Assoc Prof Russell Poulter

Biochemistry Department University of Otago P.O. Box 56 710 Cumberland St Dunedin 9054 , New Zealand Tel.: +64 3 479-7856 FAX: +64 3 479-7866 e-mail: russell@sanger.otago.ac.nz

Eukaryote mobile genetic elements

A long-term interest of this laboratory has been in the genetics of imperfect fungi (i.e. fungi without a known sexual cycle). More recently, our focus shifted to the retrotransposons present in fungi and, by extension, to retroelements in general. The five major groups of LTR retrotransposons are known as the Ty3/gypsy, Ty1/copia, DIRS1, BEL, and vertebrate retrovirus groups. The elements within each group can generally be distinguished from elements of other groups by comparisons of the sequences of their shared enzymatic domains. We have isolated a Ty1/copia retrotransposon, TCa2, from the fungal pathogen Candida albicans. TCa2 has a number of interesting features including an in-phase suppressible stop codon between ORF1 and ORF2. We have also analysed a Ty3/Gypsy element from C. albicans, Tca3. Tca3 was first identified as a widespread form that lacks a large part of its coding region; however, comparative analyses between C. albicans and C. dubliniensis allowed us to identify the closely related full-length Tcd3 element. Subsequently, we uncovered the rare full-length Tca3 elements. The potential uses of retroelements in biotechnology and their application to the analysis of fungal pathogenicity are at present being investigated. We have also contributed to the analysis of mobile elements such as DNA transposons (including tyrosine recombinase-encoding elements and Helitrons) and retroelements in other fungi, especially the basidiomycete pathogen, Cryptococcus neoformans.

During an analysis of the mobile genetic elements of Cryptococcus we detected an intein in the Cryptococcus genome. Inteins are encoded transposable elements that occur naturally as in-frame, translated insertions in the coding sequences of organisms from all three biological kingdoms. The coding sequences (encoding the exteins) of certain host genes are interrupted by inserted sequences (encoding inteins). The inteins (internal protein) disrupt the functioning of the protein and must be removed after translation to allow the host protein to function. We are engaged in in vitro and in vivo studies of intein function in collaboration with Dr Sigurd Wilbanks of this department.

We have extended our interest to the retrotransposons of vertebrates and have characterised a full-length multi-copy (x1000) LTR retrotransposon from the Fugu fish, Takifugu rubripes. This work was in collaboration with the HGMP/MRC Cambridge. The retrotransposon, sushi, is the first full length LTR retrotransposon from any vertebrate. Sushi is a member of the ty3/Gypsy group. It has many features that suggest it could represent the ancestral group from which vertebrate retroviruses were derived. Sushi, however, has closest homology to a group of fungal retrotransposons. This presents the interesting possibility that retroviruses are derived from retrotransposons that were horizontally transmitted to vertebrates from fungi. Dr Poulter and Dr John Cutfield (also from the Dept. of Biochemistry) were awarded a Marsden grant, funded by the N.Z. Royal Society, to investigate this possibility.

During this project we discovered vertebrate representatives from other retrotransposon groups. We described an element (Gmr1) from the Atlantic cod (Gadus morhua) in which the pol domains appear in the same order as in Ty1/copia elements, PRO-INT-RT-RNH, yet sequence comparisons clearly show that the element is a member of the Ty3/gypsy group. Perhaps the most distinctive LTR retrotransposons are the members of the DIRS1 group. These elements have quite different structures from all other LTR retrotransposons, encode a different set of proteins, and probably have distinct replication mechanisms. They contain genes for a putative Gag protein, RT and RNH, and a tyrosine recombinase. We discovered DIRS1-like elements in the genome of Tetraodon (a freshwater pufferfish). Another family of retrotransposons, the Ngaro1-like elements, also contain genes encoding putative tyrosine recombinases. Ngaro1-like elements differ from members of the DIRS1 group in that they consistently form a separate clade on phylogenetic trees based on alignments of RT, RNH and recombinase sequences, and they have distinct structures. The new elements thus appear to represent a second lineage of tyrosine recombinase-encoding retrotransposons. Ngaro1-like retrotransposons are found in a wide variety of eukaryotes, including plants, fungi, and animals, suggesting that they are an ancient class of element.

Dr Poulter's laboratory has a wide range of other interests, ranging from transgenic yeast technology as applied to the wine industry to the application of genetic analyses to horticulture. Dr Poulter has recently described a gene conferring resistance to powdery mildew in Lathyrus, the Sweet pea.