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C J Lyon

Publications and source records attributed to C J Lyon.

At least 19 recordsLinked to original sources
PubMed2000

The C. elegans apoptotic nuclease NUC-1 is related in sequence and activity to mammalian DNase II.

The Caenorhabditis elegans nuc-1 gene has previously been implicated in programmed cell death due to the presence of persistent undegraded apoptotic DNA in nuc-1 mutant animals. In this report, we describe the cloning and characterization of nuc-1, which encodes an acidic nuclease with significant sequence similarity to mammalian DNase II. Database searches performed with human DNase II protein sequence revealed a significant similarity with the predicted C. elegans C07B5.5 ORF. Subsequent analysis of crude C. elegans protein extracts revealed that wild-type animals contained a potent endonuclease activity with a cleavage preference similar to DNase II, while nuc-1 mutant worms demonstrated a marked reduction in this nuclease activity. Sequence analysis of C07B5.5 DNA and mRNA also revealed that nuc-1(e1392), but not wild-type animals contained a nonsense mutation within the CO7B5.5 coding region. Furthermore, nuc-1 transgenic lines carrying the wild-type C07B5.5 locus demonstrated a complete complementation of the nuc-1 mutant phenotype. Our results therefore provide compelling evidence that the C07B5.5 gene encodes the NUC-1 apoptotic nuclease and that this nuclease is related in sequence and activity to DNase II.

Amino Acid Sequence↗
PubMed1998

Characterization of a novel DNA binding domain within the amino-terminal region of the RAG-1 protein.

Rag-1 and Rag-2 are the critical components of the V-(D)-J recombinase required for site-specific recombination of the antigen receptor genes. In this study, we have examined the ability of recombinant (r) Rag-1 and Rag-2 to bind the recombination signal sequences (RSS) and have determined that rRag-1, but not rRag-2, is able to directly bind DNA. rRAG-1 DNA binding activity was found to reside within a novel amino-terminal arginine-rich (RR) domain with partial homology to a variety of nucleic acid binding domains. Although the RR-domain did not demonstrate RSS-specificity, this DNA binding domain may stabilize the interaction of RAG-1 with, or increase the affinity for, the V-(D)-J recombination signals.

Amino Acid Sequence↗
PubMed1997

Regulation of the RAG-1 promoter by the NF-Y transcription factor.

We have cloned the RAG-1 promoter region and have determined that almost all detectable promoter activity resides within a 208-bp fragment. Sequence analysis of this promoter region has identified potential recognition motifs for a number of lymphocyte-restricted and ubiquitous transcription factors. Subsequent assays have revealed that the NF-Y transcription factor interacts with a CCAAT site within the RAG-1 promoter and appears to play an important role in the positive transcriptional regulation of this gene.

3T3 Cells↗
PubMed1997

Purification and characterization of the immunoglobulin switch sequence-specific endonuclease (Endo-SR) from bovine spleen.

Mature B lymphocytes are able to specifically alter their Ig isotype expression in response to extracellular stimuli via a highly regulated, deletional recombination process called isotype switch recombination. Switch recombination breakpoints predominantly map to large (1-10 kb), G-rich and highly repetitive switch regions that are located directly upstream of immunoglobulin heavy-chain constant region genes. Switch region repeat structures vary considerably both within and between species, but all switch regions contain disproportionate numbers of two pentamer motifs, TGGGN and TGAGC, that are found at or directly adjacent to most analysed switch junctions. We have recently identified an endonuclease activity, Endo SR, that preferentially cleaves TGGGN and TGAGC switch motifs. We have purified the bovine endonuclease activity to homogeneity and have identified a protein with a molecular weight of approximately 32,000 that directly correlates with enzyme activity. As discussed in this report, we have found that murine and bovine Endo-SR are preferentially enriched in lymphoid tissue nuclear extracts and that both enzymes demonstrate highly similar physical and biochemical characteristics. However, each enzyme demonstrates related but distinctive specificities for consensus and degenerate TGGGN and TGAGC switch pentamer motifs.

Animals↗
PubMed1996

Characterization of an endonuclease activity which preferentially cleaves the G-rich immunoglobulin switch repeat sequences.

B lymphocytes can alter selectively their immunoglobulin (Ig) isotype expressed by deletional rearrangement of the first active immunoglobulin heavy-chain (IgH) constant region (C mu) gene with one of six other constant region genes. Recombination breakpoints occur within highly repetitive "switch" (S) regions located upstream of each IgH constant region gene except C delta. Analysis of rearranged switch DNA junctions has not detected a consensus sequence, although the predominance of two pentamer motifs (TGGGG and TGAGC) at or near these breakpoints and throughout all murine S region sequences has led to their advocacy as the S recombination signals. In this paper, we describe the characterization and partial purification of a lymphoid-specific endo-nuclease activity which cleaves preferentially murine S region DNA. Enzyme activity selectively produced single- and double-stranded breaks at TGAGC and TGGG motifs within murine S mu and S alpha DNA. Rare cryptic cleavage sites were detected also within non-switch sequences, although cleavage intensities at these sites were reduced greatly, relative to consensus S region cleavages. Analogous activity was found in murine tissue extracts, although among the tissues assayed only spleen and thymus contained detectable activity. Subsequent biochemical characterization of this activity demonstrated that the responsible enzyme (Endo-SR) represented a previously unreported tissue-specific mammalian endonuclease. Endo-SR-specific activity could be enhanced by addition of Mg2+ or Ca2+ and inhibited by addition of Zn2+. Maximal specific activity was detected at pH 5.5 and sharply declined within +/- 0.5 pH units. In view of this enzyme's sequence- and tissue-specificity, we propose that Endo-SR is a strong candidate for an endonuclease activity associated with the switch recombination process.

Animals↗
PubMed1970

Choice of rotation rate for the horizontal clinostat.

A series of nine rates of rotation of the clinostat were tested to determine optimal and acceptable conditions for simulating weightlessness in plants. Young seedlings of wheat (Triticum aestivum L.) developed roots and coleoptiles of equal lengths and with the same orientation angles over a range of rotation rates from 0.25 to 480 minutes per revolution. Rates from 0.25 to 3 minutes per revolution provided for maximal epinastic curvatures of leaves and branches of Coleus blumei Benth. except for a reduced mean curvature of branches at 0.25 minute per revolution, due probably to physical disturbances in their growth. Smaller epinastic curvatures developed in both leaves and branches rotated at 15 minutes per revolution or more slowly. Indoleacetic acid-2-(14)C was used for measurements of extractable radioactivity in determining the reason for smaller curvatures of Coleus branches and tomato (Lycopersicon esculentum Mill.) leaves at the rotation rate of 60 minutes per revolution than at 1 minute per revolution. The cause was determined to be movement of some auxin from the upper into the lower side of a plagiotropic leaf or branch under the transient influence of gravity during rotation on the slower clinostat. A rotation period of 1 to 3 minutes was found to be acceptable for most plants.

Journal Article↗
PubMed1967

Rotation axes for analysis of gravity effects on plant organs.

Epinastic curvatures of branches of Coleus blumei Benth. and the growth pattern of wheat (Triticum aestivum L.) seedlings on clinostats were used for bioassay of rotation methods for preventing growth responses to gravity. Tumbling a plant end over end was found to be just as effective as rotation about its horizontal axis. The results support the reliability of data from experiments in which an entire plant is rotated about a single horizontal line with only part or none of its immature tissues in horizontal orientation to gravity.

Journal Article↗
PubMed1966

Orientation of wheat seedling organs in relation to gravity.

Seedlings of wheat (Triticum aestivum L.) were grown in special holders that permitted the coleoptile and early roots to develop in moist air. The orientation of the organs of seedlings erect to gravity was compared with that of organs produced on a horizontal clinostat. Orientation was described by the angular position of each organ tip with reference to the axis of the embryo. Comparative tests were also made with barley, rye, and oat seedlings.The coleoptile of all species developed curvatures in 3 dimensions when geotropic responses were eliminated. The primary root was not precise in its positive geotropism. Seedlings grew on clinostats with much greater variations in the lateral orientation of the central root and with a tendency for it to curve away from the endosperm to a greater degree than in erect seedlings.The symmetry of root system in wheat was found to depend on a specific mechanism. Under the influence of gravity the earliest lateral roots were oriented in a plane at characteristic angles of about 57.5 degrees with the ideal primary root. The corresponding angles for lateral roots growing on clinostats were greater by about 47.5 degrees as a result of epinasty not previously reported in roots. This force also appeared to be active in the seminal roots of barley and rye but not of oats.The curvatures in coleoptiles grown without the directional effects of gravity correspond to the results of growth imbalance in Coleus stems in the absence of lateral transport of their auxin by gravity. Root epinasty appears to be based on auxin imbalance. Curvatures in the primary root are also interpreted as results of asymmetrical distribution of growth hormone.

Journal Article↗