Identification of a neuron-specific human gene, KIAA1110, that is a guanine nucleotide exchange factor for ARF1 (original) (raw)
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Molecular Brain Research, 2002
ADP-ribosylation factors (ARFs) play important roles in vesicular trafficking and cytoskeletal regulation and its activation depends on guanine nucleotide-exchange proteins (GEPs). By way of in situ hybridization histochemistry, the localization of mRNAs for subfamily members of low-molecular-weight ARF-GEPs in the rat brain was studied at embryonic and postnatal stages. In the embryonic brain, the gene expression for msec7-1 was distinct in the ventricular zone while that for msec7-1,-3 and EFA6 in the mantle zone. In early postnatal brain, the expression for msec7-1,-2,-3 and EFA6 was seen widely in various loci of the gray matter with different intensity, and the expression of msec7-1 and-2 mRNAs was evident in the cerebellar external granule cell layer. In the adult brain, the gene expression for the four ARF-GEPs decreased more or less in most gray matter and the distinct expression was maintained mainly in the hippocampal and dentate neuronal layers and cerebellar cortex. The expression of EFA6 mRNA was also evident in the molecular layer of the hippocampus and dentate gyrus. No obvious gene expression for cytohesin-4 and ARF-GEP100 was detected in the brain at any stages of development. The present findings suggest that ARF-GEPs are differentially involved in some processes essential to neuronal differentiation and maturation in association with ARFs.
Molecular Brain Research, 2001
ADP-ribosylation factors (ARFs) play crucial roles in the intracellular vesicular transport and in regulation of phospholipid-modifying enzyme activities and cytoskeletons. Using in situ hybridization histochemistry, the gene expression for six isoforms of ARF was examined during normal development of rats and in the hypoglossal nucleus following its axotomy. In the embryonic brain, the expression for ARF-1,-4,-5,-6 mRNAs was distinct in the ventricular germinal zone while that for ARF-3,-4,-5 in the mantle zone. In early postnatal brain, the expression for six ARFs was seen widely in various loci of the gray matter with different intensity, and the expression of ARF-4,-5,-6 mRNAs was evident in the cerebellar external granule cell layer. In the adult brain, the gene expression for all ARF isoforms decreased more or less in most gray matters and the distinct expression was maintained mainly in the hippocampal and dentate neuronal layers and cerebellar cortex. The expression levels of ARF-2 and-4 mRNAs in affected hypoglossal nucleus increased after 24 h up to 7 days following axotomy of the hypoglossal nerve, while no such changes were seen in the expression levels for the other ARFs. The present findings suggest that ARFs are differentially involved in some processes essential to nerve regeneration as well as neuronal differentiation and maturation.
ARF-GEP100, a guanine nucleotide-exchange protein for ADP-ribosylation factor 6
Proceedings of The National Academy of Sciences, 2001
A human cDNA encoding an 841-aa guanine nucleotide-exchange protein (GEP) for ADP-ribosylation factors (ARFs), named ARF-GEP 100, which contains a Sec7 domain, a pleckstrin homology (PH)-like domain, and an incomplete IQ-motif, was identified. On Northern blot analysis of human tissues, a Ϸ8-kb mRNA that hybridized with an ARF-GEP 100 cDNA was abundant in peripheral blood leukocytes, brain, and spleen. ARF-GEP100 accelerated [ 35 S]GTP␥S binding to ARF1 (class I) and ARF5 (class II) 2-to 3-fold, and to ARF6 (class III) ca. 12-fold. The ARF-GEP 100 Sec7 domain contains Asp 543 and Met 555 , corresponding to residues associated with sensitivity to the inhibitory effect of the fungal metabolite brefeldin A (BFA) in yeast Sec7, but also Phe 535 and Ala 536 , associated with BFA-insensitivity. The PH-like domain differs greatly from those of other ARF GEPs in regions involved in phospholipid binding. Consistent with its structure, ARF-GEP 100 activity was not affected by BFA or phospholipids. After subcellular fractionation of cultured T98G human glioblastoma cells, ARF6 was almost entirely in the crude membrane fraction, whereas ARF-GEP 100, a 100-kDa protein detected with antipeptide antibodies, was cytosolic. On immunofluorescence microscopy, both proteins had a punctate pattern of distribution throughout the cells, with apparent colocalization only in peripheral areas. The coarse punctate distribution of EEA-1 in regions nearer the nucleus appeared to coincide with that of ARF-GEP 100 in those areas. No similar coincidence of ARF-GEP100 with AP-1, AP-2, catenin, LAMP-1, or 58K was observed. The new human BFA-insensitive GEP may function with ARF6 in specific endocytic processes.
Journal of Neurochemistry, 2004
We cloned from a rat brain cDNA library a novel cDNA and named it a potential synaptic guanine nucleotide exchange factor (GEF) for Arf (synArfGEF (Po)) (GenBank Accession no. AB057643) based on its domain structure and localization. The cloned gene was 7410 bases long with a 3585-bp coding sequence encoding a protein of 1194 amino acids. The deduced protein contained a coiled-coil structure in the N-terminal portion followed by Sec7 and Plekstrin homology (PH) domains. Thus, the protein was a member of the Sec7 family of proteins, GEFs. Conservation of the ADP-ribosylation factor (Arf)-binding sequence suggested that the protein was a GEF for Arf. The gene was expressed specifically in the brain, where it exhibited region-specific expression. The protein was highly enriched in the postsynaptic density (PSD) fraction prepared from the rat forebrain. Uniquely, the protein interacted with PSD-95, SAP97 and Homer/Vesl 1/PSD-Zip45 via its C-terminal PDZ-binding motif and co-localized with these proteins in cultured cortical neurons. These results supported its localization in the PSD. The postsynaptic localization was also supported by immunohistochemical examination of the rat brain. The mRNA for the synArfGEF was also localized to dendrites, as well as somas, of neuronal cells. Thus, both the mRNA and the protein were localized in the postsynaptic compartments. These results suggest a postsynaptic role of synArfGEF in the brain.
The Journal of biological chemistry, 1992
Deletion of the amino-terminal 17 residues from human ADP-ribosylation factor (ARF) resulted in a protein ([delta 1-17]mARF1p) devoid of ARF activity but which retained the ability to bind guanine nucleotides with high affinity. Unlike the wild type, the binding of guanine nucleotides to this deletion mutant was found to be independent of added phospholipids. A chimeric protein was produced, consisting of 10% (the amino-terminal 17 amino acids) human ARF1p and 90% ARL1p, an ARF-like protein (55% identical protein sequence) from Drosophila. This chimera was found to have ARF activity, lacking in the parental ARL1 protein. Thus, the amino terminus of ARF1p was shown to be a critical component of ARF activity. A synthetic peptide, derived from the amino terminus of ARF1p, has no ARF activity. Rather, the peptide was found to be a specific inhibitor of ARF activities. This peptide was also found to be a potent and specific inhibitor of both an in vitro intra-Golgi transport assay and th...
A presynaptic role for the ADP ribosylation factor (ARF)-specific GDP/GTP exchange factor msec7-1
Proceedings of the National Academy of Sciences, 1999
ADP ribosylation factors (ARFs) represent a family of small monomeric G proteins that switch from an inactive, GDP-bound state to an active, GTP-bound state. One member of this family, ARF6, translocates on activation from intracellular compartments to the plasma membrane and has been implicated in regulated exocytosis in neuroendocrine cells. Because GDP release in vivo is rather slow, ARF activation is facilitated by specific guanine nucleotide exchange factors like cytohesin-1 or ARNO. Here we show that msec7-1, a rat homologue of cytohesin-1, translocates ARF6 to the plasma membrane in living cells. Overexpression of msec7-1 leads to an increase in basal synaptic transmission at the Xenopus neuromuscular junction. msec7-1-containing synapses have a 5-fold higher frequency of spontaneous synaptic currents than control synapses. On stimulation, the amplitudes of the resulting evoked postsynaptic currents of msec7-1-overexpressing neurons are increased as well. However, further stimulation leads to a decline in amplitudes approaching the values of control synapses. This transient effect on amplitude is strongly reduced on overexpression of msec7-1E157K, a mutant incapable of translocating ARFs. Our results provide evidence that small G proteins of the ARF family and activating factors like msec7-1 play an important role in synaptic transmission, most likely by making more vesicles available for fusion at the plasma membrane.
An ADP-Ribosylation-Factor(ARF)-Like Protein involved in Regulated Secretion
European Journal of Biochemistry, 1997
A rat ADP-ribosylation factor(ARF)-like protein named ARLI 84 was identified by cDNA cloning. The corresponding recombinant protein had an apparent molecular mass of 22000. The deduced amino acid sequence had 55 % identity with the human ARLI and four functional GTP-binding sites. Immunofluorescent confocal microscopy studies showed that ARL184 was present in the cytosol as well as in the Golgi apparatus, raising the possibility that it has a role in a secretory pathway. The involvement of this ARF-like protein in secretion was confirmed by demonstrating that ARLI 84 potentiated acetylcholine release in stably transfected PC12 cells. Collectively these results suggest that this ARL protein is a component of a regulated secretory pathway involved in Caz+-dependent release of acetylcholine.
FEBS Letters, 1999
The novel ARF-like GTPase ARL7 is a close relative of ARL4 and ARL6 (71% and 59% identical amino acids). A striking characteristic of these GTPases is their basic C-terminus which, when fused to the C-terminus of green fluorescent protein (GFP), targets the constructs to the nucleus of transfected COS-7 cells. Full length ARL4 was detected in both nuclear and extranuclear compartments, whereas a construct of ARL4 lacking its C-terminus was excluded from the nucleus. Nucleotide exchange rates of recombinant ARL4, ARL6 and ARL7 were similar and appeared considerably higher than those of other members of the ARF family (ARF1, ARP). It is concluded that ARL4, ARL6 and ARL7 form a subgroup within the ARF family with similar, possibly nuclear, function.
Journal of Biological Chemistry, 2011
ADP-ribosylation factors (ARFs) and their activating guanine nucleotide exchange factors (GEFs) play key roles in membrane traffic and signaling. All ARF GEFs share a ϳ200-residue Sec7 domain (Sec7d) that alone catalyzes the GDP to GTP exchange that activates ARF. We determined the crystal structure of human BIG2 Sec7d. A C-terminal loop immediately following helix J (loop>J) was predicted to form contacts with helix H and the switch I region of the cognate ARF, suggesting that loop>J may participate in the catalytic reaction. Indeed, we identified multiple alanine substitutions within loop>J of the full length and/or Sec7d of two large brefeldin A-sensitive GEFs (GBF1 and BIG2) and one small brefeldin A-resistant GEF (ARNO) that abrogated binding of ARF and a single alanine substitution that allowed ARF binding but inhibited GDP to GTP exchange. Loop>J sequences are highly conserved, suggesting that loop>J plays a crucial role in the catalytic activity of all ARF GEFs. Using GEF mutants unable to bind ARF, we showed that GEFs associate with membranes independently of ARF and catalyze ARF activation in vivo only when membrane-associated. Our structural, cell biological, and biochemical findings identify loop>J as a key regulatory motif essential for ARF binding and GDP to GTP exchange by GEFs and provide evidence for the requirement of membrane association during GEF activity. ADP-ribosylation factors (ARFs) 3 are key regulators of membrane traffic and intracellular signaling. ARFs cycle between an