Structural characterization of the medfly hsp83 gene and functional analysis of its proximal promoter region in vivo by germ‐line transformation

Abstract

In order to define the regulatory elements responsible for the expression of the medfly hsp83 (Cchsp83) gene, we determined the sequence of a genomic region of the gene that included 3,536 bp upstream of the transcription initiation site, the first untranslated exon of 144 bp, a 275-bp intron, and 516 bp of the second coding exon. Structural analysis of the 5' flanking region revealed the presence of a typical TATA box, 28 bp upstream of the transcription start site, and seven putative heat shock elements (HSEs) further upstream. The 5' untranslated region of the Cchsp83 mRNA was found to contain extensive secondary structure in the first 126 nucleotides. We carried out deletion functional analysis of the proximal promoter region (-380/+139) in vivo by germ line transformation using the lacZ as a reporter gene. We found that sequences in the -380/-86 region are essential for the constitutive expression of the Cchsp83 gene. Under normal conditions, the -380/+139 region was able to drive significant levels of transgene expression in all developmental stages of the medfly as well as in the ovaries and testis. In most stages, the temporal expression pattern of the reporter gene was similar to the respective pattern of the endogenous Cchsp83 gene. Although the -380/+139 promoter region contained two putative HSEs, it was found unable to confer any heat-induced expression in the reporter gene.

Cloning & sequence identification of Hsp27 gene and expression analysis of the protein on thermal stress in Lucilia cuprina

“…Interestingly, in later stages of embryogenesis, the level of Hsp27 protein expression increases considerably (Arrigo & Pauli, 1988;Arrigo & Tanguay, 1991). A similar pattern of Hsp27 expression was also found in different embryonic stages of C. capitata at high temperature Theodoraki et al, 2008). The pattern is found to be similar in vertebrates too.…”

The hsp27 gene of the Mediterranean fruit fly, Ceratitis capitata: structural characterization, regulation and developmental expression

“…In addition to Drosophila , detailed studies on the sHsps in other insect species are of interest in order to understand the role and regulation of these multifunctional proteins and reveal potential important similarities and/or differences between them. We are interested in studying the regulation of heat shock genes in the Mediterranean fruit fly Ceratitis capitata (medfly) (Komitopoulou et al ., 2004; Kalosaka et al ., 2006; Theodoraki et al ., 2008). Medfly is a notorious pest with a worldwide range and a history of fast expansion and disastrous invasions to various countries (Harris, 1989), and so far is the best‐studied fruit fly pest at the genetic and molecular level.…”

Identification of a Novel Strong and Ubiquitous Promoter/Enhancer in the SilkwormBombyx mori

“…We therefore attempted to identify the genomic region responsible for the strong expression of hsp90 . In mammals and flies, hsp90 expression is regulated by a promoter/enhancer located in an upstream and/or intronic region ( Xiao and Lis 1989 ; Shen et al 1997 ; Zhang et al 1999 ; Theodoraki et al 2008 ). Because the silkworm hsp90 lacks intron ( Figure 4 , Landais et al 2001 ), we focused on its 5′ flanking region.…”