Jennifer P. Macke

Four new members of the fibroblast growth factor (FGF) family, referred to as fibroblast growth factor homologous factors (FHFs), have been identified by a combination of random cDNA sequencing, data base searches, and degenerate PCR. Pairwise comparisons between the four FHFs show between 58% and 71% amino acid sequence identity, but each FHF shows less than 30% identity when compared with other FGFs. Like FGF-1 (acidic FGF) and FGF-2 (basic FGF), the FHFs lack a classical signal sequence and contain clusters of basic residues that can act as nuclear localization signals. In transiently transfected 293 cells FHF-1 accumulates in the nucleus and is not secreted. Each FHF is expressed in the developing and adult nervous systems, suggesting a role for this branch of the FGF family in nervous system development and function.

In Drosophila melanogaster, the frizzled gene plays an essential role in the development of tissue polarity as assessed by the orientation of cuticular structures. Through a combination of random cDNA sequencing, degenerate polymerase chain reaction amplification, and low stringency hybridization we have identified six novel frizzled homologues from mammals, at least 11 from zebrafish, several from chicken and sea urchin, and one from Caenorhabditis elegans. The complete deduced amino acid sequences of the mammalian and nematode homologues share with the Drosophila frizzled protein a conserved amino-terminal cysteine-rich domain and seven putative transmembrane segments. Each of the mammalian homologues is expressed in a distinctive set of tissues in the adult, and at least three are expressed during embryogenesis. As hypothesized for the Drosophila frizzled protein, the frizzled homologues are likely to act as transmembrane receptors for as yet unidentified ligands. These observations predict the existence of a family of signal transduction pathways that are homologous to the pathway that determines tissue polarity in Drosophila.

Expression of the yeast his3 and other amino acid biosynthetic genes is induced during conditions of amino acid starvation. The coordination of this response is mediated by a positive regulatory protein called GCN4, which binds specifically to regulatory sites upstream of all coregulated genes and stimulates their transcription. The nucleotide sequence requirements of the his3 regulatory site were determined by analysis of numerous point mutations obtained by a novel method of cloning oligonucleotides. Almost all single base pair mutations within the nine base pair sequence ATGACTCTT significantly reduce his3 induction in vivo and GCN4 binding in vitro, whereas changes outside this region have minimal effects. One mutation, which generates a sequence that most closely resembles the consensus for 15 coregulated genes, increases both the level of induction and the affinity for GCN4 protein. The palindromic nature of the optimal sequence, ATGACTCAT, suggest that GCN4 protein binds as a dimer to adjacent half-sites that possibly overlap.