Disruption of hyaluronan synthase-2 abrogates normal cardiac morphogenesis and hyaluronan-mediated transformation of epithelium to mesenchyme
Todd D. Camenisch, … , Scott E. Klewer, John A. McDonald
Todd D. Camenisch, … , Scott E. Klewer, John A. McDonald
Published August 1, 2000
Citation Information: J Clin Invest. 2000;106(3):349-360. https://doi.org/10.1172/JCI10272.
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Article

Disruption of hyaluronan synthase-2 abrogates normal cardiac morphogenesis and hyaluronan-mediated transformation of epithelium to mesenchyme

Abstract

We identified hyaluronan synthase-2 (Has2) as a likely source of hyaluronan (HA) during embryonic development, and we used gene targeting to study its function in vivo. Has2–/– embryos lack HA, exhibit severe cardiac and vascular abnormalities, and die during midgestation (E9.5–10). Heart explants from Has2–/– embryos lack the characteristic transformation of cardiac endothelial cells into mesenchyme, an essential developmental event that depends on receptor-mediated intracellular signaling. This defect is reproduced by expression of a dominant-negative Ras in wild-type heart explants, and is reversed in Has2–/– explants by gene rescue, by administering exogenous HA, or by expressing activated Ras. Conversely, transformation in Has2–/– explants mediated by exogenous HA is inhibited by dominant-negative Ras. Collectively, our results demonstrate the importance of HA in mammalian embryogenesis and the pivotal role of Has2 during mammalian development. They also reveal a previously unrecognized pathway for cell migration and invasion that is HA-dependent and involves Ras activation.

Authors

Todd D. Camenisch, Andrew P. Spicer, Tammy Brehm-Gibson, Jennifer Biesterfeldt, Mary Lou Augustine, Anthony Calabro Jr., Steven Kubalak, Scott E. Klewer, John A. McDonald

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Gene targeting of the Has2 locus. (a) Schematic of a portion of the Has2...
Gene targeting of the Has2 locus. (a) Schematic of a portion of the Has2 locus with restriction sites, exons (filled boxes), the targeting vector, and resulting targeted locus. Homologous recombination replaces the end of intron 3 and the first 60 codons of exon 4 with PGK-Neo. The BamHI and EcoRI restriction fragments confirming the structure of PCR-positive ES clones are indicated. Arrowheads indicate the direction of transcription of PGK-Neo and the diphtheria toxin A chain. (b) PCR screening with PGK-Neo and flanking primer revealing predicted amplicons of 1.8 kb in two ES clones. (c) Southern blot analysis of BamHI genomic DNA digests from the parental ES line (control), the two targeted ES clones, a representative wild-type mouse, and a heterozygous mouse. Probe 2 (box 2 in a) detected the 12.5-kb and 5.4-kb restriction fragments corresponding to the wild-type and targeted alleles, respectively. DTA, diphtheria toxin A chain.