Churchill and Sip1a repress fibroblast growth factor signaling during zebrafish somitogenesis
Article first published online: 23 DEC 2009
Copyright © 2009 Wiley-Liss, Inc.
Volume 239, Issue 2, pages 548–558, February 2010
How to Cite
Kok, F. O., Shepherd, I. T. and Sirotkin, H. I. (2010), Churchill and Sip1a repress fibroblast growth factor signaling during zebrafish somitogenesis. Dev. Dyn., 239: 548–558. doi: 10.1002/dvdy.22201
- Issue published online: 22 JAN 2010
- Article first published online: 23 DEC 2009
- Manuscript Accepted: 20 NOV 2009
- NIH. Grant Number: RO1HD043998
Additional Supporting Information may be found in the online version of this article.
|DVDY_22201_sm_SuppFig1.tif||256K||Supp. Fig. S1. sip1a splice morpholino is effective in eliminating wild-type sip1a mRNA. Reverse transcriptase-polymerase chain reaction (RT-PCR) of mRNA from staged sip1a splice MO injected embryos with sip1a specific primers reveals that the smaller misspliced product is detected by dome stage (4.3 hours postfertilization [hpf]). By 75% epiboly (8 hpf), no wild-type mRNA is detected.|
|DVDY_22201_sm_SuppFig2.tif||6675K||Supp. Fig. S2. Comparison of zebrafish Sip1a and Sip1b proteins. Genedoc software is used to align the predicted protein sequences. Predicted domains of Sip1a and Sip1b is lined with colored bars: C2H2 type zinc fingers present in both Sip1a and Sip1b (red), C2H2 type zinc finger region that is absent from Sip1a and Sip1b short forms (blue). Identical and similar amino acids conserved among all proteins are shown in black and dark gray boxes, respectively. Lighter shades of gray or no shading represent low levels of amino acid conservation and the lack of conservation, respectively.|
|DVDY_22201_sm_SuppFig3.tif||5846K||Supp. Fig. S3. sip1a splice variant targeting morpholinos efficiently eliminate short and long forms. A: Diagram of the alternative splicing within the 3′ region of the sip1a pre-mRNA. Alternative splicing of exon 8 eliminates one zinc finger (dark blue box), which is present in the longer form. Sip1a MO3 (orange bar) targeted to alternative splice site in exon 8 (denoted by black arrow) blocks the alternating splicing event and eliminates production of the short form. Sip1a MO2 (pink bar) blocks the pre-mRNA splicing event needed to generate the long form by targeting the splice site at the 3′ end of exon 8. B,C: Reverse transcriptase-polymerase chain reaction (RT-PCR) of mRNA from staged sip1a splice MO2 and MO3 injected embryos with sip1a full length specific primers (B, green arrows in A) and sip1a short form specific primers (C, orange arrows in A ). B,C: sip1 splice MO2 efficiently altered splicing so that the full length message is eliminated and only the shorter form was produced (B) while sip1 splice MO3 blocked production of the shorter form (C).|
|DVDY_22201_sm_SuppFig4.tif||18565K||Supp. Fig. S4. Pace of the “molecular clock” is not significantly altered in ChCh-compromised embryos. Living wild-type and ChCh-compromised embryos beginning at the seven-somite stage. A–J: All views are dorsal, anterior to the top. Duration of somite formation in both control (A–E) and ChCh-compromised embryos (F–J) is approximately 45 min at 23°C. Black arrows denote already formed somite boundaries and white arrows denote newly forming segmentation furrow.|
|DVDY_22201_sm_SuppFig5.tif||494K||Supp. Fig. S5. Somite malformation in ChCh and Sip1a compromised embryos can be partially rescued by fibroblast growth factor (FGF) antagonist Spry4. Bar graph representation of spry4 rescue assay in chch and sip1a ATG morphants. Injection of spry4 sense RNA into chch and sip1a ATG morphants (blue bar) reduced the penetrance of the somite phenotype with respect to their lacZ injected siblings (gray bar).|
Please note: Wiley Blackwell is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.