Multiplexin Promotes Heart but Not Aorta Morphogenesis by Polarized Enhancement of Slit/Robo Activity at the Heart Lumen

Abstract

The Drosophila heart tube represents a structure that similarly to vertebrates' primary heart tube exhibits a large lumen; the mechanisms promoting heart tube morphology in both Drosophila and vertebrates are poorly understood. We identified Multiplexin (Mp), the Drosophila orthologue of mammalian Collagen-XV/XVIII, and the only structural heart-specific protein described so far in Drosophila, as necessary and sufficient for shaping the heart tube lumen, but not that of the aorta. Mp is expressed specifically at the stage of heart tube closure, in a polarized fashion, uniquely along the cardioblasts luminal membrane, and its absence results in an extremely small heart tube lumen. Importantly, Mp forms a protein complex with Slit, and interacts genetically with both slit and robo in the formation of the heart tube. Overexpression of Mp in cardioblasts promotes a large heart lumen in a Slit-dependent manner. Moreover, Mp alters Slit distribution, and promotes the formation of multiple Slit endocytic vesicles, similarly to the effect of overexpression of Robo in these cells. Our data are consistent with Mp-dependent enhancement of Slit/Robo activity and signaling, presumably by affecting Slit protein stabilization, specifically at the lumen side of the heart tube. This activity results with a Slit-dependent, local reduction of F-actin levels at the heart luminal membrane, necessary for forming the large heart tube lumen. Consequently, lack of Mp results in decreased diastolic capacity, leading to reduced heart contractility, as measured in live fly hearts. In summary, these findings show that the polarized localization of Mp controls the direction, timing, and presumably the extent of Slit/Robo activity and signaling at the luminal membrane of the heart cardioblasts. This regulation is essential for the morphogenetic changes that sculpt the heart tube in Drosophila, and possibly in forming the vertebrates primary heart tube.

Iterative inhibition of commissural growth cone exploration, not post-crossing barrier, ensures forward midline navigation through SlitC-PlxnA1 signaling

“…Morphogenesis of the drosophila heart tube lumen was found to rely on polarized localization of the collagen XV/XVIII orthologue Multi-Plexin forming a macro-complex with Slit (Harpaz et al, 2013). In the context of midline crossing, loss of the heparan sulfate carrier (Heparan sulfate proteoglycan) Syndecan results in modification of Slit distribution and activity in drosophila (Johnson et al, 2004).…”

On the role of mechanics in driving mesenchymal-to-epithelial transitions

“…Processes involved in MET and heart tube formation in Drosophila parallel those guiding vertebrate heart formation, including the dependence of MET, but not cell identity on extracellular matrix [90–93], and the role of tension within the embryo to position precursors and enable fusion at the midline [94]. Furthermore, amnioserosa cells, which first appear to engage precursors with E-cadherin and septate junctions must disengage before cardiac cells can fully establish polarized lumenal-domains and fuse [95].…”

Slit cleavage is essential for producing an active, stable, non-diffusible short-range signal that guides muscle migration

“…Mechanistically, Slit cleavage transforms the protein into an active, stable polypeptide, which is tethered to the tendon cell, further protecting it from degradation and restricting its distribution. Whereas the nature of the protease that cleaves Slit has yet to be elucidated, the tendon-specific surface proteoglycans syndecan (Chanana et al, 2009;Coleman et al, 2010) and multiplexin (Harpaz et al, 2013;Meyer and Moussian, 2009;Momota et al, 2011Momota et al, , 2013, shown previously to bind Slit in the CNS or in the heart, respectively, might tether the cleaved Slit-N to the tendon cell membrane. This novel regulatory mechanism controlling Slit distribution may also operate in other tissues, such as the heart and blood vessels, where Slit appears to be active between neighboring cells.…”