Presented at the 6th International Congress
on Cell Biology and the 36th American Society for Cell Biology Annual Meeting,
December 7-11, 1996, San Francisco, California, Abstract #3640, Molecular
Biology of the Cell, Volume 7, Page 626a, December, 1996.
"A Specific Synthetic RNA Promotes Cardiac Myofibrillogenesis in
the Mexican Axolotl",
LF Lemanski, M Nakatsugawa, R Bhatia, N Erginel-Unaltuna, and DK
Dube.
Department of Anatomy and Cell Biology, State University of New York
Health Science Center, Syracuse, New York 13210.
Ambystoma mexicanum is an intriguing animal model for studying heart
development because it carries a mutation in gene c. Hearts of homozygous
recessive (c/c) mutant embryos do not contain organized myofibrils and
fail to beat. However, the defect can be corrected by organ-culturing in
the presence of RNA from anterior endoderm or RNA from endoderm-conditioned
medium.We constructed a cDNA library from total conditioned medium RNA
in a pcDNAII expression vector. We screened the cDNA library by an organ
culture bioassay and isolated a single clone (Cl. #4), the synthetic RNA
from which corrects the heart defect by promoting myofibrillogenesis. The
insert size of the active clone is 166 nt in length with unique nucleotide
sequence. The anti-sense RNA from Clone #4 using SP6 RNA polymerase failed
to rescue mutant hearts. The ability of a small RNA to correct the mutant
heart defect suggests that this RNA probably does not act as an mRNA, but
rather as a ligand RNA. FITC-labelled synthetic RNA can also correct the
heart defect, and its distribution indicates that the RNA is taken up by
the rescued mutant heart cells. RT-PCR analyses with RNA from different
stages indicate that the onset of the expression of this RNA is about stage
10-12 (gastrula). The expression reaches a maximum at stage 15 (early neurula)
and then declines gradually. Primer extension experiments reveal that the
length of this RNA is ~850 nt or longer. Southern blot analysis with [32P]-labelled
Cl. #4 cDNA suggests the conservation of the sequence in several mammals.
(Supported by NIH Grant HL 37702 and an AHA Grant.)
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