PubMed Health⌕ Search

Biomedical subjects

K R Lynch

Publications and source records attributed to K R Lynch.

101 records · Page 6Linked to original sources
PubMed1982

Modifications in alpha 2u globulin gene structure, transcription, and mRNA translation in hepatomas.

alpha 2u globulin synthesis ceases when a liver cell becomes malignant. We have compared the structure and transcription of the alpha 2u globulin genes in Morris hepatomas 5123D and 7793 with that of normal hepatic genes using alpha 2u globulin cDNA as a hybridization probe. No alpha 2u globulin mRNA was detected in hepatoma 7793 by cell-free translation, Northern blot, or R0t analysis. In hepatoma 5123D, however, a small number of alpha 2u globulin RNA sequences were detected by Northern blot and R0t analysis, but this RNA was not detectably translationally active either in vivo or in an in vitro cell-free translational system. No structural differences between normal liver and the hepatoma alpha 2u globulin genes were observed by Southern blot analysis of genomic DNA digested with restriction endonucleases Pst I, Bam HI, or Ava I. However, a general demethylation of cytosine residues in the alpha 2u globulin genes of hepatomas has been found using the restriction enzyme Hha I and the isoschizomeric pair of restriction enzymes Msp I and Hpa II. Therefore, loss of the alpha 2u globulin phenotype in these hepatomas is accompanied by extensive demethylation of DNA sequences within and/or immediately flanking the alpha 2u globulin genes.

Alpha-Globulins↗
PubMed1982

The role of growth hormone in alpha 2u globulin synthesis: a reexamination.

Hypophysectomy of adult male rats abolishes alpha 2u globulin synthesis; synthesis can be fully restored by daily administration of androgen, thyroid hormone, glucocorticoid and growth hormone for 12 days. It has been previously reported that growth hormone is not required to maintain alpha 2u globulin mRNA levels, and that growth hormone functions only translationally. We have reexamined the role of growth hormone in alpha 2u globulin synthesis using a cloned alpha 2u globulin cDNA probe. Measurement of alpha 2u globulin mRNA levels by hybridization and cell-free translation, and of alpha 2u globulin synthesis by pulse-labeling in vivo, demonstrates that growth hormone is required to maintain full, steady state hepatic levels of both alpha 2u globulin and its mRNA. Furthermore, the small amount of hepatic alpha 2u globulin mRNA that accumulates in the absence of growth hormone is efficiently translated in vivo. Thus we now find no evidence for growth-hormone-mediated translational control. We have also found that rats hypophysectomized prepubescently respond to the multihormonal restoration therapy to only 10% of the alpha 2u globulin mRNA levels observed in rats hypophysectomized as adults. This finding suggests that some unidentified pituitary factor(s) is required during puberty to potentiate normal alpha 2u globulin gene expression.

Alpha-Globulins↗
PubMed1981

Covalent modification and repressed transcription of a gene in hepatoma cells.

When liver cells undergo malignant transformation, certain genes cease being expressed. We have studied the structure of one such gene, whose protein product we have designated hepatic protein 22 (hp22), which is not expressed in the two Morris hepatomas studied. We have prepared a chimeric clone of pBR322 containing cDNA sequences complementary to mRNA coding for this protein. By using this cloned cDNA, we have examined changes in expression of this gene and changes in the restriction pattern of the DNA isolated from normal liver and these hepatomas. In both hepatomas, studies using the isoschizomeric pair of restriction enzymes Msp I and Hpa II have indicated hypermethylation of a cytosine residue within or proximal to the hp22 gene. Other differences in the restriction pattern between normal liver and hepatoma DNA were also detected with EcoRI and Ava I. Thus, in the nontranscribed form of this gene, the DNA has undergone covalent modification, distinguishing these two hepatomas from each other and from normal liver.

Animals↗
PubMed1981

Cloning and sequence of several alpha 2u-globulin cDNAs.

We describe a simple cloning procedure for alpha 2u-globulin that requires neither enrichment of mRNA for cloning nor purification of a specific probe for screening recombinant colonies. Total adult male liver poly(A)+RNA was used as template for cloning, and the subsequent recombinant colonies were screened by comparing hybridization to radioactive cDNA probes prepared from hepatic male and female mRNA, respectively. Almost all of the selected "male-specific" clones were later shown to contain alpha 2u-globulin sequences. This cloned alpha 2u-globulin cDNA has been shown to specifically hybridize to male rat liver RNA, which, when isolated and translated in vitro, codes for a 21,000-dalton protein (pro-alpha 2u-globulin) immunologically identical to alpha 2u-globulin. When translation occurs in the presence of pancreatic microsomes this in vitro synthesized pro-alpha 2u-globulin is processed to the 19,000-dalton mature form of alpha 2u-globulin. The nucleotide sequence of the alpha 2u-globulin cDNA has been determined, thus elucidating the complete amino acid sequence of alpha 2u-globulin and most of the hydrophobic "leader" sequence of pro-alpha 2u-globulin. The amino acid sequence deduced from the cDNA is in agreement with the partial sequence that we previously determined by sequential Edman degradation of the purified protein. alpha 2u-Globulin cDNA clones contain within the 3'-untranslated region one or both of the two putative polyadenylylation/transcription termination sites (A-A-T-A-A-A and A-A-T-T-A-A-A). Either of these can be used, generating alpha 2u-globulin mRNA species of two lengths. A codon usage analysis of the cDNA showed that, although all six leucine codons are used for the 14 leucine residues in mature alpha 2u-globulin, the seven leucines in the partial leader sequence reported are all encoded by the same codon, CTG. The primary amino acid sequence contains a unique Asn-Gly-Ser sequence, likely to be in beta-turn conformation, as the probable site of glycosylation for this glycoprotein.

Alpha-Globulins↗