Characterization of two Maize HSP90 heat shock protein genes: Expression during heat shock, embryogenesis, and pollen development

Abstract

We have isolated two genes from Zea mays encoding proteins of 82 and 81 kD that are highly homologous to the Drosophila 83-kD heat shock protein gene and have analyzed the structure and pattern of expression of these two genes during heat shock and development. Southern blot analysis and hybrid select translations indicate that the highly homologous hsp82 and hsp81 genes are members of a small multigene family composed of at least two and perhaps three or more gene family members. The deduced amino acid sequence of these proteins based on the nucleotide sequence of the coding regions shows 64-88% amino acid homology to other hsp90 family genes from human, yeast, Drosophila, and Arabidopsis. The promoter regions of both the hsp82 and hsp81 genes contain several heat shock elements (HSEs), which are putative binding sites for heat shock transcription factor (HSF) commonly found in the promoters of other heat shock genes. Gene-specific oligonucleotide probes were synthesized and used to examine the mRNA expression patterns of the hsp81 and hsp82 genes during heat shock, embryogenesis, and pollen development. The hsp81 gene is only mildly heat inducible in leaf tissue, but is strongly expressed in the absence of heat shock during the pre-meiotic and meiotic prophase stages of pollen development and in embryos, as well as in heat-shocked embryos and tassels. The hsp82 gene shows strong heat inducibility at heat-shock temperatures (37-42 degrees C) and in heat shocked embryos and tassels but is only weakly expressed in the absence of heat shock. Promoter-GUS reporter gene fusions made and analyzed by transient expression assays in Black Mexican Sweet (BMS) Maize protoplasts also indicate that the hsp82 and hsp81 are regulated differentially. The hsp82 promoter confers strong heat-inducible expression of the GUS reporter gene in heat-treated cells (60- to 80-fold over control levels), whereas the hsp81 promoter is only weakly heat inducible (5- to 10-fold over control levels).

The chlorate‐resistant and photomorphogenesis‐defective mutant cr88 encodes a chloroplast‐targeted HSP90

“…Recently, Queitsch et al (2002) showed that HSP90 may act as a capacitor, as had been proposed previously in Drosophila (Rutherfort and Lindquist, 1998). Genes encoding HSP90, along with their pattern of expression at the mRNA level, have been reported in maize (Marrs et al, 1993), barley (Walther-Larsen et al, 1993), tomato (Koning et al, 1992), periwinkle (Schroder et al, 1993), Brassica (Krishna et al, 1995;Neumann et al, 1993;Reddy et al, 1998), Arabidopsis (Conner et al, 1990;Takahashi et al, 1992), Pharbitis (Felsheim and Das, 1992), and rye (Schmitz et al, 1996), but the functional signi®cance of their expression patterns is not clear. In Arabidopsis, there are at least six HSP90 genes in addition to CR88.…”

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

“…Ia), a gene-specific oligonucleotide probe for the small heat-shock protein hsp17.9 (Marrs et al, 1993), the PCRderived fragment within the first exon of the maize GST I11 gene (GST) prepared for this study using the sequence reported by Moore et al (1986), and the cDNA plasmids for the maize C2 gene pC2-c46 (Wienand et al, 1986), the Adhl gene pB428 (Bennetzen et al, 1984), the B z l gene pMBzRl (Klein et al, 1989), and the vbcS gene pTN20 (a gift from Tim Nelson, Yale University, New Haven, CT). Probes were 32P-labeled using the oligolabeling kit supplied by Pharmacia.…”

“…As a control, RT-PCR was also performed using primers spanning the intron of the maize hsp82 gene to test the expression of an endogenous cadmium-inducible gene. Primers hsp82f and hsp82r span the single 140-bp intron of maize hsp82 (Marrs et al, 1993) and amplify a 380-bp fragment from the spliced transcript and a 520-bp fragment from the unspliced transcript. The sequences of these primers are: hsp82f (forward primer), S'-GTGCACATGGCTGGCGGTGC, and hsp82r (reverse primer), 5'-GGACTTGGTCATGCCCACGCC.…”

“…1 a) amplified a 220-bp fragment from the unspliced message and a 142-bp fragment spliced message (left); primers Actl-5 and Act 400 (Fig. 1 b) could amplify a 400-bp fragment from the unspliced message and a 316-bp fragment spliced message (middle); and, as a cadmium-inducible control (C), primers hsp821 and hsp82r could amplify transcripts from the endogenous expression of the maize hsp82 gene (Marrs et al, 1993). A 520-bp fragment was amplified from the unspliced Hsp82 message and a 380-bp fragment spliced hsp82 message (right).…”

Combinatorial Interaction of Cis Elements Specifies the Expression of the Arabidopsis AtHsp90-1Gene

“…The proteins are localized in different cell compartments, including the cytoplasm, the endoplasmic reticulum, and chloroplasts (Koning et al, 1992;Takahashi et al, 1992;Marrs et al, 1993;Schrö der et al, 1993;Krishna et al, 1995;Schmitz et al, 1996;Milioni and Hatzopoulos, 1997). The corresponding genes were shown to be specifically expressed during embryogenesis, pollen development (Marrs et al, 1993), and seed germination (Reddy et al, 1998) in young and rapidly divid-ing tissues such as shoot and root apices (Koning et al, 1992) and in flowers (Takahashi et al, 1992;Krishna et al, 1995). In oilseed rape (Brassica napus) and tomato (Lycopersicon esculentum) seedlings, HSP90 protein levels were found to increase by exogenous 24-epibrassinolide application (Dhaubhadel et al, 1999), whereas a glucosinolate-deficient Arabidopsis mutant was shown to be thermosensitive and defective in the cytosolic HSP90 expression after heat stress (Ludwig-Muller et al, 2000).…”