Signals from trunk paraxial mesoderm induce pronephros formation in chick intermediate mesoderm

We used Pax-2 mRNA expression and Lim 1/2 antibody staining as markers for the conversion of chick intermediate mesoderm (IM) to pronephric tissue and Lmx-1 mRNA expression as a marker for mesonephros. Pronephric markers were strongly expressed caudal to the fifth somite by stage 9. To determine whe ther the pronephros was induced by adjacent tissues and, if so, to identify the inducing tissues and the timing of induction, we microsurgically disse cted one side of chick embryos developing in culture and then incubated the m for up to 3 days. The undisturbed contralateral side served as a control. Most embryos cut parallel to the rostrocaudal axis between the trunk parax ial mesoderm and IM before stage 8 developed a pronephros on the control si de only. Embryos manipulated after stage 9 developed pronephric structures on both sides, but the caudal pronephric extension was attenuated on the cu t side. These results suggest that a medial signal is required for pronephr ic development and show that the signal is propagated in a rostral to cauda l sequence. In manipulated embryos cultured for 3 days in ovo, the mesoneph ros as well as the pronephros failed to develop on the experimental side. I n contrast, embryos cut between the notochord and the trunk paraxial mesode rm formed pronephric structures on both sides, regardless of the stage at w hich the operation was performed, indicating that the signal arises from th e paraxial mesoderm (PM) and not from axial mesoderm. This cut also served as a control for cuts between the PM and the LM and showed that signaling i tself was blocked in the fanner experiments, not the migration of pronephri c or mesonephric precursor cells from the primitive streak. Additional cont rol experiments ruled out the need for signals from lateral plate mesoderm, ectoderm, or endoderm. To determine whether the trunk paraxial mesoderm ca udal to the fifth somite maintains its inductive capacity in the absence of contact with more rostral tissue, embryos were transected Those transected below the prospective level of the fifth somite expressed Pax-2 in both th e rostral and the caudal isolates, whereas embryos transected rostral to th is level expressed Pax-2 in the caudal isolate only. Thus, a rostral signal is not required to establish the normal pattern of Pax-2 expression and pr onephros formation. To determine whether paraxial mesoderm is sufficient fo r pronephros induction, stage 7 or earlier chick lateral plate mesoderm was cocultured with caudal stage 8 or 9 quail somites in collagen gels. Pax-2 was expressed in chick tissues in 21 of 25 embryos. Isochronic transplantat ion of stage 4 or 5 quail node into caudal chick primitive streak resulted in the generation of ectopic somites. These somites induced ectopic proneph roi in lateral plate mesoderm, and the IRI that received signals from both native and ectopic somites formed enlarged pronephroi with increased Pax-2 expression. We conclude that signals from a localized region of the trunk p araxial mesoderm are both required and sufficient for the induction of the pronephros from the chick IM. Studies to identify the molecular nature of t he induction are in progress. (C) 2000 Academic Press.