DEV022830 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. osr1 expression in endodermal organs at 48 hpf. (A) Expression of osr1 mRNA in whole mount at 48 hpf, dorsal view. Lines represent plane of section in B, C. (B) Histological section just anterior to the fin bud shows osr1 expression in the forming liver (li). Arrowhead indicates the forming glomerulus, which is negative for osr1. (C) Histological section at the level of the fin shows osr1 expression in the gut (g).

• Supplemental Figure S2 -

  Fig. S2. osr1 loss of function does not cause general A-P patterning defects in the pronephros. Expression of trpM7 in control (A) and osr1 morphants (B) is restricted to the late proximal pronephric nephron segment. ret1 expression in control (C) and osr1 morphants (D) is unchanged in the most distal pronephric nephron segment.

• Supplemental Figure S3 -

  Fig. S3. osr1 loss of function does not affect mesoderm survival. Acridine Orange staining for apoptotic cells in control (A) and osr1 morphant (B) 18-somite embryos. Arrowheads indicate area affected in osr1 morphants. TUNEL staining of control (C) and osr1 morphant (D) 18-somite embryos show unchanged patterns of apoptosis. Arrowheads indicate the area anterior intermediate mesoderm most strongly affected in osr1 morphants.

• Supplemental Figure S4 -

  Fig. S4. Expanded vascular differentiation is not due to an altered rate of development. (A) scl (blue) and myoD (red) expression in wild-type embryos at the 12-somite stage. (B) osr1 knockdown results in expansion of scl-positive tissue, most prominently in anterior PLM. myoD is used as an internal staging control and demonstrates both control and osr1 morphants are at the 12 somite stage. (C) scl (blue) and pax2a (red) expression in wild-type embryos from the same clutch of embryos as A and B. (D) osr1 knockdown results in expansion of scl-positive tissue, most prominently in anterior PLM, and loss of pax2a-expressing cells. (C, D) are lower magnification views of A,B in Fig. 5 respectively.

• Supplemental Figure S5 -

  Fig. S5. Intermediate mesoderm patterning in pax2/8 and scl morphants. Expression of scl (blue) and myoD (red) in control (A), and pax2a/pax8 double morphant (B) demonstrates that loss of Pax gene expression does not cause scl upregulation. Arrowheads indicate the area of strongest upregulation of scl seen in osr1 morphants. Expression of pax2a in control (C), scl morphant (D) similarly shows no large expansion of pax2a expression. Expression of pax2a in osr1 morphant (E) and osr1/scl double morphant (F) shows no rescue of pax2a expression. Arrowheads in C-F indicate the proximal nephron segment lost in osr1 morphants. The data indicate that Pax genes and scl are not involved in a mutual repression mechanism.