Fig 6. Model: Dfos increases actin assembly and cross-linking through the tetraspanin TM4SF and the filamin Cher to counter surrounding tissue resistance.

We propose a speculative model for how Dfos tunes the cortical actin properties of Drosophila embryonic macrophages to aid their infiltration against the resistance of the surrounding gb tissue. We have shown that Dfos leads to an increase of the tetraspanin TM4SF and the filamin Cher. Filamins cross-link actin and have been shown to bind to RhoGEFs; Tetraspanins bind to Integrins, Rho GTPases, and Filamins in other systems (see S6 Fig). Thus, we hypothesize that in Drosophila macrophages, TM4SF and the filamin Cher could form a network at the cell surface of Integrin, actin, and upstream signaling molecules, recruiting Rho GEFs and leading to the activation of Rho1 GTPase and the actin polymerizing Formin Dia. Dia activation could occur through direct binding to active Rho1 and through direct interaction with TM4SF or Cher. Validation in Drosophila of all the protein interactions we propose awaits biochemical analysis. Through this pathway, a more cross-linked and dense F-actin network would form, aiding the macrophage in moving its cell body into the ecto–meso interface. The presence of Lamin around the nuclear membrane would not normally affect this process since the dense cross-linked cortical actin network would help macrophages withstand the load of the surrounding tissues. However, in the DfosDN-expressing macrophages, the loss of Cher and TM4SF would lead to reduced cross-linked actin levels at the cell cortex, making the stiffness of the nucleus the rate limiting step for macrophage infiltration of the gb tissue. Cher, Cheerio; ecto, ectoderm; gb, germband; mac, macrophage; meso, mesoderm.