How do cells build biological machines?
Cells are expert organisers, manufacturers, transporters and recyclers of cellular content. At the heart of all these processes is biochemistry - the chemistry of life!
In the Mali laboratory, we are interested in understanding the manufacturing process of macromolecular machines - specifically the biosynthesis of axonemal dynein motors that power the movement of eukaryotic cilia which are vital to human health.
We integrate biochemistry, cell biology and structural techniques to mechanistically understand the assembly of ciliary dynein motors.
Virtually all proteins inside a cell work with other proteins as part of large complexes to exert their biological functions. In the case of ciliary dynein motors, a whole host of cellular assembly factors work together to shephard dynein assembly. To understand their molecular mechanisms in detail, we biochemically purify dynein assembly factor complexes directly from cells and study their functions in vitro.
Image: Cellular contents get separated into distinct fractions after cells are spun at very high speeds in an ultracentrifuge. This is an essential first step in isolating pure protein complexes for further study.
Truly understanding the function of protein complexes requires a detailed look not only in vitro but also at how they behave inside a cell i.e. in vivo. In the Mali laboratory, we focus on studying the protein factors involved in building dynein motors using a diverse set of ciliated cell models including protists (Tetrahymena), green algae (Chlamydomonas) and ciliated lung and brain cells from humans and mice. Such studies provide mechanistic insights into ciliary dynein assembly within a cellular context.
Image: Axonemal dynein complexes labeled in green light up the cilia that cover the entire body of a Tetrahymena cell. The cytoplasm is labeled with a red dye and the blue cell nuclei are marked by DAPI stain.
Structure informs function in biology. Directly observing a protein complex often provides novel insights into the way it functions.
In the Mali laboratory, we use a cutting edge structural biology technique called cryo-electron microscopy along with protein structure predictions to generate 3D models of protein complexes that build dynein motors . Models are then interpreted along with functional studies which inform on their molecular mechanisms.
Image: Single molecules of a major ciliary dynein motor from different angles and orientations are shown. Images of molecules were acquired using powerful electron microscopes. After extensive image processing, a composite of many thousands of such images was used to reconstruct a first 3D structure of this motor which provided many novel functional insights.