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Fig. 2 Alk8 Is Instructive in Myelopoiesis
(A–H) Anterior expression of pu.1 was increased in ventralized embryos injected with constitutively active alk8 (CA alk8) mRNA.
(A) Embryos injected with wild-type alk8 receptor mRNA (WT alk8) (10 ng/μL) are phenotypically wild-type.
(B–D) Embryos injected with CA alk8 mRNA (10 ng/μL) are ventralized and were separated into three classes of increasing phenotypic severity: class I embryos display reduced anterior structures and small eyes (B); class II embryos have reduced anterior structures and have no eyes (C); and class III embryos have virtually no anterior structures and are severely ventralized (D) (all embryos at 8 somites).
(E) Embryos injected with WT alk8 mRNA display normal expression of pu.1 at 14 hpf.
(F) Class I embryos display mildly increased expression of pu.1 in the anterior (31%; n = 26).
(G) Class II embryos display increased intensity and expansion of pu.1 expression in the anterior (85%; n = 46).
(H) Class III embryos display a restricted region of pu.1 expression of increased intensity (94%; n = 18).
(I–K) Increased expression of pu.1 is specific to the delivery of CA alk8 mRNA in mosaic embryos.
(I) Embryos injected with lacZ mRNA only (50 ng/μL) into a single cell at the 1/16–1/32 cell stage display wild-type expression of pu.1 in lacZ-stained regions (n = 38 unilaterally lacZ-stained embryos displayed normal expression of pu.1). Embryo displayed at 8 somites. ALPM and PLPM are indicated by brackets.
(J and K) Embryos coinjected with lacZ mRNA (50 ng/μL) and CA alk8 mRNA (40 ng/μL) display specifically increased pu.1 expression in regions of lacZ staining at 10 somites (J) and 8 somites (K) (n = 18/26 CA alk8, lacZ mosaics that stained unilaterally displayed coincidentally increased pu.1 expression). Arrowheads indicate increased pu.1; boxed sections in (J) and (K) are expanded to show close association of lacZ- and pu.1-positive cells. Embryos in (E)–(K) are flatmounted and viewed dorsally; (E)–(H) are dorsal views displaying ALPM at 12 somites; (I) and (K) are dorsal views of the whole embryo at 8 somites; and (J) is a dorsal view of the whole embryo at 10 somites.