Discovery of an expanded set of avian leukosis subgroup E proviruses in chickens using Vermillion, a novel sequence capture and analysis pipeline [corrected].

Transposable elements (TEs), such as endogenous retroviruses (ERVs), are common in the genomes of vertebrates. ERVs result from retroviral infections of germ-line cells, and once integrated into host DNA they become part of the host's heritable genetic material. ERVs have been ascribed positive effects on host physiology such as the generation of novel, adaptive genetic variation and resistance to infection, as well as negative effects as agents of tumorigenesis and disease. The avian leukosis virus subgroup E family (ALVE) of endogenous viruses of chickens has been used as a model system for studying the effects of ERVs on host physiology, and approximately 30 distinct ALVE proviruses have been described in the Gallus gallus genome. In this report we describe the development of a software tool, which we call Vermillion, and the use of this tool in combination with targeted next-generation sequencing (NGS) to increase the number of known proviruses belonging to the ALVE family of ERVs in the chicken genome by 4-fold, including expanding the number of known ALVE elements on chromosome 1 (Gga1) from the current 9 to a total of 40. Although we focused on the discovery of ALVE elements in chickens, with appropriate selection of target sequences Vermillion can be used to develop profiles of other families of ERVs and TEs in chickens as well as in species other than the chicken.

Cloning and expression of a chicken alpha-amylase gene.

We have isolated and sequenced a genomic clone for a pancreatic alpha-amylase gene (amy) of the chicken (Gallus gallus). The gene is interrupted by nine introns, spans over 4 kb, and encodes a protein (AMY) of 512 aa that is 83% identical to the human pancreatic alpha-amylase enzyme. Southern blot analysis of chicken DNA revealed two distinct pancreatic amy loci. In addition, we have generated a cDNA from chicken pancreatic RNA corresponding to the coding sequence of the genomic clone. The cDNA was inserted into a yeast expression vector, and the resulting construct used to transform Saccharomyces cerevisiae cells. Transformed yeast cells synthesized and secreted active AMY enzyme, and the gel migration pattern of the alpha-amylase produced by the yeast cells was identical to that of the native chicken enzyme.

A Drosophila gene promoter is subject to glucose repression in yeast cells.

Previous work has shown that the alpha-amylase gene of Drosophila melanogaster is subject to repression by dietary glucose. Moreover, glucose repression of this gene is mediated by promoter elements that lie upstream of the transcriptional start site. In this study, we examined the activity of the glucose-repressible Drosophila promoter in transformed yeast cells. We show that the amylase promoter region can mediate glucose repression of a heterologous reporter gene in yeast. The implication of this result is that the yeast regulatory machinery can recognize the Drosophila promoter signals. This, in turn, implies an unexpectedly high degree of evolutionary conservation in the mechanism of glucose repression among eukaryotes. It also shows that genes that have acquired complex patterns of developmental regulation-e.g., the Drosophila amylase gene, can still retain, intact, more primitive forms of regulation, such as glucose repression.