Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Genes, src”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,423 records · Page 79Linked to original sources
PubMed1999

Simian immunodeficiency virus and human immunodeficiency virus type 1 nef proteins show distinct patterns and mechanisms of Src kinase activation.

The nef gene from human and simian immunodeficiency viruses (HIV and SIV) regulates cell function and viral replication, possibly through binding of the nef product to cellular proteins, including Src family tyrosine kinases. We show here that the Nef protein encoded by SIVmac239 interacts with and also activates the human Src kinases Lck and Hck. This is in direct contrast to the inhibitory effect of HIV type 1 (HIV-1) Nef on Lck catalytic activity. Unexpectedly, however, the interaction of SIV Nef with human Lck or Hck is not mediated via its consensus proline motif, which is known to mediate HIV-1 Nef binding to Src homology 3 (SH3) domains, and various experimental analyses failed to show significant interaction of SIV Nef with the SH3 domain of either kinase. Instead, SIV Nef can bind Lck and Hck SH2 domains, and its N-terminal 50 amino acid residues are sufficient for Src kinase binding and activation. Our results provide evidence for multiple mechanisms by which Nef binds to and regulates Src kinases.

Amino Acid Sequence↗
PubMed1997

Phosphorylation and high level expression of Fra-2 in v-src transformed cells: a pathway of activation of endogenous AP-1.

Chicken embryo fibroblasts (CEF) transformed with v-src were previously reported to revert to normal phenotype after the introduction of dominant-negative mutants of Fos or Jun, indicating that endogenous AP-1 activity is essential for the cellular transformation. The major changes in the expression levels of fos and jun family genes induced by v-src were the elevation of fra-2 and c-jun transcripts. We show here that extensive phosphorylation of the AP-1 component Fra-2 is a major qualitative change in v-src transformed CEF and that several Ser and Thr residues in a C-terminal region of Fra-2 (amino acids 266-323) are phosphorylated specifically. The induced kinase activity was detected at the position of 42 kDa by in gel kinase assay using the Fra-2 C-terminal region as a substrate, and it was identified as chicken ERK2. JNK1 and JNK2, other members of the MAP kinase family, were not significantly activated in v-src transformed CEF and Fra-2 was not a good substrate for JNKs. fra-2 promoter analysis indicated that this promoter activity is elevated in v-src transformed CEF via two AP-1 binding sites and CRE-like sequence. We propose that phosphorylation of Fra-2 by ERK2 converts it from an inefficient transcriptional activator to an active one and further that fra-2 expression is autoregulated in response to the phosphorylation status of its gene product.

Animals↗
PubMedDate not supplied

Altered oncogene, tumor suppressor and cell-cycle gene expression in PANC-1 cells cultured with the pleiotrophic 5-lipoxygenase inhibitor, MK886, assessed with a gene chip.

We describe a genomic response of mRNAs associated with a subset of oncogenes, tumor suppressor and cell cycle-related genes in proliferating human Panc-1 pancreatic cancer cells after 24 hours of culture with MK886, a pleotrophic 5-lipoxygenase inhibitor. Ninety-eight of these cDNAs are represented in one of the sub-arrays included in the Clontech Human cDNA Expression Array. In this initial analysis, control cells exhibited apparent widespread low levels of disparate mRNA synthesis. In cells cultured with 40 microM MK886 for 24 hr, while most expressed genes, including a number of specific proliferation-enhancing genes such as c-myc were inhibited, 19 other ones including some countervailing genes including tyrosine SRC protein kinase, cyclins B1 and D1, CDC25B phosphatase and 40s ribosomal S19, amounting to 19 percent of the cDNAs resident on the chip were up-regulated at > 1.10 experimental/control values. Therapy-induced activation of compensatory proliferative genomic responses provides an additional explanation why malignant cells can fail therapy. Among their many future uses, gene chips clearly will be an extremely powerful tool for identifying relationships between the hierarchical linear and non-linear control and implementation-related cellular events and for identifying potential molecular targets tor cancer therapy.

Blotting, Western↗
PubMed1985

Gene expression from both intronless and intron-containing Rous sarcoma virus clones is specifically inhibited by anti-sense RNA.

To distinguish the inhibitory effect of anti-sense RNA on translation from the effect on splicing, a plasmid (pLC32) was constructed from a cDNA clone of the Rous sarcoma virus (RSV) envelope gene (env) mRNA. Transcription of this plasmid results in the synthesis of RNA identical to the RSV env gene mRNA which does not require splicing to be expressed. Plasmids derived from pLC32 were also constructed in which the env gene coding sequence and 5' noncoding leader sequences were inserted in the opposite orientation relative to the RSV long terminal repeats (LTRs). pLC32 DNA transfected by the calcium phosphate coprecipitation technique efficiently rescued infectious virus from quail cells infected with an RSV mutant deleted in the env gene [R(-)Q cells], indicating that the intron sequences are dispensable in env gene expression. When the inverted constructs were cotransfected with pLC32, significantly less infectious virus was produced. The extent of the inhibition depended upon the concentration ratio of the two plasmids. The maximum inhibition (80%) occurred when the ratio of inverted constructs to pLC32 was 12:1. The inhibition is specific for the inverted orientation since cotransfection of pLC32 with several other plasmids containing viral LTRs and defective src and env genes at similar concentrations did not inhibit the production of infectious virus. In addition, the inverted constructs did not interfere with the expression of an LTR-driven chloramphenicol acetyltransferase gene. When cotransfected with a wild-type Prague A RSV DNA plasmid (pJD100), the inverted constructs also greatly inhibited expression and replication of virus in R(-)Q quail cells. These data suggest that the specific inhibition is caused by hybridization of complementary RNA transcribed from the inverted constructs to the env mRNA, thereby blocking its expression. The fact that expression of both intron-containing and intronless clones are inhibited to the same extent suggest that inhibition by anti-sense RNA from the env exon regions does not act at the level of RNA splicing.

Animals↗
PubMedDate not supplied

Loss of expression of human spectrin src homology domain binding protein 1 is associated with 10p loss in human prostatic adenocarcinoma.

The gene encoding human spectrin Src homology domain binding protein 1, or Hssh3bp1, which is a marker of macropinocytic vesicles and a potential regulator of macropinocytosis, co-localizes to a YAC containing chromosome 10p sequences at loci D10S89 and D10S111 that are frequently deleted in prostate tumors. Expression of Hssh3bp1 was evaluated at the protein level in 17 paired normal and malignant prostate tumor samples using the monoclonal antibody 2G8 to Hssh3bp1. These experiments demonstrated that 4/6 tumors (67%) with 10p deletion failed to express Hssh3bp1 protein compared to 5/11 (46%) tumors with intact 10p. Thus, loss of Hssh3bp1 expression is concordant with allelic loss of adjacent 10p sequences in human prostate tumors. In addition, two prostate tumor cell lines contain an exon skipping mutation in the Hssh3bp1 gene that leads to the abnormal splicing of the mRNA and loss of a portion of Abl tyrosine kinase SH3 domain binding site in the protein. These data are consistent with a role for Hssh3bp1 as a candidate tumor suppressor gene inactivated during prostate tumorigenesis.

Adaptor Proteins, Signal Transducing↗
PubMed1988

Control of myogenic differentiation by cellular oncogenes.

The establishment of a differentiated phenotype in skeletal muscle cells requires withdrawal from the cell cycle and termination of DNA synthesis. Myogenesis can be inhibited by serum components, purified mitogens, and transforming growth factors, but the intracellular signaling pathways utilized by these molecules are unknown. Recent studies have confirmed a role for proteins encoded by cellular proto-oncogenes in transduction of growth factor effects that lead to cell proliferation. To test the contrasting hypothesis that cellular oncogenes might also regulate tissue-specific gene expression in developing muscle cells, myoblasts have been modified by incorporation of the cognate viral oncogenes, the corresponding normal or oncogenic cellular homologs, and chimeric oncogenes, whose expression can be induced reversibly. Regulation of the endogenous cellular oncogenes also has been examined in detail. Down-regulation of c-myc is not obligatory for myogenesis; rather, inhibitory effects of myc on muscle differentiation are contingent on sustained proliferation. In contrast, activated src and ras genes block myocyte differentiation directly, through a mechanism that is independent of DNA synthesis and is rapidly reversible, resembling the effects of inhibitory growth factors. The coordinate regulation of diverse tissue-specific gene products including muscle creatine kinase, nicotinic acetylcholine receptors, sarcomeric proteins, and voltage-gated ion channels, raises the hypothesis that inhibitors such as transforming growth factor-beta and ras proteins might exert their effects through a transacting transcriptional signal shared by multiple muscle-specific genes.

Animals↗
PubMed1996

Expression and tyrosine phosphorylation of EMS1 in human breast cancer cell lines.

The EMS1 gene encodes an 80/85 kDa c-src substrate and localises with the CCND1 gene to chromosome 11q13. This locus is amplified in approximately 13% of human breast cancers. EMS1 gene amplification and expression were characterised in a panel of human breast cancer cell lines to determine at what levels expression is regulated. The degree of tyrosine phosphorylation of EMS1 protein was also determined and compared with the activity of src-family kinases. The EMS1 gene was amplified in 6 of 20 cell lines investigated: MDA-MB-134, -157, -175, -453, ZR-75-1 and MCF-7. In the MDA-MB-157 and MCF-7 cell lines, EMS1 was amplified in the absence of CCND1 gene amplification. EMS1 protein levels were increased relative to normal breast epithelial cells in 6 cell lines (ZR-75-1, MDA-MB-134, -175, 453, MCF-7 and BT-474). Of these, BT-474 is the only cell line that does not exhibit EMS1 amplification or increased EMS1 mRNA levels. EMS1 tyrosine phosphorylation was 3-fold higher in BT-474 and T-47D cells, which exhibited relatively high total src activity coupled with expression of both c-fyn and c-yes, than in MDA-MB-453 cells, which expressed only c-yes. Our results therefore demonstrate gene amplification to be the predominant mechanism underlying EMS1 over-expression in human breast cancer cell lines and identify tyrosine phosphorylation as a further level at which regulation of this protein may be perturbed.

Breast Neoplasms↗
PubMed1986

Phosphorylation and inactivation of protein phosphatase 1 by pp60v-src.

Protein phosphatase 1, one of four major protein phosphatases involved in cellular regulation, was phosphorylated in vitro by pp60v-src, the transforming gene product of Rous sarcoma virus. Phosphorylation was accompanied by a loss of protein phosphatase activity. The inactivation of protein phosphatase 1 was time-dependent and the extent of inactivation correlated closely with the stoichiometry of phosphorylation. Under optimal conditions, 0.34 +/- 0.01 mol of phosphate were incorporated per mol of protein phosphatase and the activity of the enzyme was decreased by 39 +/- 2%. The inactivation required the presence of both MgATP and pp60v-src. There was no loss of activity when adenosine 5'-[beta gamma-imido]triphosphate was used in place of ATP. Phosphorylation of protein phosphatase 1 occurred exclusively on tyrosine residues and was blocked by specific antibodies to pp60v-src. During preincubation of pp60v-src at 41 degrees C, its protein kinase activity towards casein was lost rapidly. The ability of pp60v-src to phosphorylate and inactivate protein phosphatase 1 declined in parallel with the loss of casein kinase activity. Limited chymotryptic digestion of 32P-labeled protein phosphatase 1 (Mr 37,000) resulted in its quantitative conversion to a Mr 33,000 species. Conversion to this species was accompanied by the loss of 32P-labeling and by reactivation of the protein phosphatase. When various concentrations of chymotrypsin were used in the digestion, there was a close correlation between conversion to the Mr 33,000 species and the restoration of protein phosphatase activity. pp60v-src was unable to phosphorylate or inactivate a partially proteolyzed species of protein phosphatase 1 (Mr 33,000/34,000).

Animals↗
PubMed1992

MAP kinase is constitutively activated in gip2 and src transformed rat 1a fibroblasts.

Rat 1a fibroblasts transformed by the Gi2 oncogene, gip2, exhibit a constitutively elevated mitogen-activated protein (MAP) kinase activity that correlates with enhanced tyrosine phosphorylation of the p42 MAP kinase polypeptide. The MAP kinase activity in gip2 transformed cells is 50-60% of the pertussis toxin-sensitive, thrombin-stimulated activity observed in wild-type Rat 1a cells. A similar activation of MAP kinase is observed in src but not ras or raf transformed Rat 1a cells, indicating that the persistent MAP kinase activity results from the action of the specific oncoprotein and is not the consequence of cellular transformation. The enhanced transactivation function of c-Jun characteristic of the transformed phenotype, measured using a collagenase promoter-CAT reporter gene, is observed in gip2, src, ras, and raf transformed Rat 1a cells. The regulatory networks controlled by the four transforming oncogenes therefore alter the activity of specific transcription factors, but only gip2 and src constitutively activate MAP kinase. The findings demonstrate that the catalytic activity of growth factor-regulated cytoplasmic kinases are selectively and stably activated as a consequence of specific oncogene expression.

Animals↗
PubMed1997

Severe defects in immunity and hematopoiesis caused by SHP-1 protein-tyrosine-phosphatase deficiency.

Spontaneous mouse mutations that cause severe immunodeficiency or autoimmunity are invaluable tools with which to investigate the mammalian immune system. Mutations at the 'motheaten' locus result in severe immunological dysfunction due to disruption of the structural gene encoding Src-homology 2-domain phosphatase-1 (SHP-1). This natural model for a specific protein-tyrosine-phosphatase deficiency is being widely utilized to determine the role of SHP-1 in the negative regulation of multiple signaling pathways in a number of hematopoietic lineages.

Animals↗
PubMed2003

Src family kinase-independent signal transduction and gene induction by leukemia inhibitory factor.

Members of the interleukin-6 (IL-6) family of cytokines exert their biological effects via binding to their cognate ligand-binding receptor subunit on a target cell. The subsequent recruitment of the common signal transducer glycoprotein 130 and activation of the JAK/STAT and SHP-2/Ras/mitogen-activated protein kinase (MAPK) pathways are responsible for the majority of cellular responses elicited by IL-6 cytokines. Several types of experiments suggest that the Src family of kinases (SFK) also participates in IL-6 family cytokine-mediated signaling events. SYF cells, which lack expression of SFKs Src, Yes, and Fyn, were used to determine the role of SFKs in IL-6 family cytokine signaling and gene induction. SYF and wild type (WT) control fibroblasts displayed similar activation of signaling intermediates following stimulation with leukemia inhibitory factor (LIF). LIF-stimulated tyrosine phosphorylation of SHP-2 and subsequent activation of MAPK in SYF cells were identical to that seen in LIF-stimulated WT cells. Both LIF-stimulated tyrosine phosphorylation of STAT1 and STAT3, as well as LIF-stimulated DNA binding activity of STAT-containing nuclear complexes were indistinguishable when compared in SYF and WT cells. In addition, the phosphatidylinositol 3-kinase-sensitive Akt kinase and p38 MAPK were activated by LIF in both SYF and WT cells. Furthermore, LIF-stimulated expression of c-fos, egr-1, and suppressor of cytokine signaling-3 was retained in SYF cells. The IL-6 family cytokine oncostatin M was also capable of activating MAPK, STAT3, STAT1, Akt, and p38 in both WT and SYF cells. These results demonstrate that IL-6 family cytokines can activate a full repertoire of signaling pathways and induce gene expression independent of SFKs.

Animals↗
PubMed2005

Divergent roles of c-Src in controlling platelet-derived growth factor-dependent signaling in fibroblasts.

The vast complexity of platelet-derived growth factor (PDGF)-induced downstream signaling pathways is well known, but the precise roles of critical players still elude us due to our lack of specific and temporal control over their activities. Accordingly, although Src family members are some of the better characterized effectors of PDGFbeta signaling, considerable controversy still surrounds their precise functions. To address these questions and limitations, we applied a chemical-genetic approach to study the role of c-Src at the cellular level, in defined signaling cascades; we also uncovered novel phosphorylation targets and defined its influence on transcriptional events. The spectacular control of c-Src on actin reorganization and chemotaxis was delineated by global substrate labeling and transcriptional analysis, revealing multiple cytoskeletal proteins and chemotaxis promoting genes to be under c-Src control. Additionally, this tool revealed the contrasting roles of c-Src in controlling DNA synthesis, where it transmits conflicting inputs via the phosphatidylinositol 3 kinase and Ras pathways. Finally, this study reveals a mechanism by which Src family kinases may control PDGF-mediated responses both at transcriptional and translational levels.

Actins↗
PubMed1994

Sequence specificity in the recognition of the epidermal growth factor receptor by the abl Src homology 2 domain.

The transforming activity of the abl gene product requires a functional src homology 2 (SH2) domain. An assay was developed to evaluate this function by examining binding of a bacterially-expressed abl SH2 domain to the activated EGF receptor, used as a surrogate tyrosine phosphorylated protein. The sequence specificity of this interaction has been explored with a series of point mutants of EGF receptor. Analysis of equilibrium binding reveals that substitution of Tyr1086 for Phe in the EGF receptor produced a 10-fold reduced affinity for abl SH2 domain binding as compared to the wildtype receptor. Moreover, a phosphorylated peptide modeled on the sequences surrounding Tyr1086 specifically inhibits abl SH2 binding, with an IC50 of approximately 10 microM. Evaluation of a series of additional peptides, modeled on the Tyr1086 sequence, revealed that the carboxy terminal residues directly next to the phosphotyrosine were particularly critical to this binding. Molecular modeling studies of the pTyr1086 peptide revealed the potential hydrophobic, ionic and hydrogen bonding interactions involved in the functions of the abl SH2 domain.

3T3 Cells↗
PubMed2002

Src kinase mediates phosphatidylinositol 3-kinase/Akt-dependent rapid endothelial nitric-oxide synthase activation by estrogen.

17beta-Estradiol activates endothelial nitric oxide synthase (eNOS), enhancing nitric oxide (NO) release from endothelial cells via the phosphatidylinositol 3-kinase (PI3-kinase)/Akt pathway. The upstream regulators of this pathway are unknown. We now demonstrate that 17beta-estradiol rapidly activates eNOS through Src kinase in human endothelial cells. The Src family kinase specific-inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) abrogates 17beta-estradiol- but not ionomycin-stimulated NO release. Consistent with these results, PP2 blocked 17beta-estradiol-induced Akt phosphorylation but did not inhibit NO release from cells transduced with a constitutively active Akt. PP2 abrogated 17beta-estradiol-induced activation of PI3-kinase, indicating that the PP2-inhibitable kinase is upstream of PI3-kinase and Akt. A 17beta-estradiol-induced estrogen receptor/c-Src association correlated with rapid c-Src phosphorylation. Moreover, transfection of kinase-dead c-Src inhibited 17beta-estradiol-induced Akt phosphorylation, whereas constitutively active c-Src increased basal Akt phosphorylation. Estrogen stimulation of murine embryonic fibroblasts with homozygous deletions of the c-src, fyn, and yes genes failed to induce Akt phosphorylation, whereas cells maintaining c-Src expression demonstrated estrogen-induced Akt activation. Estrogen rapidly activated c-Src inducing an estrogen receptor, c-Src, and P85 (regulatory subunit of PI3-kinase) complex formation. This complex formation results in the successive activation of PI3-kinase, Akt, and eNOS with consequent enhanced NO release, implicating c-Src as a critical upstream regulator of the estrogen-stimulated PI3-kinase/Akt/eNOS pathway.

Adenoviridae↗
PubMed2005

[Frontier of mycobacterium research--host vs. mycobacterium].

During the past decade, we have observed advance in tuberculosis research including novel vaccines, innate immunity (TLR), SNIP analysis and molecular mechanism of drug resistance. Worldwide genome project enabled the whole genome sequence of host resistant against tuberculosis as well as the whole genome sequence of M. tuberculosis H37Rv. DNA technology has also provided a great impact on the development of novel vaccine against TB. In this symposium, we have invited leading researchers in the field of the frontier study of Mycobacterium research in order to provide general overview of the cutting edge of frontier research. Molecular mechanism of drug resistance of M. tuberculosis has been clarified. On the other hand, molecular mechanism of host-defence (insusceptibility of host) against M. tuberculosis has not yet elucidated. Dr. Taro Shirakawa (Kyoto University) reviewed the susceptibility genes of host in TB infection and presented candidate genes associated with multi-drug resistant tuberculosis. Dr. Naoto Keicho (International Medical Center of Japan) tried to identify host genetic factors involved in susceptibility to pulmonary Mycobacterium avium complex (MAC) infection by candidate gene approach and genome-wide approach. In Japan, Dr. Masaji Okada (National Hospital Organization Kinki-Chuo Chest Medical Center) has been engaged actively in the development of new tuberculosis vaccines (HVJ-liposome/Hsp65 DNA + IL-12 DNA vaccine and recombinant 72f BCG vaccine). He showed basic strategy for construction of new candidate vaccines and also showed significant efficacy on the protection of tuberculosis infection using cynomolgus monkeys, which are very similar to human tuberculosis. Dr. Hatsumi Taniguchi (University of Occupational and Environmental Health) presented that M. tuberculosis mIHF and the neighbor genes went into a dormacy-like state of M. smegmatis in J774 macrophage cells. This study might provide a weapon for elucidating the mechanism of dormacy of M. tuberculosis and the development of novel therapy. Dr. Chiyoji Abe (Nippon Becton Dickinson Co.) reviewed the molecular basis of the resistance to anti-tuberculosis drugs. Most cases of resistance are related to simple nucleotide substitutions rather than to acquisition of new elements. Dr. Kiyoshi Takeda (Kyushu University) showed interesting finding. He analyzed whether Toll-like receptor (TLR)-mediated activation of innate immunity in host defense against mycobacterial infection. MyD88/TRIF double defi-indicating that innate immunity is involved in anti-mycobacterial infection. (1) SNP (single nucleotide polymorphism) analysis in association with Mycobacterium tuberculosis: Taro SHIRAKAWA (Department of Health Promotion & Human Behavior, Kyoto University Medical School, and RIKEN SRC Center) Candidate gene approach was made on 18 SNPs in 11 genes in association with M. tuberculosis. Patients with multi-drug resistance against M. tuberculosis are also subjected. SNPs in NRAMP1 gene were associated with the disease, and drug resistance, its mechanisms remain unknown. (2) Search for genes susceptible to pulmonary Mycobacterium avium complex infection: Naoto KEICHO (Department of Respiratory Diseases, Research Institute, International Medical Center of Japan) Interaction among pathogens and host factors is important for development of infectious diseases. We are trying to identify host genetic factors involved in susceptibility to nonimmunocompromized pulmonary Mycobacterium avium complex (MAC) infection by candidate gene approach and genome-wide approach. Elucidation of functional significance of susceptibility gene polymorphisms will lead to a new strategy for control and prevention of the disease. (3) T cell immunity against Tuberculosis in host and the establishment of novel vaccine: Masaji OKADA (Clinical Research Center, National Hospital Organization Kinki-Chuo Chest Medical Center) T cell (CTL, Th1) immunity including granulysin play an important role in host defense against tuberculosis (TB) in human. Patients with TB or Multi-drug resistant TB showed suppression of all these immunities. HVJ-liposome/Hsp65 DNA + IL-12 DNA vaccination was 100 fold more efficient than BCG on the elimination of Mycobacterium tuberculosis (M.TB) in the BALB/c mice. Cytotoxic T cells activity against M. TB was augmented. By using these new vaccines (Hsp 65 DNA + IL-12 DNA, recombinant 72f BCG) and the cynomolgus monkey models which are very similar to human tuberculosis, the prophylactic effect of vaccines was observed. Thus, these novel vaccines should provide a useful tool for the prevention of human TB infection. (4) Mycobacterium tuberculosis mIHF and the neighbor genes go into a dormancy-like state of M. smegmatis J15CS in J774 cells: Hatsumi TANIGUCHI (Department of Microbiology, School of Medicine, University of Occupational and Environmental Health) Mycobacterium smegmatis J15CS transformants harboring the mIHF gene or the mIHF-gmk-Rv1390 genes showed no difference in in vitro growth and acid-fastness. However, transformants harboring mIHF-gmk-Rv1390 formed short-rod cell morphology and decreased acid-fastness in the mouse macrophage-like cell line J774 compared to those of the other transformants, and the nuclei of the infected J774 cells also changed. Nevertheless, the colony forming units were similar. These indicate that mIHF and the neighbor genes of M. tuberculosis might regulate a growth of mycobacteria in macrophages. (5) Molecular basis of the resistance to anti-tuberculosis drugs: Chiyoji ABE (Nippon Becton Dickinson Company, Ltd.) Considerable progress has been made toward understanding the molecular basis of the resistance to anti-tuberculosis drugs. Most cases of resistance are related usually to simple nucleotide substitutions rather than to acquisition of new elements. Multi-drug resistant isolates of Mycobacterium tuberculosis arise a consequence of sequential accumulation of mutation conferring resistance to single therapeutic agents. The basis of resistance is not able to be explained yet in a substantial percentage of strains for other anti-tuberculosis drugs than rifampin and pyrazinamide. Further studies are required to fully understand the molecular mechanisms of resistance. (6) Toll-like receptors in anti-mycobacterial immune responses: Kiyoshi TAKEDA (Department of Molecular Genetics, Medical Institute of Bioregulation, Kyushu University) Toll-like receptors (TLRs) play an essential role in the recognition of specific patterns of microbial components. TLRs mediate activation of innate immunity and further development of antigen-specific adaptive immunity. In TLR signaling pathways, Toll/IL-1 receptor (TIR) domain-containing adaptors, such as MyD88, TIRAP, TRIF, and TRAM, have been shown to play pivotal roles. Thus, the molecular mechanisms for TLR-mediated activation of innate immunity have been largely understood. We analyzed whether TLR-mediated activation of innate immunity is involved in host defense against mycobacterial infection. MyD88/TRIF double deficient mice, in which TLR-dependent activation of innate immunity is abolished, showed high sensitivity to mycobacterial infection, indicating that innate immunity is critically involved in anti-mycobacterial responses.

Antitubercular Agents↗
PubMed2000

Isolation of transformation suppressor genes by cDNA subtraction: lumican suppresses transformation induced by v-src and v-K-ras.

We have reported that suppressive factors for transformation by viral oncogenes are expressed in primary rat embryo fibroblasts (REFs). To identify such transformation suppressor genes, we prepared a subtracted cDNA library by using REFs and a rat normal fibroblast cell line, F2408, and isolated 30 different cDNA clones whose mRNA expression was markedly reduced in F2408 cells relative to that in REFs. We referred to these as TRIF (transcript reduced in F2408) clones. Among these genes, we initially tested the suppressor activity for transformation on three TRIF genes, TRIF1 (neuronatin), TRIF2 (heparin-binding growth-associated molecule), and TRIF3 (lumican) by focus formation assay and found that lumican inhibited focus formation induced by activated H-ras in F2408 cells. Colony formation in soft agar induced by v-K-ras or v-src was also suppressed in F2408 clones stably expressing exogenous lumican without disturbing cell proliferation. Tumorigenicity in nude mice induced by these oncogenes was also suppressed in these lumican-expressing clones. These results indicate that lumican has the ability to suppress transformation by v-src and v-K-ras.

Animals↗
PubMed2003

Pp60c-src mediates ERK activation/nuclear localization and PAI-1 gene expression in response to cellular deformation.

Release of transcellular tension upon disruption of actin stress fibers with cytochalasin D (CD) and associated changes in cell morphology are reflected in the rapid transcription of "deformation-responsive" genes. For certain genes (e.g., urokinase plasminogen activator and its type-1 inhibitor PAI-1), de novo mRNA synthesis appears to require cell shape-dependent activation of the MAP kinases ERK1/2. ERK activation in response to microfilament disruption was inhibited completely by the broad-spectrum tyrosine kinase inhibitor genistein and the relatively src-kinase selective compound PP1. Such inhibitor sensitivity profiles suggested that src-family members, likely pp60(c-src), were important upstream elements in deformation-related ERK activation. pp60(c-src) kinase activity was elevated fourfold within 15 min after CD addition to quiescent R22 smooth muscle cells and declined quickly thereafter. CD-induced increases in the phosphorylation levels of both pp60(c-src) and IgG heavy chain (a substrate target in the coupled immunoprecipitation/in vitro pp60(c-src) kinase assay) were ablated completely by pretreatment with the src-type kinase inhibitor PP1. Prior PP1 exposure similarly repressed CD-stimulated PAI-1 transcript accumulation. Consistent with the pharmacologic findings, transfection of a dominant-negative pp60(c-src) expression construct (DN-Src) effectively suppressed (in a concentration-dependent manner) CD-induced PAI-1 synthesis in R22 cells. To more specifically address the potential involvement of src kinases in CD-initiated ERK mobilization, R22 cells were transiently co-transfected with DN-Src and Myc-tagged ERK2 expression constructs, serum-deprived then stimulated with CD. The effect of DN-Src expression on endogenous ERK1/2 activation and nuclear translocation was assessed in separate experiments. The phosphorylation levels of both exogenous (Myc-ERK2) and endogenous ERK1/2 targets was significantly reduced by DN-Src; nuclear accumulation of pERK1/2 was completely inhibited. These data indicate that pp60(c-src) is a critical upstream activator of the ERK cascade leading to PAI-1 transcription in response to cellular deformation stimuli.

Actin Cytoskeleton↗
PubMed1989

Localization of the SRC oncogene to chromosome band 20q11.2 and loss of this gene with deletion (20q) in two leukemic patients.

In situ hybridization of the pHul-c-src probe to metaphase cells from three normal donors and two leukemic patients showed significant labeling in the proximal region of the long arm of chromosome 20q, with modal peaks of grains consistently at band 20q11.2. A secondary peak of grains was detected in the region 20q13.2-qter, the localization of SRC suggested by previous in situ studies. The exact localization of SRC is important for understanding the del(20q) chromosomal abnormality in myeloid neoplasias. Chromosome in situ hybridization and genomic studies showed loss of one allele of SRC in two patients with the deletion (20q). These results differ from previously published findings and suggest heterogeneity of the breakpoint at 20q11.2 in interstitial deletions of 20q, which characterize myeloid disorders.

Adult↗