Wen 2020

Wen Y. 2020. Characterization of MITF gene in crayfish and their possible role in innate immunity. Master’s thesis, Department of Animal Breeding and Genetics, Sveriges lantbruksuniversitet (Swedish University of Agricultural Sciences). http://urn.kb.se/resolve?urn=urn:nbn:se:slu:epsilon-s-16418

Abstract

Microphthalmia-associated transcription factor (MITF) is a tissue-specific transcription factor (TF), with a basic helix-loop-helix leucine zipper (bHLH-LZ) domain, which binds to the canonical E-box sequence (5’-CANNTG-3’) in the promoter region of target genes. In this study, the MITF-like protein was identified in the marbled crayfish (Procambarus virginalis). The full-length cDNA coding sequence of the most similar gene to MITF from marbled crayfish is 1284bp (427 amino acids). In the secondary and tertiary structure of the deduced amino acid sequence, a conserved functional structure of bHLH-LZ was shown, which could bind to E-box. In the phylogenetic analysis, this obtained MITF-like gene showed a lager evolutionary distance to all the vertebrates and some invertebrates like lancelet (Branchiostoma belcheri), starfish (Acanthaster planci), sea urchin (Paracentrotus lividus) and (Lytechinus variegatus), but is closer than worms (Caenorhabditis elegans). The bHLH-LZ domain is located in exon 6, exon 7, exon 8, and exon 9 in the MITF gene and has a similar structure as the corresponding exons in human. Besides, in this study, a comparison of exon-intron structure of MITF between human, mouse, fruit fly and marbled crayfish was performed, it was shown that the splicing sites of the bHLH-LZ domain in MITF gene might be conserved across species. To evaluate the possible role of MITF as a TF in innate immune system regulation, a similar prophenoloxidase (proPO) DNA sequence was analysed for the presence of E-box. The proPO gene is responsible for trapping and myelinization of pathogens in invertebrate. The proPO-like gene of marbled crayfish contains eight CANNTG sequences. In addition, anti-apoptotic factor (BCL-2)-like gene was found in marbled crayfish and four CANNTG sequences were found. Our results provided evidence of the presence of MITF-like gene in crayfish species and may provide knowledge on the role of MITF in innate immune activation.

Keywords: Microphthalmia-associated transcription factor • basic Helix-Loop-Helix Leucine Zipper • crayfish • Aphanomyces astaci • immunity