Krüppel-type zinc finger (ZNF) motifs are prevalent components of transcription factor proteins in all eukaryotes. KRAB-ZNF proteins, in which a potent repressor domain is attached to a tandem array of DNA-binding zinc-finger motifs, are specific to tetrapod vertebrates and represent the largest class of ZNF proteins in mammals. To define the full repertoire of human KRAB-ZNF proteins, we searched the genome sequence for key motifs and then constructed and manually curated gene models incorporating those sequences. The resulting gene catalog contains 423 KRAB-ZNF protein-coding loci, yielding alternative transcripts that altogether predict at least 742 structurally distinct proteins. Active rounds of segmental duplication, involving single genes or larger regions and including both tandem and distributed duplication events, have driven the expansion of this mammalian gene family. Comparisons between the human genes and ZNF loci mined from the draft mouse, dog, and chimpanzee genomes not only identified 103 KRAB-ZNF genes that are conserved in mammals but also highlighted a substantial level of lineage-specific change; at least 136 KRAB-ZNF coding genes are primate specific, including many recent duplicates. KRAB-ZNF genes are widely expressed and clustered genes are typically not coregulated, indicating that paralogs have evolved to fill roles in many different biological processes. To facilitate further study, we have developed a Web-based public resource with access to gene models, sequences, and other data, including visualization tools to provide genomic context and interaction with other public data sets.

In yeast, certain resident trans-Golgi network (TGN) proteins achieve steady-state localization by cycling through late endosomes. Here, we show that chitin synthase III (Chs3p), an enzyme involved in the assembly of the cell wall at the mother-bud junction, populates an intracellular reservoir that is maintained by a cycle of transport between the TGN and early endosomes. Traffic of Chs3p from the TGN/early endosome to the cell surface requires CHS5 and CHS6, mutant alleles of which trap Chs3p in the TGN/early endosome. Disruption of the clathrin adaptor protein complex 1 (AP-1) restores Chs3p transport to the plasma membrane. Similarly, in AP-1 deficient cells, the resident TGN/early endosome syntaxin, Tlg1p, is missorted. We propose that clathrin and AP-1 act to recycle Chs3p and Tlg1p from the early endosome to the TGN.

The mitogen-activated protein kinase (MAPK) Kss1 has a dual role in regulating filamentous (invasive) growth of the yeast Saccharomyces cerevisiae. The stimulatory function of Kss1 requires both its catalytic activity and its activation by the MAPK/ERK kinase (MEK) Ste7; in contrast, the inhibitory function of Kss1 requires neither. This study examines the mechanism by which Kss1 inhibits invasive growth, and how Ste7 action overcomes this inhibition. We found that unphosphorylated Kss1 binds directly to the transcription factor Ste12, that this binding is necessary for Kss1-mediated repression of Ste12, and that Ste7-mediated phosphorylation of Kss1 weakens Kss1-Ste12 interaction and relieves Kss1-mediated repression. Relative to Kss1, the MAPK Fus3 binds less strongly to Ste12 and is correspondingly a weaker inhibitor of invasive growth. Analysis of Kss1 mutants indicated that the activation loop of Kss1 controls binding to Ste12. Potent repression of a transcription factor by its physical interaction with the unactivated isoform of a protein kinase, and relief of this repression by activation of the kinase, is a novel mechanism for signal-dependent regulation of gene expression.[Key Words: Protein-protein interaction; signal transduction; gene regulation; developmental control; invasive growth]Received June 8, 1998; revised version accepted July 24, 1998.Commitment of a cell to a developmental fate entails a change in the pattern of gene expression. In many instances, this gene regulation is controlled by signaltransduction pathways that affect the phosphorylation of transcription factors (or their associated proteins) by protein kinases (for review, see Karin 1994;Hill and Treisman 1995;Treisman 1996). Phosphorylation can control the localization or sequestration of many transcription factors, as shown for yeast Swi5, Drosophila dorsal and yan, SV40 large T antigen, and vertebrate NF-B, NFAT, STATs, and SMADs (for review, see Hill and Treisman 1995;Vandromme et al. 1996;Heldin et al. 1997). Phosphorylation by protein kinases can also influence the DNA-binding activity of transcription factors. For example, DNA-binding of Elk-1 is stimulated by the Erk and Jnk mitogen-activated protein kinases (MAPKs), whereas DNA-binding of c-Jun is inhibited by CKII (for review, see Treisman 1996). The transactivation potential of transcription factors can also be regulated independently of DNA binding. For example, protein kinase A (PKA)-mediated phosphorylation of CREB, as well as Jnk-mediated phosphorylation of c-Jun, stimulates the binding of the coactivators CREB-binding protein (CBP) and P300 (for review, see Goldman et al. 1997), and Cdkmediated phosphorylation of Rb leads to dissociation of this repressor from E2F (for review, see DePinho 1998). Finally, protein kinases can regulate transcription factor stability (Clevers and van de Wetering 1997). In all of these cases, the catalytic (phosphotransferase) activity of the protein kinase involved is an indispensable component of its regulatory action.Recent studies of...

Most genes are conserved in mammals, but certain gene families have acquired large numbers of lineage-specific loci through repeated rounds of gene duplication, divergence, and loss that have continued in each mammalian group. One such family encodes KRAB-zinc finger (KRAB-ZNF) proteins, which function as transcriptional repressors. One particular subfamily of KRAB-ZNF genes, including ZNF91, has expanded specifically in primates to comprise more than 110 loci in the human genome. Genes of the ZNF91 subfamily reside in large gene clusters near centromeric regions of human chromosomes 19 and 7 with smaller clusters or isolated copies in other locations. Phylogenetic analysis indicates that many of these genes arose before the split between the New and Old World monkeys, but the ZNF91 subfamily has continued to expand and diversify throughout the evolution of apes and humans. Paralogous loci are distinguished by divergence within their zinc finger arrays, indicating selection for proteins with different regulatory targets. In addition, many loci produce multiple alternatively spliced transcripts encoding proteins that may serve separate and perhaps even opposing regulatory roles because of the modular motif structure of KRAB-ZNF genes. The tissue-specific expression patterns and rapid structural divergence of ZNF91 subfamily genes suggest a role in determining gene expression differences between species and the evolution of novel primate traits.