Vivo-Morpholinos: Same oligos, from cultures to critters -


Zebrafish neurofibromatosis type 1 genes have redundant functions in tumorigenesis and embryonic development
Jimann Shin, Arun Padmanabhan, Eric D. de Groh, Jeong-Soo Lee, Sam Haidar, Suzanne Dahlberg, Feng Guo, Shuning He, Marc A. Wolman, Michael Granato, Nathan D. Lawson, Scot A. Wolfe, Seok-Hyung Kim, Lilianna Solnica-Krezel, John P. Kanki, Keith L. Ligon, Jonathan A. Epstein, A. Thomas Look


Neurofibromatosis type 1 (NF1) is a common, dominantly inherited genetic disorder that results from mutations in the neurofibromin 1 (NF1) gene. Affected individuals demonstrate abnormalities in neural-crest-derived tissues that include hyperpigmented skin lesions and benign peripheral nerve sheath tumors. NF1 patients also have a predisposition to malignancies including juvenile myelomonocytic leukemia (JMML), optic glioma, glioblastoma, schwannoma and malignant peripheral nerve sheath tumors (MPNSTs). In an effort to better define the molecular and cellular determinants of NF1 disease pathogenesis in vivo, we employed targeted mutagenesis strategies to generate zebrafish harboring stable germline mutations in nf1a and nf1b, orthologues of NF1. Animals homozygous for loss-of-function alleles of nf1a or nf1b alone are phenotypically normal and viable. Homozygous loss of both alleles in combination generates larval phenotypes that resemble aspects of the human disease and results in larval lethality between 7 and 10 days post fertilization. nf1-null larvae demonstrate significant central and peripheral nervous system defects. These include aberrant proliferation and differentiation of oligodendrocyte progenitor cells (OPCs), dysmorphic myelin sheaths and hyperplasia of Schwann cells. Loss of nf1 contributes to tumorigenesis as demonstrated by an accelerated onset and increased penetrance of high-grade gliomas and MPNSTs in adult nf1a+/−; nf1b−/−; p53e7/e7 animals. nf1-null larvae also demonstrate significant motor and learning defects. Importantly, we identify and quantitatively analyze a novel melanophore phenotype in nf1-null larvae, providing the first animal model of the pathognomonic pigmentation lesions of NF1. Together, these findings support a role for nf1a and nf1b as potent tumor suppressor genes that also function in the development of both central and peripheral glial cells as well as melanophores in zebrafish.



    The authors declare that they do not have any competing or financial interests.


    J.S., A.P., E.D.D., M.A.W., J.P.K., N.D.L., S.A.W., K.L.L, J.A.E. and A.T.L. conceived and designed the experiments. J.S., A.P., E.D.D., J-S.L., S.H., S.D., F.G. and M.A.W. performed the experiments. J.S., A.P., E.D.D., J.P.K., K.L.L., J.A.E. and A.T.L. analyzed the data. M.G., N.D.L., S.A.W., S-H.K., L.S-K. and K.L.L. contributed reagents, materials and analysis tools. A.P., J.S., E.D.D., J.P.K., A.T.L. and J.A.E. wrote the paper.


    This work was supported by the Department of Defense [grant numbers W81XWH-07-1-0228 and W81XWH-12-1-0125 to J.A.E. and A.T.L.] and the National Institutes of Health [grant numbers R01 HL062974 to J.A.E.; R01 HL093766 to N.D.L. and S.A.W.; T32 HL007843-15 to E.D.D.; and R01 HG002995 to L.S.-K.). A.T.L. was supported by an Innovator Award from Alex’s Lemonade Stand Foundation. A.P. was supported by a fellowship from the Sarnoff Cardiovascular Research Foundation, J.S. was supported by a fellowship from the Dana-Farber Cancer Institute (DFCI) Pediatric Low-Grade Astrocytoma Program, and J.-S.L. was supported by a Young Investigator Award from the Children’s Tumor Foundation. This work was also supported by the Vanderbilt University Academic Venture Capital Fund to L.S.-K. as well as the Spain Fund for Regenerative Medicine and the W.W. Smith Endowed Chair to J.A.E.


    Supplementary material for this article is available at

  • Received March 2, 2012.
  • Accepted May 14, 2012.

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