Mutants of Arabidopsis as tools to understand the regulation of phenylpropanoid pathway and UVB protection mechanisms.

Plants accumulate certain phenylpropanoid compounds in the vacuoles of their epidermal and subepidermal cell layers thereby protecting the underlying tissue against UVB-induced damage. However, a number of mutants of Arabidopsis thaliana are known that fail to synthesize these protective pigments, thereby allowing harmful UVB radiation to penetrate into their dermal layers. Study of several of these nonlethal mutants, defective in various aspects of flavonoid and lignin biosynthesis, has led to a better understanding of the coordinate regulation and expression of important genes as well as of mechanisms involved in plant defense against UVB radiation. The characteristics of the various phenylpropanoid mutants of Arabidopsis, viz. flavonoid mutants (banyuls [ban]; increased chalcone synthase expression 1 [icx1]; transparent testa [tt] and ultraviolet sensitive [uvs]) and hydroxycinnamic acid ester mutants (ferulic acid hydroxylase 1 [fah1] and sinapoylglucose accumulator 1 [sng1]) are discussed in detail. We have briefly touched upon, wherever relevant, the unique aspects in other plant species too.

Involvement of amino acids 361 to 364 of human topoisomerase I in camptothecin resistance and enzyme catalysis.

Camptothecins are antineoplastic drugs that specifically target the enzyme DNA topoisomerase I. Prior work has identified a human topoisomerase I mutation, F361S, that confers resistance to camptothecin. We now demonstrate that substitutions in the 361-364 region can alter DNA cleavage/ligation by the enzyme. The defective catalysis exhibited by certain mutants likely relates to an impaired interaction with DNA, since these enzymes are more sensitive to the inhibitory effects of DNA binding ligands. Moreover, studies with peptides and fusion proteins suggest that the 361-364 region may bind DNA directly. The finding that the 361-364 region is involved in both enzyme catalysis and camptothecin resistance suggests that this region is part of the active site of human topoisomerase I and that camptothecin may interact with the enzyme at this site.

Identification of a nucleolin binding site in human topoisomerase I.

DNA topoisomerase I (topo I) is involved in the regulation of DNA supercoiling, gene transcription, and rDNA recombination. However, little is known about interactions between topo I and other nuclear proteins. We used affinity chromatography with a topo I fusion protein to screen U-937 leukemic cell extracts and have identified nucleolin as a topo I-binding protein. Coimmunoprecipitation and other studies demonstrate that the interaction between topo I and nucleolin is direct. Furthermore, deletion analyses have identified the 166-210-amino acid region of topo I as sufficient for the interaction with nucleolin. Since nucleolin has been implicated in nuclear transport and in a variety of transcriptional processes, the interaction with topo I may relate to the cellular localization of topo I or to the known role of this topoisomerase in transcription.