Technology is opening up a hidden world of biology that we haven’t seen before!

All life consists of cells several orders of magnitude smaller than a grain of salt. Their seemingly simple structures hide complex molecular activities that enable them to perform life-support functions.

And researchers are starting to visualize this activity at a level of detail that they could not do before.

Biological structures can be visualized either at the level of the whole organism, or at the level of individual atoms and actions.

However, there was a gap in resolution between the smaller structures of the cell, such as the cytoskeleton, which maintains the shape of the cell, and its larger structures, such as the ribosomes, which make the proteins in the cells.

Similar to Google Maps, when scientists could see entire cities and individual houses, they didn’t have the tools to figure out how the houses fit into neighborhoods.

Observing these details at the neighborhood level is essential to understanding how the individual components work together in the environment of the cell. New tools are steadily closing this gap. The ongoing development of one technology, cryoelectron tomography or cryo-ET, may deepen how researchers study and understand how cells function in health and disease.

As Jeremy Berg, professor of biology and computing systems, assistant senior vice president of science strategy and planning at the University of Pittsburgh, former editor-in-chief of Science and researcher in the field, spent decades studying large, unimaginable protein structures. , and has made astonishing progress in developing tools that can define biological structures in detail. Just as it becomes easier to understand how complex systems work when you know what they look like, understanding how biological structures fit together in a cell is key to understanding how organisms work.

Brief history of microscopy.

In the 17th century, light microscopy first revealed the presence of cells. In the 20th century, electron microscopy provided more detail, revealing complex structures within cells, including organelles such as the endoplasmic reticulum, a complex network of membranes that plays a key role in protein synthesis and transport.

From the 1940s to the 1960s, biochemists worked to break down cells into their molecular components and learned to determine the three-dimensional structures of proteins and other large molecules at or near atomic resolution. This was done for the first time using crystallography and X-rays to image the structure of myoglobin, the protein that supplies oxygen to muscles.

Over the past decade, techniques based on nuclear magnetic resonance, which creates images based on how atoms interact in a magnetic field, and cryoelectron microscopy have increased the number of complex structures that scientists can image.

A cryoelectron microscope, or cryo-EM, uses a camera to determine how an electron beam is deflected as it passes through a sample to image structures at the molecular level.

Samples are quickly frozen to protect them from radiation damage. Detailed models of the structure of interest are created by taking multiple images of individual molecules and centering them on the 3D structure.

Cryo-ET has the same components as cryo-EM, but they use different techniques. Because most cells are too thick to be clearly imaged, the region of interest is first diluted into the cell using an ion beam.

The specimen is then tilted to take multiple pictures from different angles, similar to a CT scan of a body part. These images are then combined by a computer to create a 3D image of part of the cell. The resolution of this image is high enough for researchers or computer programs to identify individual components of various structures in the cell.

The researchers used this approach, for example, to show how proteins move and decompose inside an algae cell. Many of the steps that researchers had to follow manually to identify cellular structures have become automated, allowing scientists to identify new structures at a much faster rate.

For example, combining cryo-EM with AI software such as AlphaFold can facilitate image interpretation by predicting the structures of proteins that have not yet been characterized.

As imaging techniques and workflows improve, researchers will be able to address some of the key questions in cell biology with a variety of strategies.

The first step is to identify the cells and the locations within those cells that need to be explored. Another imaging technique called coherent light and electron microscopy, or CLEM, uses fluorescent markers to help identify regions of interest in living cells where processes of interest occur.

Comparison of genetic variation between cells can provide additional information. Scientists can look at cells that are unable to perform certain functions and see how this affects their structure. This approach could also help researchers study how cells interact with each other.

Cryo-ET is likely to remain a niche tool for a while. But further technological development and increased accessibility will allow the scientific community to explore the relationship between cellular structure and function at levels of detail previously unavailable.

Source: Science Alert