Abstract
Myosin-X (Myo10) is an unconventional myosin best known for its striking localization to the tips of filopodia. Despite the broad expression of Myo10 in vertebrate tissues, its functions at the organismal level remain largely unknown. We report here the generation of KO-first (Myo10tm1a/tm1a), floxed (Myo10tm1c/tm1c), and KO mice (Myo10tm1d/tm1d). Complete knockout of Myo10 is semi-lethal, with over half of homozygous KO embryos exhibiting exencephaly, a severe defect in neural tube closure. All Myo10 KO mice that survive birth exhibit a white belly spot, all have persistent fetal vasculature in the eye, and ~50% have webbed digits. Myo10 KO mice that survive birth can breed and produce litters of KO embryos, demonstrating that Myo10 is not absolutely essential for mitosis, meiosis, adult survival, or fertility. KO-first mice and an independent spontaneous deletion (Myo10m1J/m1J) exhibit the same core phenotypes. During retinal angiogenesis, KO mice exhibit a ~50% decrease in endothelial filopodia, demonstrating that Myo10 is required to form normal numbers of filopodia in vivo. The Myo10 mice generated here demonstrate that Myo10 has important functions in mammalian development and provide key tools for defining the functions of Myo10 in vivo.
Highlights
Myosin-X (Myo10) is an unconventional myosin expressed in most vertebrate tissues, including brain, testis, kidney, and endothelia[1]
To generate mice that completely lack all forms of Myo[10], we began by obtaining C57BL/6 derived embryonic stem cells from the Knockout Mouse Project (KOMP) that target exon 27 of Myo[10] with a KO-first cassette (www.komp.org; Myo10tm1a(KOMP)Wtsi)
To conclusively define the phenotype resulting from loss of full-length and headless Myo[10], we examined mice carrying an independent mutation (Myo10m1J) that was recently listed along with 91 other new mouse mutations identified at The Jackson Laboratory[50]
Summary
Myosin-X (Myo10) is an unconventional myosin expressed in most vertebrate tissues, including brain, testis, kidney, and endothelia[1]. This work showed that use of an alternative transcription start site located ~8.6 kb into intron 19 generates a headless specific exon consisting entirely of 5′ UTR and that splicing this to exons 20–41 generates a transcript for headless Myo[10]. This headless transcript (corresponding to NCBI reference sequence XM_192774.2) encodes a protein expected to initiate at M644 and lack most of the motor domain, but is otherwise identical to full-length Myo[10]. Headless Myo[10] does not induce filopodia and has been hypothesized to act as a natural dominant negative and/or as a scaffold that interacts with Myo[10] binding partners such as PIP3, microtubules, and β-integrins[29,30]
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