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The Science Explained

The Science Explained

Thursday, March 5, 2020

WHAT IT IS

Chaperones are proteins that are best known as regulators of protein folding. Chaperones help new proteins fold correctly, help refold misfolded proteins, and regulate disposal of proteins that cannot be refolded. Collectively, chaperones, co-chaperones, and other factors make up the so-called chaperome, which is responsible for protein homeostasis in the cell.

Two chaperones, heat shock proteins 70 (HSP70) and 90 (HSP90) play especially critical roles in protein homeostasis, since they act as the core of short-lived multi-protein scaffolds that carry out much of the folding and refolding work of the chaperome.

Work by Gabriela Chiosis, PhD, of the chemical biology program at Memorial Sloan Kettering Cancer Center in New York City, has revealed that in many types of cancer cells, these normally short-lived scaffolds become stabilized, a condition she has termed the epichaperome, to stress that, akin to epigenetic (post-translational) changes in other proteins, the change is not in the level of expression of the individual chaperones, but in the structure and behavior of the complex.

HOW IT WORKS

Because the chaperome is composed of over 200 dynamically interacting proteins, much remains to be learned about the detailed function of the chaperome in cellular health and disease. Research by Dr. Chiosis suggests that prolonged cellular stress is a trigger for stabilization of chaperone platforms—that is, creation of an epichaperome complex. The epichaperome then affects a variety of cellular pathways, depending on the cell, the type of stress, and other context-dependent factors. Neurons are among the most sensitive cell types to disruptions of internal processes, and Dr. Chiosis's research suggests that epichaperome-mediated dysfunction can affect pathways that contribute to development of Alzheimer's disease.

HOW IT IS APPLIED

Because misfolded amyloid-beta and tau are the central pathology of Alzheimer's disease, attempts have been made to manipulate individual chaperones for therapy in the disease, including through upregulating and downregulating their activity or expression. To date, these attempts have not led to clinical success. An epichaperome inhibitor is currently undergoing early clinical trials in three types of cancer. Identification of the role of the epichaperome in AD models has led to development and early clinical testing of an inhibitor of epichaperome stabilization in that disease as well.

Thursday, November 21, 2019

WHAT IT IS

The assay exploits the seeded conversion of normal prion protein to the abnormal form and therefore detects disease associated prion protein in the CSF.

HOW IT WORKS

The assay amplifies small samples of misfolded protein collected from CSF in living patients and autopsied tissue from pathologically-confirmed cases. RT-QuIC usually starts with a small body fluid sample, usually CSF, collected from patients. Nasal brushings, urine, skin and eye components have been used for the assay. In 4-repeat tau RT-QuIC assays, a small drop of diagnostic specimen that contains the tau aggregate or “seed” is mixed with a vast excess of monomers of a recombinant 4-repeat tau fragment (that is, the substrate). The aggregates incorporate the substrate and begin growing into recombinant fibrils. A fluorescent dye is used to detect the fibrils as they accumulate.

HOW IT IS APPLIED

The assay has been used to detect abnormal aggregates of prion protein and tau fragments in CJD, tauopathies, Lewy body disease, and other neurodegenerative disorders.

Wednesday, September 11, 2019

WHAT IT IS

Optogenetics is a technique for precisely controlling the firing of specific neurons. It relies on a group of microbial proteins that change their properties in response to light of a specific wavelength. In the most widely used optogenetic system, light-sensitive membrane channels (“channelrhodopsins”) are introduced into a neuron. Cation channels will depolarize the membrane, while anion channels will hyperpolarize it, effectively switching the neuron “on” or “off” within milliseconds.

HOW IT WORKS

The gene for the protein is delivered by viral vectors. Cell specificity is achieved either by including a cell-specific promoter in the gene package, or, as in this paper, combining microinjection targeting with a genetic “homing” system for the transgene. Light is introduced with a fine optical fiber implanted into the target neurons.

HOW IT IS APPLIED

Optogenetics has been widely used throughout the brain to map neural circuits, by stimulating or inhibiting one type of neuron and recording the response in another. It has also been used to understand the inhibitory versus excitatory role of a specific neuronal type within a circuit, and to explore temporal patterns of communication between neurons. Therapeutic applications of optogenetics, for instance as a type of deep brain stimulator, are being explored, but will require gene therapy to deliver the proteins to the target neurons.

Wednesday, September 11, 2019

WHAT IT IS

Whole genome sequencing (WGS) is a comprehensive method that can rapidly sequence large amounts of DNA and provide genetic analysis of an individual's entire genome, including coding and non-coding regions of nuclear DNA. In the current study, the researchers used this technique to study the genome of single cells.

HOW IT WORKS

The scientists start with neuronal nuclei from postmortem brain. They individually sort single nuclei into wells on a microtiter plate. Then, then they amplify the entire genome from a single DNA copy using a highly efficient DNA polymerase that makes millions of copies of the original genome. They generate a tube of DNA as if it came from the blood of an individual, but it is obtained from a single cell's genomes. Then, they sequence that DNA sample from the single cell. The sequences are then mapped to the consensus genome and the variants that pop up in the sequencer that are different from the genome reference are annotated.

HOW IT IS APPLIED

WGS has been used mostly as a research tool to identify inherited disorders, mutations, and large and small variants that put people at risk for disease. More recently, costs for WGS have dropped substantially: It cost $350,000 in 2005 and today it can cost a few thousand dollars. It is being used clinically to diagnose disorders.


Wednesday, September 11, 2019

WHAT IT IS

Phage display immunoprecipitation sequencing helps identify antibodies that bind to human proteins in cerebrospinal fluid or serum.

HOW IT WORKS

Scientists lay out chunks of every human protein (about 700,000 autoantibodies across all human proteins) and then clone and express them in phage. Then, they incubate the patient’s CSF with the phage, allowing the antibodies to bind to phage expressing their peptide target or antigen. The scientists capture only the phage that stick to the patient antibodies by using magnetic beads, and they wash the rest away. They then observe which proteins were bound by the patient’s antibodies by sequencing the DNA in the phage.

HOW IT IS APPLIED

The technique enables researchers to display the complete human peptidome and discover autoantigen biomarkers common to different disorders, including breast cancer, neuroinfectious and autoimmune diseases.