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Topic Name: Researchers Unveiled the Activity of a Specific Family of Nanometer-Sized Molecular Motors

Category: Biomedical

Research persons: . Michael Ostap, Ph.D., Henry Shuman, PhD

Location: University of Pennsylvania School of Medicine, United States

Details

Researchers at the University of Pennsylvania School of Medicine discovered that the activity of a specific family of nanometer-sized molecular motors called myosin-I is regulated by force. The motor puts tension on cellular springs that allow vibrations to be detected within the body. This finely tuned regulation has important implications for understanding a wide variety of basic cellular processes, including hearing and balance and glucose uptake in response to insulin.

"This is the first demonstration that myosin-I shows such dramatic sensitivity to tension," says senior author E. Michael Ostap, Ph.D., Associate Director, Pennsylvania Muscle Institute and Associate Professor of Physiology. "It is surprising that a molecular motor can sense such small changes in force."

Myosin-I is a biological motor that uses the chemical energy made by cells to ferry proteins within cells and to generate force, powering the movement of molecular cargos in nearly all cells.

In two specific cases, myosin I puts tension on the specialized spring-like structures in human ears that enable hearing and maintenance of balance, and also has a role in delivering the proteins that pump glucose into cells in response to insulin. "However, why a tension-sensing molecular motor is needed for this function is unknown," says Ostap.

In collaboration with Henry Shuman, PhD, Associate Professor of Physiology, the research team used optical tweezers -- a combination focused laser beam and microscope, of sorts -- to measure incredibly small forces and movements (on the piconewton and nanometer level) to discover that myosin I motors are regulated by force. The motors pull on their cellular cargos until a certain tension is attained, after which they stop moving, but will hold the tension. If something happens in the cell to decrease this tension, the motor will restart its activity and will restore the lost tension.

Myosins use the energy from ATP to generate force and motion. Humans have 40 myosin genes that sort into 12 myosin families. Members of the myosin family have been found in every type of cell researchers have examined. The Ostap lab is investigating the biochemical properties of several members of the myosin family to better understand movement in cells, which is important in development, wound healing, the immune response, and the spread of cancer, among other functions. These new findings shed light on the role of myosin I in cells, supporting the notion that this molecular motor is more important in generating and sustaining tension rather than transporting protein cargo.

The research team will now apply these results to better understand how cells use these tension sensors to carry out their physiological functions.

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Note for Myosin
Myosins are a large family of motor proteins found in eukaryotic tissues. They are responsible for actin-based motility. The wide variety of myosin genes found throughout the eukaryotic phyla were named according to different schemes as they were discovered. The nomenclature can therefore be somewhat confusing when attempting to compare the functions of myosin proteins within and between organisms.

Skeletal muscle myosin, the most conspicuous of the myosin superfamily due to its abundance in muscle fibers, was the first to be discovered. This protein makes up part of the sarcomere and forms macromolecular filaments composed of multiple myosin subunits. Similar filament-forming myosin proteins were found in cardiac muscle, smooth muscle, and non-muscle cells. However, beginning in the 1970s researchers began to discover new myosin genes in simple eukaryotes encoding proteins that acted as monomers and were therefore entitled Class I myosins. These new myosins were collectively termed "unconventional myosins" and have been found in many tissues other than muscle. These new superfamily members have been grouped according to phylogenetic relationships derived from a comparison of the amino acid sequences of their head domains, with each class being assigned a Roman numeral. The unconventional myosins also have divergent tail domains, suggesting unique functions.

Analysis of the amino acid sequences of different myosins shows great variability among the tail domains but strong conservation of head domain sequences. Presumably this is so the myosins may interact, via their tails, with a large number of different cargoes, while the goal in each case - to move along actin filaments - remains the same and therefore requires the same machinery in the motor. For example, the human genome contains over 40 different myosin genes.

These differences in shape also determine the speed at which myosins can move along actin filaments. The hydrolysis of ATP and the subsequent release of the phosphate group causes the "power stroke," in which the "lever arm" or "neck" region of the heavy chain is dragged forward. Since the power stroke always moves the lever arm by the same angle, the length of the lever arm determines how fast the cargo will move. A longer lever arm will cause the cargo to traverse a greater distance even though the lever arm undergoes the same angular displacement - just as a person with longer legs can move farther with each individual step. Myosin V, for example, has a much longer neck region than myosin II, and therefore moves 30-40 nanometers with each stroke as opposed to only 5-10.

Note for Optical Tweezer
An optical tweezer is a scientific instrument that uses a focused laser beam to provide an attractive or repulsive force, depending on the index mismatch (typically on the order of piconewtons) to physically hold and move microscopic dielectric objects. Optical tweezers have been particularly successful in studying a variety of biological systems in recent years.

Optical tweezers are capable of manipulating nanometer and micrometer-sized dielectric particles by exerting extremely small forces via a highly focused laser beam. The beam is typically focused by sending it through a microscope objective. The narrowest point of the focused beam, known as the beam waist, contains a very strong electric field gradient. It turns out that dielectric particles are attracted along the gradient to the region of strongest electric field, which is the center of the beam. The laser light also tends to apply a force on particles in the beam along the direction of beam propagation. It is easy to understand why if you imagine light to be a group of tiny particles, each impinging on the tiny dielectric particle in its path.

Optical traps are very sensitive instruments and are capable of the manipulation and detection of sub-nanometer displacements for sub-micrometre dielectric particles. For this reason, they are often used to manipulate and study single molecules by interacting with a bead that has been attached to that molecule. DNA and the proteins and enzymes that interact with it are commonly studied in this way.

For quantitative scientific measurements, most optical traps are operated in such a way that the dielectric particle rarely moves far from the trap center. The reason for this is that the force applied to the particle is linear with respect to its displacement from the center of the trap as long as the displacement is small. In this way, an optical trap can be compared to a simple spring, which follows Hooke's law.

This research was supported by the National Institute of General Medical Sciences and the National Institute of Arthritis and Musculoskeletal and Skin Diseases.