Evaluation of developmental phenotypes produced by morpholino antisense targeting of a sea urchin Runx gene.
BACKGROUND
Runx transcription factors are important regulators of metazoan development. The sea urchin Runx gene SpRunt was previously identified as a trans-activator of the CyIIIa actin gene, a differentiation marker of larval aboral ectoderm. Here we extend the functional analysis of SpRunt, using morpholino antisense oligonucleotides (morpholinos) to interfere with SpRunt expression in the embryo.


RESULTS
The developmental effects of four different SpRunt-specific morpholinos were evaluated. The two morpholinos most effective at knocking down SpRunt produce an identical mitotic catastrophe phenotype at late cleavage stage that is an artifact of coincidental mis-targeting to histone mRNA, providing a cautionary example of the insufficiency of two different morpholinos as a control for specificity. The other two morpholinos produce gastrula stage proliferation and differentiation defects that are rescued by exogenous SpRunt mRNA. The expression of 22 genes involved in cell proliferation and differentiation was analyzed in the latter embryos by quantitative polymerase chain reaction. Knockdown of SpRunt was found to perturb the expression of differentiation markers in all of the major tissue territories as well as the expression of cell cycle control genes, including cyclin B and cyclin D.


CONCLUSIONS
SpRunt is essential for embryonic development, and is required globally to coordinate cell proliferation and differentiation.
Coffman
James A
JA
Stowers Institute for Medical Research, 1000 E, 50th Street, Kansas City, MO 64110, USA. jac@stowers-institute.org
Dickey-Sims
Carrie
C
Haug
Jeffrey S
JS
McCarthy
John J
JJ
Robertson
Anthony J
AJ
D000818:Animals; D004625:Embryo, Nonmammalian / Q000187:drug effects* / Q000196:embryology; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D018390:Gene Targeting; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012617:Sea Urchins / Q000196:embryology* / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; C098677:SpRunt-1 protein, Strongylocentrotus purpuratus; D014157:Transcription Factors