Isoforms of the human ets-1 protein: generation by alternative splicing and differential phosphorylation

The ets-1 gene belongs to the ets gene family (ets-1, ets-2, erg, and elk) and is homologous to the v-ets oncogene found in the avian leukemia virus E26. The ets-1 gene products were characterized using a specific monoclonal antibody developed against a bacterially expressed v-ets protein. The ets-1 gene product in the human T-cell line CEM was found to consist of at least six species: four major species with apparent molecular weights of 51 kDa (p51), 48 kDa (p48), 42 kDa (p42), and 39 kDa (p39); and two minor species of 52 kDa (pp52) and 49 kDa (pp49), which are demonstrated to be the phosphorylated forms of p51 and p48, respectively. All of the ets-1 proteins are related to each other and are considered products of the ets-1 gene. Subcellular localization showed that the pp52 and p51 are found mainly in the cytoplasm, while p48 and p39 are found mainly in the nucleus. Specific antibodies against various exons of ets-1 showed that both p42 and p39 lack a region corresponding to exon VII. Polymerase chain reaction analyses revealed the presence of an additional RNA product that corresponds to mRNA lacking exon VII. These results suggest that the human ets-1 gene encodes multiple proteins that are generated by at least two distinct mechanisms: alternative splicing of mRNA and protein phosphorylation.     

Identification and characterization of human GUKH2 gene in silico

Drosophila Guanylate-kinase holder (Gukh) is an adaptor molecule bridging Discs large (Dlg) and Scribble (Scrib), which are implicated in the establishment and maintenance of epithelial polarity. Here, we searched for human homologs of Drosophila gukh by using bioinformatics, and identified GUKH1 and GUKH2 genes. GUKH1 was identical to Nance-Horan syndrome (NHS) gene, while GUKH2 was a novel gene. FLJ35425 (AK092744.1), DKFZp686P1949 (BX647246.1) and KIAA1357 (AB037778.1) cDNAs were derived from human GUKH2 gene. Nucleotide sequence of GUKH2 cDNA was determined by assembling 5'-part of FLJ35425 cDNA and entire region of DKFZp686P1949 cDNA. Human GUKH2 gene consists of 8 exons. Exon 5 (132 bp) of GUKH2 gene was spliced out in GUKH2 cDNA due to alternative splicing. GUKH2-REPS1 locus at human chromosome 6q24.1 and GUKH1-REPS2 locus at human chromosome Xp22.22-p22.13 are paralogous regions within the human genome. Mouse Gukh2 and zebrafish gukh2 genes were also identified. N-terminal part of human GUKH2, mouse Gukh2 and zebrafish gukh2 proteins were completely divergent from human GUKH1 protein. Human GUKH2 and GUKH1, consisting of eight GUKH homology (GKH1-GKH8) domains and Proline-rich domain, showed 28.5% total-amino-acid identity. GKH1, GKH4, GKH5, GKH7 and GKH8 domains were conserved among human GUKH1, human GUKH2 and Drosophila Gukh. Because human homologs of Drosophila dlg (DLG1-DLG7) as well as human homologs of Drosophila scrib (SCRIB, ERBB2IP and Densin-180) are cancer-associated genes, human homologs of Drosophila gukh (GUKH1 and GUKH2) are predicted cancer-associated genes.     

Identification and characterization of human LLGL4 gene and mouse Llgl4 gene in silico

Drosophila Discs large (Dlg), Scribble (Scrib) and Lethal giant larvae (Lgl) act in concert as regulators of epithelial polarity, and human homologs of Drosophila dlg, scrib, and lgl are cancer-associated genes. LLGL1, LLGL2, and LLGL3/STXBP5 genes, encoding LGL1, LGL2, and LGL3/Tomosyn, respectively, are human homologs of Drosophila lgl gene. Here, we identified and characterized LLGL4 (also known as STXBP5L) gene encoding LGL4 protein, by using bioinformatics. Uncharacterized human KIAA1006 cDNA (AB023223) was derived from human LLGL4 gene. LLGL4 mRNA was expressed in kidney, brain hippocampus, and also in lung carcinoid, and germ cell tumors. LLGL4 gene, consisting of 28 exons, was mapped to human chromosome 3q13.33. Mouse A830015P08Rik cDNA (NM_172440.1) was a 3'-truncated partial Llgl4 cDNA. Nucleotide sequence of full-length mouse Llgl4 cDNA was determined in silico by assembling A830015P08Rik cDNA, BU609516 EST and last two exons of Llgl4 gene within mouse genome clone RP24-174G4 (AC118742.3). Human LGL4 showed 95.8% total-amino-acid identity with mouse Lgl4, and 68.4% total-amino-acid identity with human LGL3. LGLH1 domain (codon 1-11 of LGL4), LGLH2 domain (codon 52-98) and LGLH3 domain (codon 994-1054) were identified as novel conserved regions among LGL family members. LGL1 and LGL2 consist of LGLH1, LGLH2, LGLH3 domains and five WD40 repeats, while LGL3 and LGL4 consist of LGLH1, LGLH2, LGLH3 domains, five WD40 repeats and the C-terminal Syntaxin-binding SNARE domain. This is the first report on identification and characterization of human LLGL4 and mouse Llgl4 genes.     

The human erg gene maps to chromosome 21, band q22: relationship to the 8; 21 translocation of acute myelogenous leukemia

There is accumulating evidence to support that genes on chromosome 21 play an important role in the development of pathologies associated with leukemia, Down's syndrome, and Alzheimer's disease. We have previously described erg, a human gene related to the ets oncogene. In this study, we have regionally assigned the erg gene to chromosome 21q22.3 by using somatic cell hybrids and in situ hybridization analysis. In light of this chromosome assignment, the relationship of erg to the 21q translocation breakpoint characteristic of acute myelogenous leukemia (AML) was considered. By using a DNA probe that is specific for the erg gene, a panel of rodent-human cell hybrids was analyzed by the Southern technique to study specific chromosome translocations occurring in acute myeloblastic leukemia. The erg gene was found to translocate from chromosome 21 to 8 in the t(8; 21) (q22; q22), a non-random translocation found in patients with acute myelogenous leukemia of the subgroup M2 (AML-M2). The localization of the erg gene to chromosome 21q22 raises the possibility that this gene may be involved in the pathogenesis of AML-M2.     

Sequence and functional properties of Ets genes in the model organism Drosophila

Detailed molecular and genetic studies, coupled with the recent sequencing of the fly genome, have identified eight Ets-related genes in the model organism Drosophila. All show homology to genes in vertebrate species. Functional analyses of some of the Drosophila ets genes have revealed their essential roles in developmental processes such as metamorphosis, oogenesis, neurogenesis, myogenesis, and eye development. Such studies have yielded important insights into our understanding of the genetic control of hormonally-regulated gene expression, programmed cell death, and signal transduction during cell fate determination and differentiation. The developmental roles of E74 (ELF1), pointed (Ets 1), yan (TEL), and D-elg (GABPalpha) will be reviewed in this article. The context of their participation in signal transduction and gene regulation will also be discussed. The information should be of significant value to the study of related processes in higher organisms due to the growing evidence for the cross species conservation of developmental mechanisms.