Extended Data Figure 10: Functions of H. magnipapillata Ndr and Pitx in bud initiation.
From: Nodal signalling determines biradial asymmetry in Hydra
Gene-specific siRNAs were co-transfected with a GFP-specific siRNA, and bud formation from the side of transfection (GFP-depleted side) was monitored. In the mock-transfected hydra, GFP fluorescence remained intact after electroporation. As Ndr and Pitx are expressed in different germ layers (that is, in the ectoderm and endoderm, respectively), we used transgenic Hydra strains in which either the ectoderm or endoderm is labelled with GFP or red fluorescent protein (RFP), respectively35. Bud formation from the GFP-depleted side was significantly suppressed by co-transfection with an Ndr-specific siRNA (a) or a Pitx-specific siRNA (b). Note that the RFP fluorescence was not affected, indicating that the decrease in the GFP fluorescence was not due to tissue damage caused by electroporation. The siRNAs were electroporated more effectively into the ectoderm than the endoderm, because the GFP-positive region of the endoderm remained more expanded at the non-transfected side (a, b). Bud formation from the side of the GFP-positive endoderm was not affected in Pitx-specific siRNA transfectants (c, d), indicating that localized expression and function of Pitx at the budding region is required. The data are presented as the mean ± s.e.m. of three independent experiments.
