Temporal control of imaginal disc growth

Growth of Drosophila melanogaster imaginal disc cells proceeds rapidly during larval stages but declines as the larva approaches pupariation. This growth decline is critical for defining the final size of adult appendages, yet we know little about how it is regulated. In the nematode C.elegans aspects of developmental timing are controlled by the small temporal RNA (stRNA) let-7. Mutations in let-7 prevent the onset of adult stages of development, causing reiteration of the L4 larval stage. The functional 22 nucleotide let-7 sequence is completely conserved in D. melanogaster and D. pseudoobscura. Developmental timing of let-7 expression is also conserved in D. melanogaster, turning on initially in late larval stages, peaking 42 hours after puparium formation, and declining again at the pupa to adult transition. Since the timing of D. melanogaster let-7 expression coincides with the slowing of growth of imaginal disc cells as the larva approaches pupariation, we are investigating whether let-7 plays a role in regulating this process.
Small temporal RNAs are a class of microRNAs. MicroRNAs regulate gene expression by binding to semi-complementary regions within the 3'UTR of target genes and inhibiting translation of the mRNA. We have identified several cell cycle and growth regulatory genes whose products or activity decline during the latter part of larval development. Using a bioinformatics approach, we have identified possible let-7 target sites within the 3'UTRs of these candidate genes. To empirically confirm the ability of D. melanogaster let-7 to bind to and regulate the candidate genes we have constructed regulatory sensors containing EGFP followed by the 3'UTRs of the candidate genes. We are using these sensors in combination with overexpression constructs to examine the role of let-7 and other microRNAs in temporal control of imaginal discs growth.