Steel-Dickie mutation encodes a c-kit ligand lacking transmembrane and cytoplasmic domains.

Abstract

Mice homozygous for the viable Si allele steel-Dickie (Si") are sterile, severely anemic, and black-eyed white. The nature of the Sid mutation was investigated at the molecular level and was found to be due to a 4.0-kilobase intragenic deletion in mast cell growth factor (MGF) genomic sequences, providing conclusive evidence that SI encodes MGF.As a consequence ofthis deletion, Sidis only capable ofencoding a soluble truncated growth factor that lacks both transmembrane and cytoplasmic domains. Northern analysis indicates that Sid mRNA is expressed at approximately wild-type levels in adult tissues, and yeast expression studies suggest that the Sid protein is as biologically active as wild-type soluble MGF.These studies provide a molecular basis for explaining the Sid phenotype, a description of a germ-line mutation in the transmembrane and cytoplasmic domains of a membrane-bound growth factor, and in vivo evidence for the importance of membrane-bound forms of growth factors in mammalian development.Mice carrying mutations at the dominant white spotting (W) and steel (So) loci have deficiencies in pigment cells, germ cells, and blood cells (reviewed in ref. 1). While phenotypically similar, W mutations act cell-autonomously whereas Si mutations exert their effects in the extracellular environment. The W locus encodes the c-kit protooncogene product (cKit), a tyrosine kinase receptor (2, 3). A ligand for c-Kit has recently been identified which is variously known as mast cell growth factor (4-6) (subsequently referred to here as MGF), stem cell factor (7-9), and c-Kit ligand (10, 11). This growth factor is a good candidate for the SI gene product (5, 9, 11). Like several other growth factors characterized to date (12-18), MGF exists in both membrane-bound and soluble forms (6,8,11). The primary translation products of these growth factors are membrane-bound, whereas the soluble form(s) is derived by proteolytic cleavage.We and others have shown (5, 9, 11) that MGF coding sequences are totally deleted in mice homozygous for lethal SI alleles, whereas no gross structural alterations are associated with most viable SI alleles. These results suggest, but do not prove, that MGF is encoded by SI. A simple interpretation of these data is that the lethal SI alleles result from the complete absence of MGF protein and that homozygous viable alleles either encode a mutant form ofMGF or produce lower levels of the wild-type gene product. This interpretation is consistent with a previous study showing that an intragenic deletion in c-kit produces a homozygous lethal phenotype (19). It remains possible, however, that MGF deletion mutants are viable but other genes mapping near MGF are deleted in mice carrying lethal Si alleles and that these genes are responsible for the lethality.In this report we show that one of the viable mutant SI alleles, steel-Dickie (Sld), has a 4.0-kilobase (kb) intragenic deletion that truncates the SI coding sequence. These results provide direct evidence that Si encodes MGF and a molecul...

Human Stem Cell Factor Dimer Forms a Complex with Two Molecules of the Extracellular Domain of Its Receptor, Kit

“…5 shows the results of a simultaneous fit of Model 1 to data for sKit plus E. coli-derived SCF mixtures made up at both the two sKit per SCF dimer and one sKit per SCF dimer molar ratios, and at three different rotor speeds. This fit is excellent and returns an intrinsic dissociation constant for each site of 17 [12][13][14][15][16][17][18][19][20][21] nM. As implied by Fig.…”

“…The alternate splicing leads to a shorter form which lacks the exon 6 amino acids including the proteolytic cleavage site and therefore tends to remain membrane bound (12,13). From several different approaches, there have been indications of functional differences between soluble and membrane-bound SCFs (12,13,(15)(16)(17). Soluble SCF is found at substantial levels in human serum (18).…”

c-Kit is required for growth and survival of the cells of origin of Brca1-mutation-associated breast cancer

“…In vivo, only the membrane bound form supports survival of long-term haematopoietic progenitors (Gommerman et al, 2000;Driessen et al, 2003). The Steel-Dickie mutation in mice generates an SCF protein lacking both transmembrane and cytoplasmic domains and results in production of only soluble SCF (Brannan et al, 1991), supporting the importance of the membrane bound form in a number of biological processes. It is not yet clear which form predominates in the mammary epithelium.…”

Rôle des facteurs de croissance et cytokines dans le testicule normal et pathologique

“…De plus, les formes solubles et membranaires du SCF n'auraient pas les m6mes propri6t6s. En effet, la mutation S1 d (Steel Dickie : d616tion du domaine transmembranaire, [6]) provoque la production de SCF sous forme soluble uniquement, les cellules germinales primordiales ont migr6 dans la gonade fcetale, cependant, l'animal est st6rile car h la pubert6, les cellules germinales ne se multiplient pas. Ces donn6es exp~rimentales sugg~rent que le SCF (forme soluble et surtout membranaire) aurait un r61e c16 dans la migration et la prolif6ration des cellules de la lign6e germinale.…”