Biomedical subjects

S Kozlov

Publications and source records attributed to S Kozlov.

10 recordsLinked to original sources

PubMed2002

Novel insecticidal toxins from the venom of the spider Segestria florentina.

Three insecticidal polypeptide toxins (F5.5, F5.6, F5.7) with molecular masses 4973, 4993 and 5159Da were isolated from the venom of the central Asian spider Segestria florentina. These toxins caused the complete flaccid paralysis of Heliothis virescens (Lepidoptera: Noctuidae) larvae (LD(50) 4-10 microg/g), whereas they were inactive upon intravenous injections into mice. On the basis of N-terminal amino acid sequences a family of eight genes encoding highly homologues polypeptides (SFI1-SFI8) was revealed, some of which encode polypeptides actually demonstrated to be present in S. florentina venom. All deduced polypeptides consist of 46 amino acids residues. Comparison of primary structures of SFI1-SFI8 with other spider toxins suggests that this family might share structural and functional relationships with other small spider neurotoxins, several of which are known to be highly selective agonists/antagonists of different voltage-dependent Ca(2+) channels.

Amino Acid Sequence↗

PubMed2001

Thyroid hormone regulates TAG-1 expression in the developing rat brain.

TAG-1 is a member of the immunoglobulin superfamily of cell adhesion molecules thought to play important roles in neuronal differentiation and the establishment of connectivity during brain development. Because these are processes also affected by hypothyroidism, we studied the effects of thyroid hormone deprivation and administration on TAG-1 expression in the developing rat brain. By in situ hybridization, immunohistochemistry and Western blotting we found that TAG-1 RNA and protein levels are upregulated in the hypothyroid brain. From embryonic day 20 to postnatal day (P) 15, elevated TAG-1 RNA was found in several areas including the cerebral cortex, hippocampus and olfactory bulb. In agreement with this, TAG-1 protein was overexpressed in the major fibre tracts arising from these structures, including the corpus callosum, anterior and hippocampal commissures and lateral olfactory tract. A similar overexpression of TAG-1 by hypothyroidism was detected in the cerebellum, but starting only at P15. In all cases, elevation of TAG-1 RNA and protein expression could be reversed by thyroid hormone treatment. These results show that the deregulation of TAG-1 might contribute to the alterations caused by the lack of thyroid hormone during brain development.

Aging↗

PubMed2000

Expression and imprinting of MAGEL2 suggest a role in Prader-willi syndrome and the homologous murine imprinting phenotype.

Prader-Willi syndrome (PWS) is caused by the loss of expression of imprinted genes in chromosome 15q11-q13. Affected individuals exhibit neonatal hypotonia, developmental delay and childhood-onset obesity. Necdin, a protein implicated in the terminal differentiation of neurons, is the only PWS candidate gene to reduce viability when disrupted in a mouse model. In this study, we have characterized MAGEL2 (also known as NDNL1), a gene with 51% amino acid sequence similarity to necdin and located 41 kb distal to NDN in the PWS deletion region. MAGEL2 is expressed predominantly in brain, the primary tissue affected in PWS and in several fetal tissues as shown by northern blot analysis. MAGEL2 is imprinted with monoallelic expression in control brain, and paternal-only expression in the central nervous system as demonstrated by its lack of expression in brain from a PWS-affected individual. The orthologous mouse gene (Magel2) is located within 150 kb of NDN:, is imprinted with paternal-only expression and is expressed predominantly in late developmental stages and adult brain as shown by northern blotting, RT-PCR and whole-mount RNA in situ hybridization. Magel2 distribution partially overlaps that of NDN:, with strong expression being detected in the central nervous system in mid-gestation mouse embryos by in situ hybridization. We hypothesize that, although loss of necdin expression may be important in the neonatal presentation of PWS, loss of MAGEL2 may be critical to abnormalities in brain development and dysmorphic features in individuals with PWS.

Amino Acid Sequence↗

PubMed2000

Purification and cDNA cloning of an insecticidal protein from the venom of the scorpion Orthochirus scrobiculosus.

Injection of crude venom from the scorpion Orthochirus scrobiculosus into larvae of Heliothis virescens (Lepidoptera: Noctuidae) caused trembling and uncoordinated movement before development of a progressive and prolonged flaccid paralysis. The isolation of the toxin (OsI-1) responsible for this effect of O. scrobiclosus venom is described. The molecular mass of OsI-1 toxin was 6994 Da, as determined by desorption mass spectroscopy. The complete primary structure of OsI-1 was deduced from the sequence of cDNA clones obtained by rapid amplification of cDNA ends (RACE) PCR. Comparison of the deduced amino acid sequence of OsI-1 with those of other insecticidal scorpion toxins indicates that it is a sodium (Na+) channel active depressant insect-selective toxin. The analysis of amino acid sequence of the toxin in conjunction with mass spectroscopy data indicates post-translational modification in maturation with the removal of 3 C-terminal amino acids and amidation of the C-terminus.

Amino Acid Sequence↗

PubMed2000

ATM-dependent phosphorylation of nibrin in response to radiation exposure.

Mutations in the gene ATM are responsible for the genetic disorder ataxia-telangiectasia (A-T), which is characterized by cerebellar dysfunction, radiosensitivity, chromosomal instability and cancer predisposition. Both the A-T phenotype and the similarity of the ATM protein to other DNA-damage sensors suggests a role for ATM in biochemical pathways involved in the recognition, signalling and repair of DNA double-strand breaks (DSBs). There are strong parallels between the pattern of radiosensitivity, chromosomal instability and cancer predisposition in A-T patients and that in patients with Nijmegen breakage syndrome (NBS). The protein defective in NBS, nibrin (encoded by NBS1), forms a complex with MRE11 and RAD50 (refs 1,2). This complex localizes to DSBs within 30 minutes after cellular exposure to ionizing radiation (IR) and is observed in brightly staining nuclear foci after a longer period of time. The overlap between clinical and cellular phenotypes in A-T and NBS suggests that ATM and nibrin may function in the same biochemical pathway. Here we demonstrate that nibrin is phosphorylated within one hour of treatment of cells with IR. This response is abrogated in A-T cells that either do not express ATM protein or express near full-length mutant protein. We also show that ATM physically interacts with and phosphorylates nibrin on serine 343 both in vivo and in vitro. Phosphorylation of this site appears to be functionally important because mutated nibrin (S343A) does not completely complement radiosensitivity in NBS cells. ATM phosphorylation of nibrin does not affect nibrin-MRE11-RAD50 association as revealed by radiation-induced foci formation. Our data provide a biochemical explanation for the similarity in phenotype between A-T and NBS.

Ataxia Telangiectasia↗

PubMed2000

Zac1 (Lot1), a potential tumor suppressor gene, and the gene for epsilon-sarcoglycan are maternally imprinted genes: identification by a subtractive screen of novel uniparental fibroblast lines.

Imprinted genes are expressed from one allele according to their parent of origin, and many are essential to mammalian embryogenesis. Here we show that the epsilon-sarcoglycan gene (Sgce) and Zac1 (Lot1) are both paternally expressed imprinted genes. They were identified in a subtractive screen for imprinted genes using a cDNA library made from novel parthenogenetic and wild-type fibroblast lines. Sgce is a component of the dystrophin-sarcoglycan complex, Zac1 is a nuclear protein inducing growth arrest and/or apoptosis, and Zac1 is a potential tumor suppressor gene. Sgce and Zac1 are expressed predominantly from their paternal alleles in all adult mouse tissues, except that Zac1 is biallelic in the liver and Sgce is weakly expressed from the maternal allele in the brain. Sgce and Zac1 are broadly expressed in embryos, with Zac1 being highly expressed in the liver primordium, the umbilical region, and the neural tube. Sgce, however, is strongly expressed in the allantoic region on day 9.5 but becomes more widely expressed throughout the embryo by day 11.5. Sgce is located at the proximal end of mouse chromosome 6 and is a candidate gene for embryonic lethality associated with uniparental maternal inheritance of this region. Zac1 maps to the proximal region of chromosome 10, identifying a new imprinted locus in the mouse, homologous with human chromosome 6q24-q25. In humans, unipaternal disomy for this region is associated with fetal growth retardation and transient neonatal diabetes mellitus. In addition, loss of expression of ZAC has been described for a number of breast and ovarian carcinomas, suggesting that ZAC is a potential tumor suppressor gene.

Animals↗

PubMed1998

ATM associates with and phosphorylates p53: mapping the region of interaction.

The human genetic disorder ataxia-telangiectasia (AT) is characterized by immunodeficiency, progressive cerebellar ataxia, radiosensitivity, cell cycle checkpoint defects and cancer predisposition. The gene mutated in this syndrome, ATM (for AT mutated), encodes a protein containing a phosphatidyl-inositol 3-kinase (PI-3 kinase)-like domain. ATM also contains a proline-rich region and a leucine zipper, both of which implicate this protein in signal transduction. The proline-rich region has been shown to bind to the SH3 domain of c-Abl, which facilitates its phosphorylation and activation by ATM. Previous results have demonstrated that AT cells are defective in the G1/S checkpoint activated after radiation damage and that this defect is attributable to a defective p53 signal transduction pathway. We report here direct interaction between ATM and p53 involving two regions in ATM, one at the amino terminus and the other at the carboxy terminus, corresponding to the PI-3 kinase domain. Recombinant ATM protein phosphorylates p53 on serine 15 near the N terminus. Furthermore, ectopic expression of ATM in AT cells restores normal ionizing radiation (IR)-induced phosphorylation of p53, whereas expression of ATM antisense RNA in control cells abrogates the rapid IR-induced phosphorylation of p53 on serine 15. These results demonstrate that ATM can bind p53 directly and is responsible for its serine 15 phosphorylation, thereby contributing to the activation and stabilization of p53 during the IR-induced DNA damage response.

Ataxia Telangiectasia Mutated Proteins↗

PubMed1997

Isolation of full-length ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype.

A gene mutated in the human genetic disorder ataxia-telangiectasia (A-T), ATM, was recently identified by positional cloning. ATM is a member of the phosphatidylinositol-3-kinase superfamily, some of which are protein kinases and appear to have important roles in cell cycle control and radiation signal transduction. We describe herein, to our knowledge, for the first time, the cloning of a full-length cDNA for ATM and correction of multiple aspects of the radio-sensitive phenotype of A-T cells by transfection with this cDNA. Overexpression of ATM cDNA in A-T cells enhanced the survival of these cells in response to radiation exposure, decreased radiation-induced chromosome aberrations, reduced radio-resistant DNA synthesis, and partially corrected defective cell cycle checkpoints and induction of stress-activated protein kinase. This correction of the defects in A-T cells provides further evidence of the multiplicity of effector functions of the ATM protein and suggests possible approaches to gene therapy.

Ataxia Telangiectasia↗

PubMed1997

Interaction between ATM protein and c-Abl in response to DNA damage.

The gene mutated in the autosomal recessive disorder ataxia telangiectasia (AT), designated ATM (for 'AT mutated'), is a member of a family of phosphatidylinositol-3-kinase-like enzymes that are involved in cell-cycle control, meiotic recombination, telomere length monitoring and DNA-damage response. Previous results have demonstrated that AT cells are hypersensitive to ionizing radiation and are defective at the G1/S checkpoint after radiation damage. Because cells lacking the protein tyrosine kinase c-Abl are also defective in radiation-induced G1 arrest, we investigated the possibility that ATM might interact with c-Abl in response to radiation damage. Here we show that ATM binds c-Abl constitutively in control cells but not in AT cells. Our results demonstrate that the SH3 domain of c-Abl interacts with a DPAPNPPHFP motif (residues 1,373-1,382) of ATM. The results also reveal that radiation-induction of c-Abl tyrosine kinase activity is diminished in AT cells. These findings indicate that ATM is involved in the activation of c-Abl by DNA damage and this interaction may in part mediate radiation-induced G1 arrest.

Ataxia Telangiectasia↗

PubMed1997

Cellular localisation of the ataxia-telangiectasia (ATM) gene product and discrimination between mutated and normal forms.

The recently cloned gene (ATM) mutated in the human genetic disorder ataxia-telangiectasia (A-T) is involved in DNA damage response at different cell cycle checkpoints and also appears to have a wider role in signal transduction. Antibodies prepared against peptides from the predicted protein sequence detected a approximately 350 kDa protein corresponding to the open reading frame, which was absent in 13/23 A-T homozygotes. Subcellular fractionation, immunoelectronmicroscopy and immunofluorescence showed that the ATM protein is present in the nucleus and cytoplasmic vesicles. This distribution did not change after irradiation. We also provide evidence that ATM protein binds to p53 and this association is defective in A-T cells compatible with the defective p53 response in these cells. These results provide further support for a role for the ATM protein as a sensor of DNA damage and in a more general role in cell signalling, compatible with the broader phenotype of the syndrome.

Antibodies↗