The purpose of this study is to understand the differences between the two nonmuscle myosin II (NM II) isoforms, NM IIA and NM IIC1 by studying whether NM IIC1 can functionally replace NM IIA during development. To genetically replace NM IIA with NM IIC1 in mice , nonmuscle myosin heavy chain NM IIA was ablated by inserting cDNA encoding NMHC IIC1-GFP into the first coding exon of the gene encoding NMHC IIA. Heterozygous AC1*-gfp/A+ mice, expressing 50% wild-type NM IIA, are normal. Homozygous AC1*gfp/AC1*gfp mice die at approximately E10.5 demonstrating that NM IIC1 can rescue the failure of visceral endoderm function that results in embryonic day (E)6.5 lethality of A−/A−mice. This confirms an isoform-independent requirement for NM II in supporting visceral endoderm function and consequent gastrulation. In contrast, lethality at E 10.5 of the AC1*gfp/AC1*gfp embryos appears to be due to increased apoptosis and abnormalities in the placenta which shows a defect in vasculogenesis and angiogenesis. Abnormalities in cell migration, focal adhesion formation and defects in the actomyosin cytoskeleton structure are found in cultured AC1*gfp/AC1*gfp MEF cells and help to explain the placental defects. We have modeled these placental vascular defects in allantois explants to confirm a role for NM IIA and NM II phosphorylation in vascular development. Thus NM IIC1, which is not normally expressed in mice until E11, can restore cell-cell adhesions and normal visceral endoderm function at E6.5 but cannot replace NM II As roles in cytoskeleton organization and placenta vascular formation. Previously we modeled 3 different mutations that are found in human NM IIA in mice (R702C, D1424N and E1841K). We reported that all three lines phenocopied human MYH9-Related disease. In addition, male mice that are homozygous for the mutation E1841K exhibit infertility due to a lack of mature and motile sperm. Electron microscopy on the mutant testes shows evidence for germ cell apoptosis, multinucleated giant cells and a lack of late stage elongated spermatids

Project Start
Project End
Budget Start
Budget End
Support Year
21
Fiscal Year
2013
Total Cost
$446,623
Indirect Cost
Name
National Heart, Lung, and Blood Institute
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