ABSTRACT
A human thiamine pyrophosphokinase cDNA clone (hTPK1) was isolated and sequenced. When the intact hTPK1 open reading frame was expressed as a histidine-tag fusion protein in Escherichia coli, marked enzyme activity was detected in the bacterial cells. The hTPK1 mRNA was widely expressed in various human tissues at a very low level, and the mRNA content in cultured fibroblasts was unaffected by the thiamine concentration of the medium. The chromosome localization of the hTPK1 gene was assigned to 7q34.
Subject(s)
DNA, Complementary/genetics , Thiamin Pyrophosphokinase/genetics , Amino Acid Sequence , Anemia, Megaloblastic/enzymology , Anemia, Megaloblastic/genetics , Blotting, Northern , Chromosomes, Human, Pair 7 , Cloning, Molecular , DNA, Complementary/chemistry , DNA, Complementary/isolation & purification , Escherichia coli/genetics , Escherichia coli/metabolism , Fibroblasts/enzymology , Humans , In Situ Hybridization, Fluorescence , Kidney/enzymology , Leukocytes/enzymology , Molecular Sequence Data , Myocardium/enzymology , RNA, Messenger/analysis , RNA, Messenger/metabolism , Sequence Alignment , Thiamin Pyrophosphokinase/biosynthesis , Thiamin Pyrophosphokinase/chemistryABSTRACT
We isolated cDNAs encoding a novel RING finger protein (LUN), the mRNAs of which were expressed at high levels in the lung. In situ hybridization revealed that LUN mRNAs were expressed in the alveolar epithelium of the lung. The LUN gene locus was assigned to chromosome 9p21, which contains candidate tumor suppressor genes associated with loss of heterozygosity in more than 86% of small cell lung cancers. We clarified that LUN is localized to the nucleus and reveals Zn(2+)-dependent DNA binding activity. The region from amino acids 51 to 374 of LUN is responsible for DNA binding. Furthermore, we identified a novel palindromic binding consensus (5'-TCCCAGCACTTTGGGA-3') for the LUN binding. Interestingly, this LUN binding palindromic sequence is found in the upstream transcriptional regulatory region of the E-cadherin gene and two intervening regions of the talin gene. Our results suggested that LUN might be an important trans-acting transcriptional regulator for lung cancer-associated genes including E-cadherin and talin genes.
Subject(s)
Carrier Proteins , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/genetics , Lung/metabolism , Neoplasm Proteins , Nuclear Proteins , Trans-Activators/chemistry , Trans-Activators/genetics , Transcription Factors , Amino Acid Sequence , Base Sequence , Binding Sites , Blotting, Northern , Cadherins/metabolism , Cations , Cell Nucleus/metabolism , Chromosome Mapping , Chromosomes, Human, Pair 9 , Cloning, Molecular , DNA/metabolism , DNA, Complementary/metabolism , DNA-Binding Proteins/metabolism , Epithelium/metabolism , Forkhead Transcription Factors , Gene Library , Glutathione Transferase/metabolism , HeLa Cells , Humans , In Situ Hybridization , In Situ Hybridization, Fluorescence , Loss of Heterozygosity , Molecular Sequence Data , Protein Binding , Protein Structure, Tertiary , Pulmonary Alveoli/metabolism , RNA, Messenger/metabolism , Recombinant Fusion Proteins/metabolism , Sequence Homology, Amino Acid , Sequence Homology, Nucleic Acid , Tissue Distribution , Trans-Activators/biosynthesis , Transcription, Genetic , Transfection , Ubiquitin-Protein Ligases , Zinc/metabolismABSTRACT
Cell adhesion kinase beta (CAK beta) is the second protein-tyrosine kinase of the focal adhesion kinase subfamily with large N- and C-domains in addition to the central kinase domain. The cDNA of the human CAK beta has been cloned and used as a probe for the assignment of this gene by fluorescence in situ hybridization. CAK beta is sublocalized on chromosome 8p21.1, a locus frequently involved in allelic losses in colorectal cancers and prostate carcinomas.