Zasp PDZ Domain Proteins Cooperate in Z-Disc Formation and Myofibril Assembly

Abstract

Zasp52 is a multi-domain protein, composed of an N-terminal PDZ domain, ZM motif, and four LIM domains, and is found predominantly in Drosophila muscle. It is part of the PDZ-LIM domain protein family, which comprises Enigma, ENH, ZASP, Alp, CLP36, RIL, and Mystique in vertebrates. In Drosophila embryos, Zasp52 was shown to co-localize with α-actinin at Z-discs and with integrins at myotendinous junctions. Mutations in Zasp52 cause first instar larval lethality with defects in Z-disc assembly and maintenance as well as muscle attachment. During Drosophila indirect flight muscle (IFM) development in the pupa, Zasp52 is present at very early stages of myofibril assembly. It localizes with α-actinin in rudimentary Z-bodies along the assembling myofibril, before other muscle proteins show any periodicity. RNA interference against the last exon encoding the most C-terminal LIM domain results in the depletion of all high molecular weight isoforms. This gives rise to viable but flightless adult animals. IFM sarcomeres show thin and interrupted Z-discs as well as distorted H-zones from early stages of myofibril assembly onwards. These findings suggest that Zasp52 is required for the establishment of normal Z-discs in the IFM and subsequent sarcomere stability after onset of contractility. A closely related PDZ domain containing protein, Zasp66, is also required for Z-disc formation and stability. Simultaneous knock down of Zasp52 and Zasp66 lead to more severe, synergistic myofibril defects, demonstrating that both proteins act together during Z-disc formation, likely because both biochemically bind α-actinin.