Dorsoventral Patterning of the Zebrafish Embryo
95
6.8 Chordino Acts via Bmp2b and Bmp7
To test whether Chordino functions via an inhibition of ventralizing
proteins of the Bmp family, as suggested by' biochemical analyses
carried out in amphibia (Piccolo et al. 1996), chordino-bmp2b (Hammerschmidt et al. 1996b) and chordino-bmp7 double mutants were
generated. Such double mutants appear dorsalized much like bmp2b and
bmp7 single mutants, indicating that Bmp2b and Bmp7 are epistatic to
Chordino. This suggests that Chordino is not an actively dorsalizing
agent, but does indeed act as an inhibitor of Bmp2b and Bmp7. Clearly,
the absence of such an inhibitor has no further effects when its substrate
is missing anyway.
The analysis of the bmp2b, bmp4, and bmp7 expression patterns in
chordino, bmp2b, and bmp7 mutant embryos suggests that Chordino is
required for the establishment of the putative morphogenetic Bmp2/417
gradient. bmp2 and bmp7 are initially expressed in a uniform and broad
fashion, sparing only the region of the presumptive Spemann organizer
where chordino is expressed. During further development, this pattern is
transformed into a graded pattern (see above and Fig. 3). In chordino
mutants, however, the initial broad and uniform expression pattern is
maintained (see Fig. 3E for bmp4), indicating that Chordino is required
for the progressive clearing of bmp21417 expression from dorsal to
ventral. This clearing is achieved indirectly via the inhibition of
Bmp2/417 activity. Bmps are positive autoregulators of their own expryssion, as indicated by the progressive loss of bmp21417 transcripts in
swr bmp2b and snh bmp7 double mutant embryos (see Fig. 3F for bmp4
in swr). Thus, by binding and inhibiting Bmp proteins, Chordino attenuates this positive autoregulation, leading to decreased bmp21417 transcription. Because of the positive autoregulation, the levels of bmp21417
transcripts can be regarded as a readout of Bmp2/417 activity. Therefore,
the observed gradient of bmp21417 transcripts could be a direct reflection of the proposed gradient of Bmp2/417 activity.
95
6.8 Chordino Acts via Bmp2b and Bmp7
To test whether Chordino functions via an inhibition of ventralizing
proteins of the Bmp family, as suggested by' biochemical analyses
carried out in amphibia (Piccolo et al. 1996), chordino-bmp2b (Hammerschmidt et al. 1996b) and chordino-bmp7 double mutants were
generated. Such double mutants appear dorsalized much like bmp2b and
bmp7 single mutants, indicating that Bmp2b and Bmp7 are epistatic to
Chordino. This suggests that Chordino is not an actively dorsalizing
agent, but does indeed act as an inhibitor of Bmp2b and Bmp7. Clearly,
the absence of such an inhibitor has no further effects when its substrate
is missing anyway.
The analysis of the bmp2b, bmp4, and bmp7 expression patterns in
chordino, bmp2b, and bmp7 mutant embryos suggests that Chordino is
required for the establishment of the putative morphogenetic Bmp2/417
gradient. bmp2 and bmp7 are initially expressed in a uniform and broad
fashion, sparing only the region of the presumptive Spemann organizer
where chordino is expressed. During further development, this pattern is
transformed into a graded pattern (see above and Fig. 3). In chordino
mutants, however, the initial broad and uniform expression pattern is
maintained (see Fig. 3E for bmp4), indicating that Chordino is required
for the progressive clearing of bmp21417 expression from dorsal to
ventral. This clearing is achieved indirectly via the inhibition of
Bmp2/417 activity. Bmps are positive autoregulators of their own expryssion, as indicated by the progressive loss of bmp21417 transcripts in
swr bmp2b and snh bmp7 double mutant embryos (see Fig. 3F for bmp4
in swr). Thus, by binding and inhibiting Bmp proteins, Chordino attenuates this positive autoregulation, leading to decreased bmp21417 transcription. Because of the positive autoregulation, the levels of bmp21417
transcripts can be regarded as a readout of Bmp2/417 activity. Therefore,
the observed gradient of bmp21417 transcripts could be a direct reflection of the proposed gradient of Bmp2/417 activity.