Cell-cell fusion is a fascinating cellular process in the development and physiology of multicellular organisms. It is critical for fertilization, skeletal muscle development, bone remodeling, immune response, placental formation, and stem cell-mediated tissue regeneration. Failure in cell-cell fusion leads to a plethora of defects such as infertility, congenital myopathy, osteopetrosis, immune deficiency, and preterm birth. Despite the diversity of cell types that undergo fusion, all cell-cell fusion events commence from the recognition and adhesion of two fusion partners and result in the merging of their plasma membranes and union of their cytoplasm. We are interested in uncovering the general mechanisms underlying cell-cell fusion and understanding its dysregulation in human diseases.
During skeletal muscle development, mononucleated myoblasts fuse to form multinucleated, contractile muscle fibers. Myoblast fusion is an evolutionarily conserved process that occurs from insects to mammals. We are using three different model organisms, Drosophila, zebrafish and mouse, to study mechanisms of myoblast fusion. We use Drosophila as a powerful genetic model to conduct unbiased genetic screens to identify new genes in myoblast fusion and characterize their functions. We use zebrafish as a vertebrate model to visualize the fusion process in the transparent zebrafish embryos. And we use mouse as a mammalian model to study the role for myoblast fusion in skeletal muscle regeneration and disease.
Podosomes and Invasive Protrusions
Our genetic and cell biological studies led to the discovery of an actin-enriched podosome-like structure (PLS) at the site of Drosophila myoblast fusion. The PLS is only generated in one of the two fusing cells (attacking cell). It projects finger-like invasive protrusions into the receiving cell, increasing membrane contact area and facilitating membrane juxtaposition and fusion. The one-sided PLS invasion makes the site of fusion an asymmetric structure, which we termed the asymmetric fusogenic synapse. Similar invasive protrusions mediate the induced fusion of cultured Drosophila non-muscle cells, and the fusion of mammalian myoblasts, osteoclasts and macrophages. Thus, invasive protrusions are used as a conserved and general mechanism to promote cell-cell fusion.
The actin cytoskeleton is critical for propelling invasive membrane protrusions. Genetic screens identified essential functions for the actin nucleation-promoting factors of the Arp2/3 complex in myoblast fusion. Our molecular, cell biological, biochemical, cryo-ET studies revealed novel mechanisms underlying the regulation of the actin cytoskeleton. We identified a regulatory protein that modulates the actin cytoskeletal dynamics by controlling the stability of the WASP-WIP complex, as well as a kinase that acts downstream of Rac to organize the actin filaments within the PLS. Our recent study of the large GTPase dynamin revealed the mechanism by which dynamin bundles actin to increase the mechanical strength of the actin network and promote invasive protrusions.
Using both Drosophila myoblast fusion and our reconstituted cell-fusion culture system, we discovered mechanosensory responses in the receiving fusion partner. Both the actin motor Myosin II and the membrane skeleton protein Spectrin exhibit mechanoresponsive accumulations at the fusogenic synapse in the receiving cell. Our genetic, cell biological, biophysical and mathematical modelling analyses demonstrate that the accumulated Myosin II increases actomyosin contraction in the receiving cell to resist the invasive forces, whereas the accumulated Spectrin network functions as a cellular sieve to constrict the invasive protrusions, thereby increasing the mechanical tension of the fusogenic synapse. The high mechanical tension, in turn, helps to overcome energy barriers for membrane apposition and drives cell membrane juxtaposition and fusion.