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© The Rockefeller University Press, 0021-9525/1999//151 $5.00
The Journal of Cell Biology, Volume 144, Number 1, , 1999 151-160

Articles

Absence of Basement Membranes after Targeting the LAMC1 Gene Results in Embryonic Lethality Due to Failure of Endoderm Differentiation



Neil Smyth*,{ddagger}, H. Seda Vatansever*, Patricia Murray*, Michael Meyer§, Christian Frie{ddagger}, Mats Paulsson{ddagger}, and David Edgar*

* Department of Human Anatomy and Cell Biology, University of Liverpool, Liverpool L69 3GE, United Kingdom; {ddagger} Institute for Biochemistry, University of Cologne, D-50931 Cologne, Germany; and § Department of Neurochemistry, Max-Planck-Institute of Psychiatry, D-82152 Martinsried, Germany

The LAMC1 gene coding for the laminin {gamma}1 subunit was targeted by homologous recombination in mouse embryonic stem cells. Mice heterozygous for the mutation had a normal phenotype and were fertile, whereas homozygous mutant embryos did not survive beyond day 5.5 post coitum. These embryos lacked basement membranes and although the blastocysts had expanded, primitive endoderm cells remained in the inner cell mass, and the parietal yolk sac did not develop. Cultured embryonic stem cells appeared normal after targeting both LAMC1 genes, but the embryoid bodies derived from them also lacked basement membranes, having disorganized extracellular deposits of the basement membrane proteins collagen IV and perlecan, and the cells failed to differentiate into stable myotubes. Secretion of the linking protein nidogen and a truncated laminin {alpha}1 subunit did occur, but these were not deposited in the extracellular matrix. These results show that the laminin {gamma}1 subunit is necessary for laminin assembly and that laminin is in turn essential for the organization of other basement membrane components in vivo and in vitro. Surprisingly, basement membranes are not necessary for the formation of the first epithelium to develop during embryogenesis, but first become required for extra embryonic endoderm differentiation.

Key Words: extracellular matrix • epithelium • embryogenesis • endoderm • laminin

Abbreviations used in this paper: ES, embryonic stem; pc, post coitum; TUNEL, terminal dUTP–biotin nick end labeling.

We are greatly indebted to H. Thoenen (Max Planck Institute for Psychiatry, Munich, Germany) for providing encouragement, advice, and facilities for the initial stages of this work, to H. Thorun and B. Kunkel (both from Max Planck Institute for Psychiatry) who provided skilled technical assistance with tissue culture and microinjection, and to A. Fichard (Max Planck Institute for Psychiatry) who was involved in preliminary experiments leading to this project. We thank U. Mayer and R. Timpl (both from Max Planck Institute for Biochemistry) for generously making their antibodies available to us and also P. Soriano (Fred Hutchinson Cancer Center Research Center, Seattle, WA) and A. Nagy (Samuel Lunenfeld Research Institute, Mt. Sinai Hospital, Toronto, Canada) who provided the IRES construct and R1 ES cells, respectively.


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