Posts tagged protein

ScienceDaily (June 20, 2012) — A protein required to regrow injured peripheral nerves has been identified by researchers at Washington University School of Medicine in St. Louis.

These are images of axon regeneration in mice two weeks after injury to the hind leg’s sciatic nerve. On the left, axons (green) of a normal mouse have regrown to their targets (red) in the muscle. On the right, a mouse lacking DLK shows no axons have regenerated, even after two weeks. (Credit: Jung Eun Shin)

The finding, in mice, has implications for improving recovery after nerve injury in the extremities. It also opens new avenues of investigation toward triggering nerve regeneration in the central nervous system, notorious for its inability to heal.

Peripheral nerves provide the sense of touch and drive the muscles that move arms and legs, hands and feet. Unlike nerves of the central nervous system, peripheral nerves can regenerate after they are cut or crushed. But the mechanisms behind the regeneration are not well understood.

In the new study, published online June 20 in Neuron, the scientists show that a protein called dual leucine zipper kinase (DLK) regulates signals that tell the nerve cell it has been injured — often communicating over distances of several feet. The protein governs whether the neuron turns on its regeneration program.

"DLK is a key molecule linking an injury to the nerve’s response to that injury, allowing the nerve to regenerate," says Aaron DiAntonio, MD, PhD, professor of developmental biology. "How does an injured nerve know that it is injured? How does it take that information and turn on a regenerative program and regrow connections? And why does only the peripheral nervous system respond this way, while the central nervous system does not? We think DLK is part of the answer."

The nerve cell body containing the nucleus or “brain” of a peripheral nerve resides in the spinal cord. During early development, these nerves send long, thin, branching wires, called axons, out to the tips of the fingers and toes. Once the axons reach their targets (a muscle, for example), they stop extending and remain mostly unchanged for the life of the organism. Unless they’re damaged.

If an axon is severed somewhere between the cell body in the spinal cord and the muscle, the piece of axon that is no longer connected to the cell body begins to disintegrate. Earlier work showed that DLK helps regulate this axonal degeneration. And in worms and flies, DLK also is known to govern the formation of an axon’s growth cone, the structure responsible for extending the tip of a growing axon whether after injury or during development.

The formation of the growth cone is an important part of the early, local response of a nerve to injury. But a later response, traveling over greater distances, proves vital for relaying the signals that activate genes promoting regeneration. This late response can happen hours or even days after injury.

But in mice, unlike worms and flies, DiAntonio and his colleagues found that DLK is not involved in an axon’s early response to injury. Even without DLK, the growth cone forms. But a lack of DLK means the nerve cell body, nestled in the spinal cord far from the injury, doesn’t get the message that it’s injured. Without the signals relaying the injury message, the cell body doesn’t turn on its regeneration program and the growth cone’s progress in extending the axon stalls.

In addition, it was shown many years ago that axons regrow faster after a second injury than axons injured only once. In other words, injury itself increases an axon’s ability to regenerate. Furthering this work, first author Jung Eun Shin, graduate research assistant, and her colleagues found that DLK is required to promote this accelerated growth.

"A neuron that has seen a previous injury now has a different regenerative program than one that has never been damaged," Shin says. "We hope to be able to identify what is different between these two neurons — specifically what factors lead to the improved regeneration after a second injury. We have found that activated DLK is one such factor. We would like to activate DLK in a newly injured neuron to see if it has improved regeneration."

In addition to speeding peripheral nerve recovery, DiAntonio and Shin see possible implications in the central nervous system. It is known for example, that some of the important factors regulated and ramped up by DLK are not activated in the central nervous system.

"Since this sort of signaling doesn’t appear to happen in the central nervous system, it’s possible these nerves don’t ‘know’ when they are injured," DiAntonio says. "It’s an exciting idea — but not at all proven — that activating DLK in the central nervous system could promote its regeneration."

Source: Science Daily

ScienceDaily (June 7, 2012) — Scientists have discovered a new function for a protein that protects cells during injury and could eventually translate into treatment for conditions ranging from cardiovascular disease to Alzheimer’s.

Researchers report online June 7 in the journal Cell that a type of protein called thrombospondin activates a protective pathway that prevents heart cell damage in mice undergoing simulated extreme hypertension, cardiac pressure overload and heart attack.

"Our results suggest that medically this protein could be targeted as a way to help people with many different disease states where various organs are under stress,” said Jeffery Molkentin, PhD, lead investigator and a researcher at Cincinnati Children’s Hospital Medical Center and the Howard Hughes Medical Institute. "Although more study is needed to determine how our findings might be applied clinically, a possible therapeutic strategy could include a drug or gene therapy that induces overexpression of the protein in tissues or organs undergoing injury."

Thrombospondin (Thbs) proteins are produced by the body in cells where tissues are being injured, reconfigured or remodeled, such as in chronic cardiac disease. They appear in part of the cell’s internal machinery called the endoplasmic reticulum. There, Thbs triggers a stress response process to regulate production of other proteins and help correct or rid cells of proteins that misfold and lose their form and intended function. Misfolded proteins help drive tissue damage and organ dysfunction.

The researchers zeroed in on how one thrombospondin protein (Thbs4) activates cellular stress responses in mice bred to overexpress the protein in heart cells. They compared how the hearts of the Thbs4-positive mice responded to simulated stress and injury to mice not bred to overexpress cardiac-specific Thbs4.

Overexpression of Thbs4 had no effect on the animals prior to cardiac stress — although during simulated hypertension and cardiac infarction the protein reduced injury and protected them from death. Mice not bred for Thbs4 overexpression were extremely sensitive to cardiac injury, according to Molkentin, a member of the Division of Molecular Cardiovascular Biology and Cincinnati Children’s Heart Institute.

The researchers reported that overexpressed Thbs4 enhanced the ability of heart cells to secrete helpful proteins, resolve misfolded proteins and properly reconstruct extracellular matrix — connective tissues that help give the heart functional form and structural integrity.

Critical to the stress response process was Thbs4 activating and regulating a transcription factor called Aft6alpha. Transcription factors help decode genetic instructions of other genes to control their expression. In the case of Aft6alpha in the heart, it helps mediate repair processes. When Aft6alpha is activated by Thbs4, the endoplasmic reticulum in cells expands and the production of chaperone molecules and other repair proteins is enhanced.

Mice bred not to overexpress cardiac Thbs4 did not exhibit activated Aft6alpha or robust repair processes following cardiac injury, leading to their poor outcomes.

Molkentin said the research team continues to examine the Thbs-dependent stress response pathway to better understand the involved processes. This includes seeing how the pathway affects laboratory models of neurodegenerative diseases like Parkinson’s, Alzheimer’s and amyotrophic lateral sclerosis.

Source: Science Daily

February 19, 2012 

In two landmark papers in the journal Nature this week, scientists at The Scripps Research Institute report that they have identified a class of proteins that detect “painful touch.”

Scientists have known that sensory nerves in our skin detect pressure, pain, heat, cold, and other stimuli using specialized “ion channel” proteins in their outer membranes. They have only just begun, however, to identify and characterize the specific proteins involved in each of these sensory pathways. The new work provides evidence that a family of sensory nerve proteins known as piezo proteins are ion channel proteins essential to the sensation of painful touch.

The experiments in the new study were conducted in fruit flies, a model system for the sensory nervous system of mammals, where piezo proteins are also expressed, as well as in certain cell types in the ear, kidney, heart, and other tissues. Future studies will focus on the roles of piezo proteins in sensing sound, blood pressure, and related stimuli that press and/or stretch cell membranes.

"Researchers in this field have been trying for decades to identify pressure-transducing ion channel proteins that exist in mammals, and these piezo proteins are exceptionally strong candidates," said Ardem Patapoutian, a professor in the Department of Cell Biology and the Dorris Neuroscience Center at Scripps Research, and a senior investigator for both papers. "We now have solid clues that we can follow up to learn how the mechanotransduction pathway works and how it is disrupted in diseases."

The two papers appear online in Nature on February 19, 2012.

Following the Path of Clues

Patapoutian’s laboratory specializes in the study of sensory ion-channel proteins. When hit by a stimulus to which it is sensitive, one of these proteins typically will open its structure to allow charged calcium, sodium, or potassium molecules (“ions”) to flow from the fluid outside the cell into the cell’s interior. Ion channels that sense mechanical pressure are thought to open when the membrane in which they are embedded is distorted past a certain threshold. The resulting flow of charge can trigger other signals inside the cell, for example a nerve impulse within sensory neurons—and in a human, a sufficient number of these nerve impulses would be interpreted by the brain as a touch- or pressure-related feeling.

 In a highly cited paper published in Science in late 2010, Patapoutian and his colleagues reported that two mouse proteins of previously unknown function exhibited properties of mechanotransducers. Cells to which these proteins were added drew in positively charged ions when subjected to mechanical pressure. Bertrand Coste, the first author of the paper, named the two closely related proteins piezo1 and piezo2—the prefix “piezo-” being derived from the ancient Greek word for pressure or squeezing.

"Since these proteins bore little resemblance to known ion channel proteins, the next step for us was to confirm that they are indeed ion channel proteins," Patapoutian said. The new studies take this step and more.

In the first of the new studies, lead authors Bertrand Coste, Bailong Xiao, and their colleagues confirmed that piezo proteins are indeed ion channel proteins, and very large ones. “It assembles into a ‘tetramer’ complex of four piezo proteins, which appears to be the biggest plasma membrane ion channel yet discovered,” said Coste, a research associate in the Patapoutian lab. The protein sequences within piezo also suggest that its ion channel structure weaves through the cell membrane more than 100 times.

Collaborating researchers in the laboratory of Mauricio Montal, a Distinguished Professor of Neurobiology at the University of California, San Diego, found that even in the absence of other proteins, piezo proteins could self-assemble into this tetramer complex, forming ion channels in artificial membranes known as lipid bilayers.

The second of the new studies involved experiments with the fruit fly Drosophila. Sung Eun Kim, first author of the study, genetically engineered a line of Drosophila that does not express the Drosophila piezo (dpiezo) gene. “We found that their larvae showed a severe loss of responsiveness to mechanical stimuli that would be expected to generate pain-related signals, though they responded normally to other kinds of stimuli such as heat and mild pressure,” she said. Kim is a graduate student who divides her time between the Patapoutian lab and the lab of Scripps Research Assistant Professor Boaz Cook, who was co-principal investigator of this study.

Kim also used genetic “knockdown” techniques in Drosophila to show that interrupting dpiezo expression in certain sensory neurons could reproduce this loss of sensitivity. Finally, when she artificially reinstated dpiezo expression in larvae that had been born without the gene, they displayed normal sensitivity to strong pressure. “It’s the first demonstration of a specific physiological function of a piezo family protein,” said Cook.

The Patapoutian lab now is now conducting detailed follow-up studies of piezo and other possible mechanotransduction proteins. “In the next several years, we’ll be trying to determine all the biological processes and diseases in which these pressure-sensing proteins play a role,” he said.

More information: “Piezos Are Pore-Forming Subunits of Mechanically Activated Channels,” Nature (2012).

Provided by The Scripps Research Institute

Source: medicalxpress.com