Cryo-Em of Z-Discs Isolated from Honeybee Flight Muscle

Abstract

Muscle Z-discs were originally thought to have the purely mechanical function of transmitting contractile force along myofibrils, connecting thin filaments in adjacent sarcomeres through Z-bridges composed principally of α-actinin. However, the Z-disc is now known to be considerably more complex, with ∼40 different proteins, some of which are transient. Other Z-disc functions identified include stress sensing into signalling pathways involving muscle growth, remodelling and degradation. Z-discs vary widely in different muscles, including differences in thickness and symmetry. Electron microscope transverse sections of vertebrate muscle show Z-discs with tetragonal symmetry, but with two appearances, called small-square and basketweave, with the basketweave lattice 10% smaller. The relative proportions of these states can be modulated by several factors affecting the state of the muscle, but the significance of this transition is not known. Invertebrate Z-discs have hexagonal symmetry and have been most studied in insect flight muscle. The structure of the Z-disc is known only in outline, to ∼7 nm resolution, whereas ∼2 nm resolution is required to recognise protein shapes and accurately dock crystal structures. Isolated Z-discs are potentially useful for EM studies because they are naturally thin and do not have to be cut into thin sections, which causes damage and loss of resolution. Reports of methods to prepare and purify Z-discs date back 50 years, but such preparations have not been examined by cryo-EM or tomography, which may improve resolution. Isolated Z-discs may also be valuable for composition studies, perhaps including genetic modification. We have obtained preliminary cryo-EM data from honeybee flight muscle Z-discs prepared by high salt treatment of myofibrils.