Characterization of Gmhsp26-A, a stress gene encoding a divergent heat shock protein of soybean: heavy-metal-induced inhibition of intron processing.

Abstract

We determined the DNA sequence and mapped the corresponding transcripts of a genomic clone containing the Gmhsp26-A gene of soybean. This gene is homologous to the previously characterized cDNA clone pCE54 (E. Czarnecka, L. Edelman, F. Schoffl, and J. L. Key, Plant Mol. Biol. 3:45-58, 1984) and is expressed in response to a wide variety of physiological stresses including heat shock (HS). Si nuclease mapping of transcripts and a comparison of the cDNA sequence with the genomic sequence indicated the presence of a single intron of 388 base pairs. Intron removal from pre-mRNA was preferentially inhibited by treatment of soybean seedlings with either CdCl2 or CuS04. Analysis of the 5' termini of transcripts indicated the presence of one major and at least two minor start sites. In each case, initiation occurred 27 to 30 base pairs downstream from a TATA-like motif, and thus each initiation site appears to be promoted by the activity of a separate subpromoter. The three subpromoters are all associated with sequences showing low homology to the HS consensus element of Drosophila melanogaster HS genes and are differentially induced in response to various stresses. Within the carboxyl-terminal half of the protein, hydropathy analysis of the deduced amino acid sequence indicated a high degree of relatedness to the small HS proteins. A comparison of the primary amino acid sequence of hsp26-A with sequences of the small HS proteins suggested that this stress protein is highly diverged and may therefore be specialized for stress adaptation in soybean.The heat shock (HS) response is found in a wide range of organisms and is typically triggered when cells are exposed to hyperthermia or a variety of other stresses (1). In soybean the low-molecular-mass small heat shock proteins (hsps) represent the most abundant class of proteins expressed during HS (23) and include 30 to 50 proteins which range in molecular mass from 15 to 27 kilodaltons (kDa). The lowmolecular-mass family of hsps is more diverse than the high-molecular-mass hsps (68, 70, and 83 kDa) and can be subdivided into at least three subgroups on the basis of size and conservation of amino acid sequences (22,30,40,41).Analysis of hybridization of soybean seedling RNA to cloned HS-specific cDNAs indicates that most representatives of the low-molecular-mass hsps show various degrees of induction by numerous stress agents, but generally at levels much lower than with HS (8; L. Edelman, E. Czarnecka, and J.

Isolation and characterization of an auxin-inducible glutathione S-transferase gene of Arabidopsis thaliana

“…The protein encoded by the AtlO3-1a gene clearly belongs to the family of type III GSTs and shows homology with other members of this family (NT103, G M H S P 2 6 from soybean [2] and PRP1 from potato [ 16]) ranging from 46 to 48~o. This homology was significantly higher than that with GSTs that were identified in Arabidopsis so far, where amino acid identity is only 23~o to 27~o ( Table 1).…”

“…Like the Ntl03 [ 18], as well as the prpl gene [16] and the Gmhsp26-A gene [2] the gene con- Table 1. Percentage of amino acid identity of the protein encoded by the AtlO3-1a gene compared to the proteins encoded by the tobacco Nt103, [ 19] Nt107 and Nt114 [ 12] genes, the Gmhsp26-A gene of soybean [2] and the prpl gene of potato [ 16]. Also proteins encoded by different GSTs of arabidopsis were compared to the AT103-1A protein.…”

The Role of Octadecanoids and Functional Mimics in Soybean Defense Responses

“…Total RNA of treated soybean cells was prepared according to Schröder et al (1979) and 15 µg per lane were blotted onto Biodyne-A membranes (Pall, Dreieich, Germany). Hybridization probes used were: 3 kbp soybean actin gene, bean (Phaseolus vulgaris) chalcone synthase 1 (CHS) cDNA (Ryder et al, 1984), soybean reductase co-acting with CHS (CHR) partial cDNA clone p1 (Welle et al, 1991), soybean flavonoid 6-hydroxylase (F6H) gene-specific cDNA fragment Latunde-Dada et al, 2001), soybean 3,9-dihydroxypterocarpane 6a-hydroxylase (D6aH) gene-specific probe generated by PCR from the cDNA , soybean Gmhsp-26A glutathione S-transferase (GST) cDNA (Czarnecka et al, 1988, kindly provided by R. Tenhaken, University of Kaiserslautern, Germany), and partial soybean lipoxygenase 4 (LOX4, corresponding to swissprot accession P38417) cDNA (J. Fliegmann, unpublished).…”

Expression and RNA Splicing of the Maize Glutathione S-Transferase Bronze2 Gene Is Regulated by Cadmium and Other Stresses

“…Recently, it has been recognized that a number of "stress" genes of previously unidentified functions make u p a plant GST subclass (for reviews, see Droog et al, 1993;Marrs, 1996). These type I11 GSTs are induced by numerous environmental stress conditions, including those arising from pathogen attack (Taylor et al, 1990), oxidative stress (Wingate et al, 1988;Levine et al, 1994;Tenhaken et al, 1995), and heavy metal toxicity (Czarnecka et al, 1988;Hagen et al, 1988;Wingate et al, 1988;Mauch and Dudler, 1993). The type I11 GSTs also play a role in the cellular response to both natural and synthetic auxins (Droog et al, 1993;Hahn and Strittmatter, 1994;Ulmasov et al, 1994Ulmasov et al, , 1995Droog, 1995;Droog et al, 1995).…”