Characterization of the BhC4-1-lacZ mRNA pattern of expression in embryos of the (-824/-187), (-488/-187) and (-850/+40) series in D. melanogaster transgenic lines

• Grant number: 16/00412-5
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: July 01, 2016
• End date: June 30, 2018
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Nadia Monesi
• Grantee: João Vitor Cardoso Uliana
• Host Institution: Faculdade de Ciências Farmacêuticas de Ribeirão Preto (FCFRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

The DNA puff C4 BhC4-1 gene of Bradysia hygiene is amplified and expressed in a regulated manner in two larval tissues, the prothoracic gland, and the salivary gland. Functional studies in Drosophila melanogaster revealed that the molecular mechanisms that regulate BhC4-1 expression are conserved in D. melanogaster and resulted in the identification of cis-regulatory modules that regulate the expression of the sciarid gene. Among the identified modules, the (-253/-187) module regulates BhC4-1-lacZ expression in the ring gland of late embryos, while the distal cis-repressor module (-3314/-1294) represses BhC4-1-lacZ ring gland expression at developmental times prior to 0h prepupae and the proximal cis-repressor module (-1293/-254) represses BhC4-1-lacZ ring gland expression at developmental stages prior to the second larval instar. The lines (-824/-187) and (-488/-187) were previously obtained in our laboratory and were designed to extend the functional characterization of the proximal repressor module. In these lines, the ring gland cis-regulatory module (-253/-187) is placed downstream of either the fragment (-824/-254) or (-488/-254) and reporter gene expression occurs both in the ring gland of second instar larvae and ectopically in embryonic epidermal cells. The levels of ectopic expression of the reporter protein in the epidermis of embryos appear to be higher in lines (-824/-187) and (-488/-187), when compared to those observed in lines (-850/+40), which may be indicating the existence of a module that represses the ectopic expression of the transgene in the (-186/+40) fragment. These lines were only analyzed for the pattern of expression of the reporter protein. This project intends to refine the characterization of the pattern of expression of the transgene in lines (-824/-187), (-488/-187), and (-850/+40) employing in situ hybridization experiments. With these experiments it will be possible to compare the expression levels of the transgene mRNA in the epidermal cells of the embryo of lines (-824/-187) with those observed in lines (-850/+40), which will confirm the presence of cis-regulatory modules in the (-186/+40) fragment that repress the ectopic pattern of expression. Together, the proposed experiments are an important step towards refining the characterization of the BhC4-1 promoter, will constitute a starting point for the design of future experiments, and will contribute with relevant information about eukaryote cis-regulatory modules. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

CARDOSO ULIANA, JOAO VITOR; PEREIRA BRANCINI, GUILHERME THOMAZ; CASTELLI-GAIR HOMBRIA, JAMES; DIGIAMPIETRI, LUCIANO ANTONIO; ANDRIOLI, LUIZ PAULO; MONESI, NADIA. Characterizing the embryonic development of B. hygida (Diptera: Sciaridae) following enzymatic treatment to permeabilize the serosal cuticle. MECHANISMS OF DEVELOPMENT, v. 154, p. 270-276, DEC 2018. (16/25325-8, 16/00412-5, 15/24305-0, 14/14318-5)