SIMILARITIES TO A LINE ELEMENT SHARED BY ANOPHELINE AND CULICINE MOSQUITOS MAP TO THE DISTAL END OF DIHYDROFOLATE-REDUCTASE AMPLICONS IN AEDES-ALBOPICTUS MOSQUITO CELLS

To extend our understanding of amplicon structure in methotrexate-resi stant Mtx-5011-256 Aedes albopictus mosquito cells, we examined a seri es of cosmids containing genomic DNA corresponding to the unique 3'-en d of the Type I dihydrofolate reductase amplicon. Cosmid pWED118 conta ined five EcoRI fragments ranging from 2 to 5 kb (A, B, C, F, G) that hybridized to cDNA from methotrexate-resistant cells. Of these, fragme nts B and F hybridized weakly to first-strand cDNA from sensitive cell s and shared considerable sequence identity. Fragment G occurred twice in the map of pWED118; one copy mapped within a 10 kb BssHII core fra gment from the Type I amplicon and a second copy mapped downstream in the 48 kb BssHII core fragment. Hybridization signals among fragments contained in overlapping cosmids suggested that a branch point definin g two or more subtypes of the Type I amplicon occurs within or near th e 10 kb BssHII genomic DNA fragment. A 1.8 kb sequence common to fragm ents B and F included an similar to 0.4 kb region that shared sequence similarities with a LINE element from Aedes aegypti and with a repeat ed sequence from Anopheles gambiae. In addition, these elements shared amino acid similarity to a reverse transcriptase from the nematode, C aenorhabditis elegans. Shared sequence between Aedes and Anopheles ele ments supports the hypothesis that an ancestral LINE-like element was active in mosquito genomes prior to the divergence of the subfamilies Culicinae and Anophelinae. The presence of homologies to LINE-like ele ments near a branch point in the dihydrofolate reductase amplicon is c onsistent with a possible role of repeated sequences in amplicon short ening. (C) 1998 Elsevier Science Ltd. All rights reserved.