UC San Diego

ATP-binding cassette (ABC) transporters are large membrane proteins found in all organisms. They use ATP to power transport of diverse substrates across membranes, and the ABC-B, -C, and -G families make up the multidrug resistance (MDR) transporters. Here, I survey the expression of MDR transporters during development of the purple sea urchin, Strongylocentrotus purpuratus, and identify and characterize one transporter, Sp-ABCC5a (C5a) that has a role in developmental signaling required for hindgut morphogenesis.

From the unfertilized egg through the prism stage (~58 hours post-fertilization, hpf), I detected transcripts of 20 MDR transporters with predicted roles in cell signaling, lysosomal and mitochondrial homeostasis, potassium channel regulation, pigmentation, and xenobiotic efflux. The protective transporter ABCB1a is ubiquitously expressed throughout embryogenesis in all cells, and the protein localizes to apical membranes at the interface between the embryo and external environment. In contrast, expression of C5a, a previously uncharacterized transporter, is first detected at hatching and peaks after gastrulation, and the protein localizes to basolateral membranes. I propose that profiling spatiotemporal expression and subcellular protein localization may identify transporters with roles in protection, homeostasis, and signaling.

As expected, efflux activity of C5a does not support a role in protection; C5a is not a broad-spectrum chemical effluxor, and it discriminates between structurally similar compounds. C5a expression is restricted to aboral non-skeletogenic mesoderm (NSM) cells (pigment cells) and is dependent on delta/notch signaling from the skeletogenic mesoderm (SM) as well as the aboral NSM regulatory gene, glial cells missing. During gastrulation, C5a protein traffics to the plasma membrane of pigment cells, while these cells migrate away from the archenteron to embed in the aboral ectoderm.

Knockdown of C5a leads to embryos with differentiated pigment cells but abnormal protrusion of the hindgut, which is seen in ~90% of embryos by the late prism stage (~eight hours after C5a protein expression peaks). The ABCC5 substrate cAMP rescues the prolapse in a dose-dependent manner, and in control embryos causes hyper-invagination. The cAMP-producing enzyme soluble adenylyl cyclase (sAC) is expressed in tandem with C5a, with peak expression in pigment cells just after gastrulation. Blocking sAC activity with the specific inhibitor KH7 impairs gastrulation. Together these data suggest that C5a-mediated efflux of sAC-derived cAMP from pigment cells controls late invagination of the hindgut. This study elucidates a novel role for an ABCC/MRP transporter in mesoderm-endoderm signaling during embryogenesis.