The Influence of Drosophila Spire and Myosin V During Mid-Oogenesis Is Independent of Their Direct Interaction.

Abstract

Cooperativity between cytoskeletal proteins is crucial for spatiotemporal coordination in biological processes, like oogenesis. In mammalian and Drosophila oogenesis, proper assembly and function of actin networks require coordination between actin assembly factors Spire and formins, as well as actin-associated proteins like myosins and Rab GTPases. Here, we investigate the interaction between Spire and Myosin V (MyoV) in Drosophila oogenesis. We combine in vitro biochemical assays with immunofluorescence and genetics to probe the interaction and its impact on polarity establishment and the actin mesh that fills the oocyte during mid-oogenesis. Expressed Spire and MyoV constructs colocalize in punctae during mid oogenesis, with considerable enrichment near the oocyte cortex. Purified constructs interact directly in vitro, and we find that Spire can weakly activate MyoV ATPase activity. Cytoplasmic flows, critical for polarity establishment, are faster and more coordinated in the absence of MyoV, although not to the extent of fast streaming. This intermediate streaming has not been observed before. Interestingly, this MyoV-dependent change in ooplasm dynamics is sensitive to Spire levels. Despite this interplay, the actin mesh and polarity establishment appear normal when binding mutants of Spire and MyoV are expressed in the Drosophila germline. These findings suggest that direct interaction between Spire and MyoV is not essential for their primary roles at this stage of development.