The sporulation-essential gene spolIG of the S1 nuclease mapping experiments indicate that ORF3 is initially cotranscribed with spoliG from about 1 to 4 hr into the sporulation process and that later on ORF3 is transcribed independently from a new site located between spolIG and ORF3. The role of ORF3 was investigated by constructing a deletion mutation in its structural gene. The mutant exhibits normal growth but is unable to produce heat-resistant spores. We propose that the ORF3 gene product is a af factor or a related peptide essential for sporulation at a late stage of development.

The extracellular portion of the granulocyte colonystimulating factor (G-CSF) receptor has a mosaic structure of six domains (each approximately 100 amino acid residues) consisting of an immunoglobulin-like (Ig) domain, a cytokine receptor homologous region subdivided into amino-terminal (BN) and carboxyl-terminal (BC) domains, and three fibronectin type III repeats. In the present study, we expressed the Ig-BN and the BN-BC regions and purified them to homogeneity as monomers using G-CSF affinity column chromatography. Using gel filtration high performance liquid chromatography, we investigated the molecular composition of receptor-ligand complexes formed between G-CSF and purified BN-BC or Ig-BN domains. In contrast to the well characterized example of the human growth hormone (

An extracellular portion of granulocyte colony-stimulating factor (G-CSF) receptor, which contains an immunoglobulin-like (Ig) domain and cytokine receptor homologous (CRH) region, was secreted into the medium using Trichoplwia ni-Autographa californica nuclear polyhedrosis virus system. The gene product was purified to homogeneity mainly as a dimer (85 kDa) using G-CSF affinity column chromatography and gel filtration HPLC, although the product existed as a monomer (45 kDa) in the medium. Scatchard analyses suggested that only the dimer had high aBinity ligand binding (& = about 100 PM), which is comparable with the Kd value of the cell surface. receptor. The binding of G-CSF to Ig-CRH induced its tetramerization (200-250 kDa). The molecular composition of the tetrameric complex showed a stoichiometry of four ligands bound to four Ig-CRH. These results suggested that the oligomeric mechanism of the G-CSF receptor differs from that reported for growth hormone (GH) receptor, although CD spectrum spectroscopy suggested that the Ig-CRH has a GH receptor-like structure.

No abstract

We have determined the NMR structure of a ligand-binding domain of the granulocyte colony-stimulating factor (G-CSF) receptor, containing the highly conserved WSxWS motif. The domain consists of seven beta-strands with the fibronectin type III-like topology seen in several cytokine receptors. Comparisons between the spectra of the 15N-labelled domain with and without G-CSF indicate that the major ligand-recognition site is on the FG loop just upstream of the WSxWS sequence, and not on the BC loop which is mainly used in the growth hormone system. The WSxWS residues are suggested to contribute to ligand-recognition and to the protein architecture of the G-CSF receptor.