Summary Conditional β 1-integrin gene deletion in neural crest cells causes severe developmental alterations of the peripheral nervous system

The ontogeny of neural crest cells (NCC) is a major morphogenetic process involving various biological steps, such as epithelium-mesenchyme transition, cell migration and cell aggregation. NCC differentiate to generate a diversity of cell types, including craniofacial structures, the peripheral nervous system (PNS) and melanocytes (Le Douarin, 1982; Le Douarin and Kalcheim, 1999). NCC interact with the extracellular matrix (ECM) during their differentiation. Integrins are α /β heterodimers that bind ECM components and cell-surface receptors, and control cell adhesion, migration, differentiation and survival (Hynes, 2002). Previous studies have shown that integrins play a major role in the control of NCC adhesion and migration. In vitro, these cells express a large repertoire of integrins, including α1β1, α3β1, α4β1, α5β1, α6β1, α8β1, αVβ1, αVβ3 and αVβ8 (Kil et al., 1996; Desban and Duband, 1997; Testaz et al., 1999). In vivo, functional analyses with integrin antibodies, antisense oligonucleotides and peptide competitors have shown that mouse and avian NCC use multiple β1-integrins (Boucaut et al., 1984; Bronner-Fraser, 1986; Poole and Thiery, 1986; Kil et al., 1998). Constitutive knockout of the β1-integrin gene leads to the loss of twelve members of the integrin family and leads to death of the embryo during the peri-implantation period due to inner cell mass failure (Fässler and Meyer, 1995; Stephens et al., 1995). This constitutive β1-knockout model demonstrates the essential role of β 1-integrins during early embryogenesis, but is of no use for investigating integrin functions during later stages of development or in differentiated cells, such as NCC derivatives. By contrast, neither the invalidation of individual α-integrin subunits (for reviews, see Bouvard et al., 2001; Hynes, 2002) nor the production of knockout chimaeric mice (Fässler and Meyer, 1995) has made it possible to delineate precisely the roles of specific integrins during NCC development, with the exception of α 5β1-integrins, which have been implicated in cranial NCC survival (Goh et al., 1997), and α5β1and α4β1-integrins, which have been implicated in the proliferation or survival of glial cell precursors (Haack and Hynes, 2001). The Cre-LoxP conditional gene disruption system has been used to investigate the effect of integrins on NCC ontogeny in more detail. P0-Cre mice crossed with mice carrying a floxed β1-integrin allele generated animals with the conditional mutation targeted specifically to Schwann cells (SC), which are derived from NCC (Feltri et al., 2002). This conditional knockout revealed the role played by β1-integrin receptors in postnatal PNS development. The loss of β1-integrins impedes interactions between SC and axons, causing dysmyelinating neuropathy (Feltri et al., 2002). We recently developed the Ht-PA-Cre transgenic mouse line, in which Cre recombinase activity is targeted specifically to NCC at the start of migration (Pietri et al., 2003). We report here the effects of specific inactivation of the β 1-integrin gene in migratory neural crest cells. The crossing of Ht-PA-Cre mice with β1 floxed mice generated animals in which the NCC produce no β1-integrin but do produce β -galactosidase under the control of the endogenous β1-integrin promoter (Potocnik et al., 2000). The mutation targets the glial and sensory neuronal compartments of the PNS as well as the other NCC derivatives. Integrins are transmembrane receptors that are known to interact with the extracellular matrix and to be required for migration, proliferation, differentiation and apoptosis. We have generated mice with a neural crest cell-specific deletion of the β1-integrin gene to analyse the role of β 1integrins in neural crest cell migration and differentiation. This targeted mutation caused death within a month of birth. The loss of β1-integrins from the embryo delayed the migration of Schwann cells along axons and induced multiple defects in spinal nerve arborisation and morphology. There was an almost complete absence of Schwann cells and sensory axon segregation and defective maturation in neuromuscular synaptogenesis. Thus, β1integrins are important for the control of embryonic and postnatal peripheral nervous system development.