assistance of 181 researchers from 145 scientific institutes, including medical centers in Rotterdam, Groningen, Leiden and Utrecht, and the universities of Rotterdam and Groningen. Read more »
Research led by Northeastern's scientists is trying to answer how metastasis begins whether can we stop it. The paper, published in Biophysical Journal, reveals how the abnormal protein-fiber scaffolding of tumors and the agility of the cancer cells themselves come together in a perfect storm to enable the escape. The quantitative method the researchers developed to understand the cells' sliding ability could also lead to a new way to screen for effective cancer drugs and help diagnose the stage of a cancer early on.
To do so they developed a model environment that mimics protein fibers. First they stamped stripes of fibronectin on glass plates, making sure to represent various widths. Then they deposited the cells -- alternately hundreds of breast cancer cells and hundreds of normal cells -- on these fiber-like stripes and used a time lapse microscope to observe and quantify their behavior.
On fibers that were 6 or 9 microns wide -- the typical size of fibers in tumors -- half the breast cancer cells elongated and slid around the cells they collided with. Conversely, 99 percent of the normal breast cells did an about face.
In the future, the system could also alert cancer patients and Clinicians before metastasis starts. Studies with patients have shown that the structure of a tumor's protein-fiber scaffolding can indicate how far the disease has progressed. The researchers found that certain aggressive genetic mutations enabled cells to slide on very narrow fibers, whereas cells with milder mutations would slide only when the fibers got much wider. Clinicians could biopsy the tumor and meansure the width of the fibers to see if that danger point were approaching. Read more »
Salts in our brain decide whether we are asleep or awake. For the first time, researchers have shown that the level of salts in our body and brain differ depending on whether we are asleep or awake. A new study from the University of Copenhagen reveals that by influencing the level of salts, it is possible to control a mouse's sleep-wake cycle. The research has recently been published in the scientific journal, SCIENCE.
The authors claimed that those salts play a much larger and much more decisive role than previously imagined. The discovery reveals a completely new layer of understanding of how the brain functions. First and foremost, they learned more about how sleep is controlled. It may, however, also open up for a better future understanding of why some people suffer convulsive fits when staying awake all through the night.
The researchers have used mice to test whether injecting salt into the brain enables control of the mouse's sleep-wake cycle -- independently of neuromodulators. Neuromodulators are compounds such as, adrenalin, which plays a decisive role in our waking up every morning. The study shows that adrenalin and other neuromodulators change the level of salts surrounding the neurons and that the salt balance then decides whether the neurons are sensitive to stimulation in the shape of a touch. When we are awake, the salt balance makes neurons highly sensitive to stimulation, as opposed to the salt balance in the brain during sleep, where the level of salts makes it harder to activate the neurons. Read more »
Upon entry into a cell, the RNA genome of HIV is converted into DNA and becomes entwined with the cellular DNA. From here, CRISPR/Cas9 can be programmed to target a DNA sequence and cleave viral DNA. The problem is that HIV is notoriously good at surviving and thriving with new mutations, so while many viruses are killed by the targeted approach, those viruses that escape CRISPR/Cas9 treatment become more difficult to target.
The scientists who conducted this study claimed that when they sequenced the viral RNA of escaped HIV, the majority of the mutations that the virus has are nicely aligned at the site where Cas9 cleaves the DNA, which indicates that these mutations, instead of resulting from the errors of viral reverse transcriptase, are introduced by the cellular non-homologous end joining machinery when repairing the broken DNA.
The study, a collaborative effort involving researchers at McGill University, the University of Montreal in Canada, the Chinese Academy of Medical Sciences, and the Peking Union Medical College in China, serves as a cautionary tale for those who hope to apply CRISPR/Cas9 as an antiviral. The researchers do not believe the effort is futile; however, there are strategies that could overcome this limitation, such as targeting multiple sites with CRISPR/Cas9 or using other enzymes aside from Cas9. Once a solution is identified, the next barrier will be identifying ways to deliver the treatment to patients. Read more »
The Study to Assess Longer-term Opioid Medication Effectiveness (SALOME) found hydromorphone (HDM) to be as effective as diacetylmorphine (pharmaceutical-grade prescription heroin) for people who have not benefited from previous treatments, such as methadone or suboxone.
Led by researchers from Providence Health Care (PHC), the Centre for Health Evaluation and Outcome Sciences (CHÉOS) at St. Paul's Hospital and the School of Population and Public Health of the University of British Columbia (UBC), SALOME is the only clinical trial of its kind in the world.
A total of 202 participants in Vancouver were randomized in a 6-month double blind study to receive injectable hydromorphone or diacetylmorphine (DAM). The medication was administered at PHC's Crosstown Clinic under the supervision of an interdisciplinary team of physicians, nurses, social workers, and counselors.
Key findings of the research include:
Injectable hydromorphone is as effective as injectable diacetylmorphine for long-term street opioid users not currently benefitting from available treatments (estimated to be approximately 10% of the opioid-dependent population not currently in treatment).
Study participants on both medications reported far fewer days of street-heroin and other opioid use at 6 months (3-5 days per month) compared to almost daily illicit opioid use prior to being enrolled in the study.
Participants also reported a significant reduction in days of illegal activities (from an average of 14.1 days per month to < 4 days).
Nearly 80% of the participants were retained in treatment at 6 months.
Hydromorphone and diacetylmorphine are both safe when taken in a clinical setting. Of a total of 88,451 injections, there were 14 overdoses and 11 seizures, all of which were successfully managed in the clinic. If these events had occurred on the street, the outcomes may have been fatal. Read more »
Low levels of total vitamin D and bioavailable vitamin D can help predict a person's risk of major adverse cardiovascular events such as myocardial infarction, stroke, heart failure or death, according to a first-of-its-kind study from the Intermountain Medical Center Heart Institute in Salt Lake City.
The study evaluated 4200 participants between 52 and 76 years of age. One-fourth of the study participants were diabetic and 70% had coronary artery disease.
Clinicians measured the participants' vitamin D metabolite levels, which included components of vitamin D that are formed during metabolism, to determine the association of the metabolite with future major adverse cardiovascular events. Bioavailable vitamin D results from vitamin D being absorbed into the bloodstream without binding to surrounding proteins.
During metabolism, only 10-15% of total vitamin D is available in the body to act on target cells, as most vitamin D is bound to vitamin D binding proteins. Therefore, evaluating whether or not the proportion of vitamin D that can be used may be important as unbound vitamin D alone, such as bioavailable vitamin D, is available to act on target cells.
The study tested many different types of vitamin D, but found that measuring total and bioavailable vitamin D were most accurate in predicting harmful cardiovascular events. Read more »
Passive immunization against misfolded toxic proteins is a promising approach to treat neurodegenerative disorders. For effective immunotherapy against Alzheimer's disease, recent clinical data indicate that monoclonal antibodies directed against the amyloid-β peptide should be administered before the onset of symptoms associated with irreversible brain damage. It is therefore critical to develop technologies for continuous antibody delivery applicable to disease prevention.
A group of scientists addressed this question using a bioactive cellular implant to deliver recombinant anti-amyloid-β antibodies in the subcutaneous tissue. An encapsulating device permeable to macromolecules supports the long-term survival of myogenic cells for > 10 months in immunocompetent allogeneic recipients. The encapsulated cells are genetically engineered to secrete high levels of anti-amyloid-β antibodies. Peripheral implantation leads to continuous antibody delivery to reach plasma levels that exceed 50 µg/ml. In a proof-of-concept study, the scientists showed that the recombinant antibodies produced by this system penetrate the brain and bind amyloid plaques in two mouse models of Alzheimer's pathology. When encapsulated cells are implanted before the onset of amyloid plaque deposition in TauPS2APP mice, chronic exposure to anti-amyloid-β antibodies dramatically reduce amyloid-β40 and amyloid-β42 levels in the brain, decrease the amyloid plaque burden, and most notably, prevent phospho-tau pathology in the hippocampus. These results support the use of encapsulated cell implants for passive immunotherapy against the misfolded proteins, which accumulate in Alzheimer's disease and other neurodegenerative disorders. Read more »
RNA-programmed genome editing using CRISPR/Cas9 from Streptococcus pyogenes has enabled rapid and accessible alteration of specific genomic loci in many organisms. A flexible means to target RNA would facilitate alteration and imaging of endogenous RNA transcripts analogous to CRISPR/Cas-based genomic tools, but most RNA targeting methods rely on incorporation of exogenous tags. A group of scientists demonstrated that nuclease-inactive S. pyogenes CRISPR/Cas9 can bind RNA in a nucleic acid-programmed fashion and allow endogenous RNA tracking in living cells. The analysis showed that nuclear-localized RNA-targeting Cas9 (RCas9) is only exported to the cytoplasm in the presence of sgRNAs targeting mRNA, and accumulation of ACTB, CCNA2, and TFRC mRNAs was observed in RNA granules that correlates with fluorescence in situ hybridization. Read more »
The Aedes aegypti mosquito is a significant public health threat because the Aedes aegypti mosquito is the main vector for dengue and chikungunya viruses. Disease control efforts can be enhanced through reproductive manipulation of these vectors. Previous work revealed a relationship between male seminal fluid proteins transferred to females during mating and female post-mating physiology and behavior.
To better understand this interplay, a group of scientists used short-read RNA sequencing to identify gene expression changes in the lower reproductive tract of females in response to mating. The scientists characterized mRNA expression in virgin and mated females 0, 6, and 24 hours post-mating (hpm) and identified 364 differentially-abundant transcripts between the mating status groups. Surprisingly, 60 transcripts were more abundant at 0 hpm compared to virgin females, suggesting transfer from males. Twenty of these transcripts encoded known A. aegypti seminal fluid proteins. Transfer and detection of male accessory gland-derived mRNA in females at 0 hpm was confirmed by measurement of eGFP mRNA in females mated to eGFP-expressing males. In addition, 150 transcripts were up-regulated at 6 and 24 hpm, while 130 transcripts were down-regulated at 6 and 24 hpm. Gene ontology (GO) enrichment analysis revealed that proteases, a protein class broadly known to play important roles in reproduction, were among the most enriched protein classes. RNAs associated with the immune system and antimicrobial function were also up-regulated at 24 hpm.
Collectively, the results suggest that copulation initiates broad transcriptome changes across the mosquito female reproductive tract, "priming" her for important subsequent processes of blood feeding, egg development, and immune defense. Further transcriptome analysis provides a vital foundation for future studies of the consequences of mating on female biology and will aid studies seeking to identify specific gene families, molecules, and pathways that support key reproductive processes in the female mosquito. Read more »
Everyone gets hiccups. The majority of the time hiccups are completely harmless and are more of an irritant than a symptom of an underlying condition, but hiccups that last > 48 hours could potentially signal serious health complications.
Hiccups occur when the diaphragm and respiratory organs have a sudden, involuntary spasm. This spasm is usually followed by closure of the glottis (the slit-like opening between the vocal cords and the larynx) and a characteristic sound like that of a cough. Persistent hiccups are hiccups that last > 48 hours but < 30 days, while intractable hiccups are classified as hiccups that last> 30 days.
Occasional hiccups are mostly harmless.
Anything that causes your stomach to become distended can cause hiccups. Drinking alcohol, for example, can induce hiccups because alcohol consumption irritates the esophagus and may result in a flare-up of acid reflux.
If your hiccups last > 2 days, talk to your physician.
Sometimes intractable hiccups are diagnosed with tumors involving the brain, lymph nodes, or stomach, but hiccups can also indicate a stroke. It is unclear why many of these incidents occur.
How to test if your hiccup is a sign of illness
To quickly ease your occasional hiccup woes, you can hold your breath (for a short period of time) or breathe into a bag to ease hiccups Other methods to extinguish hiccups include putting a cotton swab in the back of your throat to induce a gag reflex, gargling with ice water, swallowing granulated sugar, biting a lemon, and pulling your knees to the chest to compress the chest. Pressing lightly on the eyeballs will also activate the vagus nerve and result in a reflex that hinders the spasm of hiccups.
Transcription factors dictate what genes are turned on or off in a cell during embryo development, thereby controlling the type of organ the cell will form. In a recent study published in Cell, three transcription factors that are crucial for heart development (NKX2-5, TBX5, and GATA4) were tracked, and it was revealed for the first time how the transcription factors interact on genomic and physical levels. Indeed, mutations in the genes that regulate these proteins are implicated in congenital heart disease.
To determine how transcription factors interact during development, the researchers created mouse embryos that were missing one or two of the proteins. A loss of one transcription factor resulted in known heart defects, while embryos missing two transcription factors only had a rudimentary heart, demonstrating the importance of both factors. Next, the scientists created cardiac cells missing the same transcription factors to determine how the transcription factors interact to affect gene expression. It turns out that transcription factors are like adolescents in love; the transcription factors exert a strong influence on one another, and if are separated, the transcription factors "get into trouble". The transcription factors are often adjacent to each other on the genome and the presence of both transcription factors is required to bind to DNA. If one of the proteins is absent, the other transcription factors go rogue on occasion, binding to places on the genome where transcription factors are not supposed to be located. This migration turns on genes that are supposed to remain off and silence other genes when the genes should have been activated.
The scientists claimed that the transcription factors must stick together, otherwise the other transcription factor "gets into trouble". Not only are those transcription factors vital for turning on specific genes, but transcription factor interaction is important to keep the transcription factors from going to the wrong place and turning on a set of genes that does not belong in a heart cell. Read more »
Intellectual faculties are the sum of many factors, the majority of which are genetic and inherited from parents. Each first-degree relative (parents, offspring, and siblings) has 50% of their genetic code in common, therefore 50% of the genetic factors that partially determine cognition. For example, depending on the additional factors involved, a 25 point drop in intelligence quotient (IQ) can determine whether or not a person has crossed the threshold of intellectual disability.
A group of scientists in Canada discovered that a missing copy of a region in chromosome 16 results in a 25 point drop in IQ in carriers. Addition of a copy in the same genomic region results in an IQ drop of approximately 16 points. To reach these conclusions, the researchers measured the intelligence of 700 family members who had at least 1 relative carrying the same genetic mutation on chromosome 16, which is known to predispose to autistic spectrum disorders. Even in the study participants with IQs considered to be normal, the researchers found a substantial 25 point drop in IQ associated with 16p11.2 gene deletions. Indeed, it is quite common for mutation carriers to show no mental health problems.
Further studies are needed to quantify the effects of all mutations associated with autism and characterize the additive effects that lead to this psychiatric disorder. "No single mutation can cause the whole set of clinical signs shown by these patients", concluded the scientists. Read more »
Using innovative technology, scientists from France have filmed the process by which an AIDS vaccine candidate triggers the immune response in vivo. This previously unseen footage clearly shows how the vaccine recruits the immune cells needed to destroy infected cells. These results, published in the journal Nature Medicine on 21 December 2015, shed new light on the mode of action and potential of this vaccine.
First, the scientists administered the vaccine to healthy mice, then observed in real time how cells from the immune system were mobilized to the lymph nodes, the organ where the vaccine response is developed, in just a few hours.
The inflammasome promotes the maturation of the chemical messenger, interleukin (IL)-1, but also induces macrophage death, thereby releasing this inflammatory messenger in the lymph nodes. This signal triggers a chain reaction which assembles several key players of the immune system in the lymph nodes, including killer cells, which are vital for the vaccine response.
These in vivo films have given scientists a detailed picture of the main stages in the mechanism of action of this vaccine and highlighted an important pathway that orchestrates the effective mobilization of the immune response. Read more »
Influenza causes serious illness among millions of people each year, resulting in 250,000-500,000 deaths, according to the World Health Organization. Those most at risk include infants < 6 months of age because infants cannot be vaccinated against the disease. A group of researchers have identified a naturally occurring protein that, when added to the flu vaccine, may offer protection to babies during their first months of life.
Infants < 6 months of age do not make antibodies when given the flu vaccine because the immune systems of these very young babies do not respond to the adjuvant, or additive, within the vaccine that boosts the body's immune response when confronted with a virus.
The adjuvant used in most vaccines is aluminum hydroxide, or ALUM. ALUM is an additive that essentially acts as an irritant to attract white blood cells (neutrophils) to the vaccination site. Neutrophils secrete the protein lactoferrin, which works with the immune system to impede the virus's ability to survive in the body. In premature and term infants, however, ALUM does not make immature immune cells work better. In this very young group, only the smaller amount of naturally occurring lactoferrin found near the vaccination site improves the immune response. Read more »
Movement and strength comes from the muscle fibers that make up a muscle group contracting and generating tension. Muscle weakening happens when the fibers contract slower and with less force. A team of Canadian researchers hypothesized that the constant physical training of older elite runners preserved their muscle fibers such that the fibers behaved like those of young adults. Muscle fiber samples were obtained from the quadriceps of older elite runners and non-athlete adults in the same age range. Those individuals were in their 80s and 90s and actively competed in the world masters track and field championships; everyone placed in the top four of their respective events.
Fiber contraction speed and force were compared to fibers from 23-year-old non-athlete adults. Muscle fibers from older non-athletes contracted considerably slower and weaker than fibers from young non-athletes. To the researchers' surprise, the muscle fibers of masters athletes contracted at a speed and force similar to older non-athlete adults, rather than the young adults. Success in high-performance sports in old age does not appear to be due to the maintained contraction capability of the fibers.
This study suggests that aging is associated with decreased muscle quality regardless of physical activity status; however, other studies have shown that muscle fibers can be arranged in a variety of ways to optimize strength, speed, and power of the whole muscle, so there are many structural ways to compensate for the reduced performance at the fiber level to maintain performance at the whole muscle level. Read more »
Two-dimensional gel electrophoresis (2DE) and SDS-PAGE are the most useful methods in protein separation. Proteins separated by 2DE or SDS-PAGE are then transferred to membranes, using a variety of methods, such as electrophoretic transfer, heat-mediated transfer, or non-electrophoretic transfer, for specific protein detection and/or analysis. Specific proteins can be detected using a chemiluminescence method. In a previous study, a group of scientists claimed to reuse transfer buffer containing methanol at least five times when transferring proteins from SDS-PAGE to a polyvinylidene difluoride (PVDF) membrane. It was mentioned that the scientists added 150–200 ml of fresh transfer solution each time for extended use as a result of a loss of transfer buffer. Finally, they tested the efficiency of each protein transfer by chemiluminescence detection; however, some researchers believe this method is neither accurate nor useful for protein analysis and believe it can cause background as well as inaccurate protein analysis.
First, losing 200 ml of buffer for every transfer is not normal.
Second, it is possible that methanol could be lost as a result of evaporation after each transfer.
Finally, it is important to mention that proteins in the gel, prior to electrophoretic transfer, are not fixed.
Therefore, reusing transfer buffer multiple times could cause background binding and inaccurate analysis results, and is therefore not recommended. Read more.
Moderate alcohol consumption has been associated with a lower risk of developing and dying from heart disease and stroke, but alcohol is known to damage brain cells, and given that dementia is a neurodegenerative disorder, drinking alcohol might be harmful in those with dementia. A group of researchers therefore wanted to determine if the same potentially positive association between alcohol and a reduced risk of cardiovascular death could be applied to 321 people with early-stage Alzheimer's disease. The research team set out to assess the impact of a 12-month program of psychosocial counselling and support, and tracked the progress for 3 years thereafter, accumulating a considerable amount of data. During the monitoring period, 53 (16.5%) of those with mild Alzheimer's disease died. Surprisingly, consumption of 2-3 units of alcohol every day was associated with a 77% lower risk of death compared with a tally of 1 or fewer daily units. Read more.
Some subtle artifacts can have significant deleterious effects on carefully planned and executed protein electrophoresis experiments. These artifacts include proteases that act at room temperature upon proteins in the sample buffer prior to heating, cleavage of the Asp-Pro bond upon prolonged heating of proteins at high temperatures, contamination of sample or sample buffer with keratin, leaching of chemicals from disposable plastic ware, and contamination of urea with ammonium cyanate. In addition, researchers are capable of committing mistakes, as follows: (a) calculating the cross-linking factor of a gel; (b) polymerization temperature and time for a polyacrylamide gel; (c) inducing aggregates in samples for electrophoresis; (d) titrating the running buffer in electrophoresis; (e) proper sample preparation; (f) amount of protein to be loaded on a gel; (g) sample buffer-to-protein ratios; (h) incompletely removing phosphate buffered saline from cells prior to cell lysis; and (i) overfocusing of IPG strip in two-dimensional gel electrophoresis. Taking proper precautions to all these factors can greatly help generate perfect experimental results. Read more.
The androgen receptor transcript is targeted by mi-124, acting as a tumor suppressor to broadly limit the growth of prostate cancer (CaP). A group of scientists recently unraveled the mechanisms by which miR-124 acts in this setting. CaP cell proliferation is inhibited by miR-124 in vitro and sensitizes CaP cells to inhibitors of androgen receptor signaling (ARSI). Notably, miR-124 could restore the apoptotic response of cells resistant to enzalutamide, a drug approved for the treatment of castration-resistant CaP. These scientists used xenograft models to examine the effects of miR-124 in vivo when complexed with polyethylenimine (PEI)-derived nanoparticles. Intravenous delivery of miR-124 was sufficient to inhibit tumor growth and to increase tumor cell apoptosis in combination with enzalutamide. Mechanistic investigations revealed that miR-124 directly down-regulated AR splice variants (AR-V4 and V7), along with oncogenic targets (EZH2 and Src) that have been reported to contribute to CaP progression and treatment resistance. Taken together, the results offer a preclinical rationale to evaluate miR-124 for cancer treatment. Read more.
Riboswitches are noncoding RNA structures located in messenger RNAs that bind endogenous ligands such as a specific metabolite or ion to regulate gene expression. As such, riboswitches serve as a novel, yet largely unexploited, class of emerging drug targets. Demonstrating this potential, however, has proven difficult and is restricted to structurally similar antimetabolites and semisynthetic analogues of their cognate ligand, thus greatly restricting the chemical space and selectivity sought for such inhibitors. A group of scientists reported the discovery and characterization of ribocil, a highly selective chemical modulator of bacterial riboflavin riboswitches that was identified in a phenotypic screen and acts as a structurally distinct synthetic mimic of the natural ligand, flavin mononucleotide, to repress riboswitch-mediated ribB gene expression and inhibit bacterial cell growth. These findings indicate that noncoding RNA structural elements may be more broadly targeted using synthetic small molecules than previously expected. Read more.
The global spread of dengue virus (DENV) has increased viral genetic diversity and appears to be associated with greater epidemic potential. The mechanisms governing viral fitness in epidemiological settings, however, remain poorly defined. A group of scientists identified a determinant of fitness in a foreign dominant (PR-2B) DENV serotype 2 (DENV-2) clade, which emerged during the 1994 epidemic in Puerto Rico and replaced an endemic (PR-1) DENV-2 clade. Furthermore, PR-2B DENV-2 produced increased subgenomic flavivirus RNA (sfRNA) levels during replication, as compared with genomic RNA. PR-2B sfRNA showed sequence-dependent binding to and prevention of tripartite motif 25 (TRIM25) deubiquitylation, which is critical for sustained and amplified retinoic acid–inducible gene 1 (RIG-I)-induced type I interferon expression. These findings demonstrate a distinct viral RNA-host protein interaction that is able to evade innate immune responses, thus demonstrating increased epidemiological fitness. Read more.
Protein gels are usually stained with colorimetric or fluorescent dyes; however, colorimetric and fluorescent dyes are toxic and require proper disposal. A group of scientists have claimed that curry spice can stain protein without destaining. Moreover, the spice, curcumin, is completely environmental-friendly. Although curcumin is insoluble in water, heat solubilization can make the staining sensitivity of curcumin equal to Coomassie blue, but curcumin does not require destaining. If you are using SDS-PAGE frequently and want to be an environmental-friendly scientist, please read this article. Read more.
People who live in tropical regions tend to adapt well to humid and hot conditions. The indigenous people of Greenland, the Inuits, have lived in extreme Arctic conditions for a long time. Can this climate adaptation be encrypted into genetic signatures? After a rigorous study, a group of researchers claimed that such encryption could occur. A scan of Inuit genomes for signatures of adaptation revealed that select alleles modulate fatty acid composition that could affect the regulation of growth hormones. The Inuits have been shown to have genetic and physiologic adaptations to a diet rich in omega-3 polyunsaturated fatty acids (PUFAs). Want to learn more about this interesting finding? Click here to Read more.
Western blot is a standard detection method to check a specific antigen or protein. Normally, a standard Western blot will require an overnight blotting step for antibodies to be fully incubated. A number of commercial Western kits have claimed to shorten the Western blot significantly, while a group of scientists reported a novel process that carried out an immunoblot in 30 min, allowing the entire process of electrophoresis, electrotransfer, and immunoblot to be completed in 1 hour. Want to check out how this system works? Click here to Read more.
Histidine phosphorylation remains unexplored in mammalian signaling pathways because of the phosphoramidate bond. However, a specific antibody that binds to phosphorylated histidine is a more direct method to study histidine phosphorylation. A group of scientists developed a set of monoclonal antibodies that specifically recognize only 1-pHis and 3-pHis, without binding to pTyrs. These antibodies can be used in immunoblotting, IF, and even affinity purification. Additional research using these antibodies has already revealed the roles of 1-pHis and 3-pHis in phagocytosis and mitosis. Want to learn more about this interesting finding? Click here to Read more.
Protein SDS-PAGE is a standard detection method; however, the staining and destaining process is the most time-consuming step. A number of reagent suppliers have developed non–Coomassie dyes to speed up experiments, and heating the gel after electrophoresis is an even more efficient way that is accepted by a lot of researchers.
A group of scientists tested the effect of heating SDS-PAGE gel and found that destaining requires a significantly longer time than staining; surprisingly, a 1.5-mm gel can take up to 20 min for destaining, even at a temperature of 70°C. If the thickness of your gel were 0.8 mm, you would need at least 8 min if heating at 65°C. Want to see how this system works? Click here to Read more.
Human IgG has been a forefront tool in clinical therapeutics over the recent decade and remains a key target in pathway research. From a therapeutics perspective, bispecific IgG is unique but is challenging to manufacture at industrial levels. A group of scientists developed a method to scale up the purification of bispecific IgG without any modifications that may appear. This IgG was called κλ-body by the authors because it has one heavy chain and two different light chains, one κ and one λ. This antibody is believed to have light-chain domains that are able to undergo robust downstream processing. Want to learn more about this interesting finding? Click here to Read more.
Immunoprecipitation (IP) is a technique to isolate a specific protein out of a solution
using an
antibody that binds to it. Antibody-protein complexes are removed from the solution with the addition
of an insoluble form of an antibody binding protein, such as Protein A or Protein G conjugated to
newly popular Magnetic Beads…
Learn more »
One of the most important factors for achieving successful crystallization is the
quality
of your
protein. Optimizing a protein's purity and homogeneity, while maintaining its composition and
structure will increase your chances for successful crystallization. Prior to crystallization, these
qualities should be evaluated…
Learn more »